Understand the Role of the Nurse

Introduction The purpose of this assignment is to explore the professional responsibilities of the nurse and their role in safeguarding vulnerable patients, all of which are based around a fictitious scenario. The scenario is centred on a patient named Margaret and is going to be used to identify any vulnerability issues Margaret may have. Margaret could be identified as been vulnerable, as she is an 89 year old lady who is suffering from a heart condition. Margaret is currently living with her daughter and has said that her mother’s health has begun to deteriorate.Margaret is becoming very confused and forgetting to take her medication which has resulted in numerous admissions to hospital. No consent was needed on this occasion as it is a fictitious patient. Based around this fictitious scenario, the aim is to discuss and identify vulnerability and any issues arising from it. Exploring all aspects of vulnerability involving all models and discussing confidentiality, looking a t the importance of it and why it should be maintained.This assignment will also look at the mental capacity act and how the nurse and other healthcare professionals assess the patient for capacity and use the models of assessment. All these are in place to protect the patient’s health and well-being and patient rights. Communication is another important skill for any nurse to have. Good communication skills are essential in providing a high standard of care, which will be explored and discussed in detail.It will also explore what accountability and responsibility is within the nurse’s role. The nurse being accountable for their actions and adhere to the Nursing and Midwifery Councils (NMC), Code. â€Å"The core function of the Nursing and Midwifery Council is to establish standards of education, training, conduct and performance for nurses and midwives to ensure those standards are maintained, thereby safeguarding the health and well-being of the public† (NMC, code, 2009).It will then identify and explain how the patient’s autonomy might be promoted by the nurse, highlighting the support available and with the nurse building a therapeutic relationship with the patient, relating to scenario whilst maintaining appropriate professional behaviour and interventions. Overall this assignment will explore and discuss many of the NMC guidelines about the above topics.It will give you an understanding of the nurse’s role involved in delivering the best possible care to the public. Firstly to explore accountability within the nurse’s role The Nursing and Midwifery Council states, â€Å"Accountability is often defined as responsibility, but there is a difference between the two. Responsibility is concerned with answering for what you do, whereas accountability is being answerable for the ‘consequences’ of what you do† (NMC, 2009).

Economic Problems in Brazil

Aphrodite was the goddess of love, beauty and sexuality, she was often naked The Romans called her Venus, because of her beauty. Aphrodite was born of the foam sea when cronus cut of Uranus gentiles and threw them into the sea, Aphrodite lived on Mount Olympus with the other supreme deities. In which she married a homely craft man god named Hephaestus, her parents where Uranus or Zeus and Dione, she hardly where clothes and she cause the Trojan War.Aphrodite thinks noting but love and her work is pleasure, often Aphrodite would have in affairs with other guys. When the hero Peleus was married to the sea-nymph Thetis, all the gods were invited to the ceremony, but one god. So she got a golden apple and rolled it to the three gods who were sitting down and on that golden apple it had a note saying â€Å" For the fairest† the three gods began to fight over the apple so they go up to Zeus to tell him pick which of them is the most beautiful one.Zeus had a hard time choosing who, s ince all three of them were dear to him so he let the handsome youth god the Trojan Prince Paris choose. The goddess where bribing him except for Aphrodite was trying to seduced him, telling him â€Å"I will give you the most beautiful god†, so he think about it and he go’s with Aphrodite’s bet.

A Case Study in Applying Theories to Practice

A Case Study in Applying Theories to Practice Introduction The aim of this assignment is to show how a Social Worker would apply a Behavioural Method as an understanding and intervention on the case study supplied. I will do this by explaining what Behavioural Social Work is, how it is used in practice, how it meets the needs of the service user and identify personal challenges along the way. I will try to challenge my own views and ideals on the service user, and how these can be challenged for Anti-Oppressive Practice. The History of Behavioural Social Work â€Å"Most of what makes us truly human, most of what makes us individuals rather than ‘clones’, most of what gives us a discernible personality – made up of characteristic patterns of behaviour, emotion and cognition – is the result of learning† (Sheldon and McDonald, 2008). The Behavioural Perspective focuses on the individual and the relationship between stimuli in an environment and how it determines behaviour through learning (Westen, 2001). This is also how Behavioural Social Work is executed, by focusing on behaviour that is observable and changeable. Like other forms of social work methods it has been adopted from other disciplines, and in particular Psychology, but has been adapted to achieve measured outcomes and effective practice for Social Work (Watson and West, 2006). There are four main Theories that are relevant and used in Behavioural Social Work; The first behavioural theory is Respondent Conditioning , first introduced by Ivan Pavlov, who used experiments on dogs to discover how to condition a response after the presence of a certain stimuli has been removed. The second major behavioural theorist is B.F Skinner and his Operant Conditioning , he observed that the behaviour of organisms can be controlled by environmental consequences that either increase (reinforce) or decrease (punish) the likelihood of the behaviour occurring (Westen, 2001). He claimed that the outcome of behaviour was volunt ary and goal directed, and always controlled by the consequences the behaviour would lead to. The next behavioural theory identified is Social Learning Theory , which extends behavioural ideas and claims that most learning is gained by copying others around them, rather than them being reinforced as skinner claimed. That behaviour is shaped by observing others and interpreting it (Payne, 1997). The fourth behavioural theory is Cognitive Learning Theory and was introduced by Albert Ellis and Aaron Beck in the 1960’s, this is a theory that focuses on the way people perceive, process and retrieve information (Westen, 2001). A perception of the environment from previous experience. It was during the 1980’s that Social Work adopted behavioural theory as a method of working with service users, part of the reason for this was the ability of the theory to achieve realistic outcomes (Watson and West, 2006). The learning theory used in social work is a combination betwe en all four theories listed above; Respondent Conditioning, Operant Conditioning, The Social Learning Theory and Cognitive Learning. These are used to enable the social worker to observe behaviour and therefore intervene appropriately (Watson and West, 2006). The importance of behavioural social work is that the behaviour is learned and can therefore be unlearned. Cognitive learning theory focuses on this specifically and its engagement with cognitive processes which produce thoughts and feelings (Sheldon, 1995). Behavioural social work allows the service user to modify and change their behaviour through a process of reinforcement, both positive and negative, to produce a likeliness of a wanted behaviour occurring (Watson and West, 2006).

Why Are Some Fruits Sweet and Some Sour and how does the Ripening of Essay

Why Are Some Fruits Sweet and Some Sour and how does the Ripening of Fruit Affect It - Essay Example Other sour tasting fruits include raw mangoes. According to Hulme (1970) fruits consist of different substances, including vitamins, sugars, starch, proteins, and organic acids in different proportions. The taste of a particular fruit depends on the concentrations of these substances. In sweet tasting fruits, the concentration of fructose, which is a simple monosaccharide, is higher than the concentration of sour substances such as organic acids. Fructose is a simple sugar and is highly soluble in water and hence it is a major component of fruit juices. Naturally, bananas have high concentration of starch and as they ripen, it is converted into simple sugars such as fructose (Childers, 1978). This explains why ripe bananas have sweet taste while green bananas are not. In fruits, fructose could be in its simplest form or as a component of sucrose. Sour taste in fruits is caused by high concentration of hydrogen ions present in organic acids (Hulme, 1970). Some common organic acids in sour fruits include citric, malic and tartaric acids. Ripening of fruits affects the process of seed dispersal in various ways. According to Mauseth, (2003) the major functions of the fruit are to protect the developing seed and enhance its dispersal. As the seed matures, the fruit ripens simultaneously. Fruits undergo different ripening processes that determine the eventual method of dispersal. In wind-dispersed seed, the fruit undergoes dehydration or desiccation, which reduces the weight of the seed. In other wind-dispersed seeds, the dehydration process enhances development of flight structures such as wings, which increases the floating ability of the seed in air (Mauseth, 2003). Therefore, the desiccation process during the ripening of wind-dispersed seeds reduces the weight of the seed, which makes it possible for wind to carry and disperse the seed away from the parent plant. In animal dispersed seeds, the ripening process increases the attraction of the fruit to the consumi ng animal (Biale, 1954). During ripening of such fruits, the plant releases enzymes within the plant that enhances conversion of complex compounds such as starch into sweet tasting sugars. The enzymes enhance production of sweet smelling aromatic compounds to attract animals during the ripening process. Some fruits such as tomatoes release ethylene that induces ripening, characterized by production of pink, reddish coloration and softening of the fruit (Mauseth, 2003). The appealing color, sweet taste and aromatic smell attract animals, which eat the fruit and disperse the seeds in the process. Therefore ripening of fruit enhances seed dispersal by promoting development of characteristics appropriate to the particular method of dispersal. Human beings interfere with the method of seed dispersal by consuming grains and fruits. Consumption of immature seeds and fruits interferes with natural process of seed growth and development (Hulme, 1970). Therefore, when people eat the immature fruits and seeds, the natural process of seed dispersal and the eventual germination is interfered with because the seeds lose their viability to germinate. Besides consumption, human beings interfere with the process of seed dispersal by subjecting fruits and seeds to various techniques such as food processing and storage. Most food processing methods such as drying removes water in the seeds and the fruits. This interferes with critical seed development processes that depend on water (Hulme, 1970). For example, drying seeds and fruits interferes with enzymatic and hormonal processes and reactions that promote ripening, development and eventual seed dispersal. Storage of seeds and fruits prevent other agents of dispersal such as wind and other animals from carrying

The Job Satisfaction in Hasbro Company Case Study - 20

The Job Satisfaction in Hasbro Company - Case Study Example The job satisfaction in the company will depend on how the mission statements of the company will help the employers in achieving their profit levels and also their personal enhancement. At Hasbro, there are two sets of people working on designing the games. One who focuses on the traditional methods and are happy with their routine jobs. The other set is the innovators who develop new strategies to the existing games in order to make it more appealing. Thus, the satisfaction level depends on the type of job the person is engaged in. People like Peaches Belanger are the company loyalists who are happy with their existing pattern of routine jobs. They take pride in owning a job and develop a sense of belonging to their company. The company can increase their pay scales, provide them with additional perks and benefit and acknowledge their work in the company to increase their job satisfaction level. Rob Daviau’s job satisfaction level is more than that of Peaches Belanger though the latter’s is more dedicated towards the company. This can be explained using Locke’s Value Discrepancy Theory. This theory explains that satisfaction in a job results from the satisfaction of the desires than from the deprived needs (Singh, p.259). Daviau’s sought to introduce changes in the existing standard of the games that were manufactured by the company. He was satisfied with the fact that there were very few game planners like him in the entire country. This personal feeling helps in boosting the job satisfaction in a person and his overall performance.

THE MEANING AND LEGACY OF THE KENNEDY ELECTION Assignment

THE MEANING AND LEGACY OF THE KENNEDY ELECTION - Assignment Example icals of the previous era is in their peaceful way of gaining the power, not revolutionary one, for their major doctrine is rights and freedoms of man as key value and legal platform of social order. One of the major events in the late 1950s that awakened the nation is the struggle for the rights of blacks headed by prophet Martin Luther King, which has gradually begun to gain popularity in the society of whites.   1960 appears to be the year of John F. Kennedy becoming the President of America due to several reasons, namely: support of the great number of registered Democrats (for economic recession of Republicans), as well as one of Catholics; focusing of his efforts on densely populated States of the North-East; benefit of well-financed campaign, and television translation. Kennedy’s campaign platform has contained the program of socio-economic reforms, forward to new frontier, including reduction of taxation, adoption of a series of legislative measures to ensure the civil rights of colored people, health insurance for the elderly, raising the minimum wage, monopoly control, and acceleration of the pace of economic development. The major events determined the elections are participation of voters from two new states of America, that is, Hawaii and Alaska, as well as public advocacy of electoral campaign by both presidential candidates. â€Å"An unprecedented series of four television  debates  between the two nominees constituted the highlight of the campaign† (Levy).   As for the major political and economic policies of Eisenhower administration, they are characterized by the concept of creating of favorable state framework for individual economic activities, the end of the practice of persecution for leftist persuasion, and putting a stake on achieving nuclear superiority over the USSR (the Eisenhower’s administration General line of foreign policy of the

The Decameron Essay Example | Topics and Well Written Essays - 1250 words

The Decameron - Essay Example The book shows that even the most noble or holy people in title are still prone to sin and vice. Living the best life possible, according to The Decameron, is a personal endeavor which can best be undertaken by fortifying one's mind with tales of the moral shortfalls of others. The story begins with a narrator discussing the various ways in which the people of Florence were coping with the overwhelming death toll and destruction of social order brought about by the bubonic plague. Some people chose to behave "as though each day was to be their last" (16). The narrator notes that "nowadays, laws relating to pleasure are somewhat restrictive, whereas at that timethey were exceptionally lax." In contrast, however, the seven ladies of our story are found "more or less in a circle, in one part of the church, reciting their paternosters" (17). The eldest of the women, Pampinea, determines it is in their best interests to leave town to protect their lives and virtue. She states that the townspeople, "prompted by their appetites, they will do whatever affords them the greatest pleasureIt is not only of lay people I speak, but also of those enclosed in monasteries[that have] given themselves over to carnal pleasures" (18-19) She asks the other six women, "If this b e so (and we plainly perceive that it is), what are we doing here" (19). In this manner, the seven women decided to leave town, asking "three young men of courage and intelligence" to accompany them as "guides and servants" (21). In this manner, the group sets itself apart from the others in Florence that are just interested in fulfilling sinful desires before their imminent demise. Upon arriving at a well-appointed and vacant country villa, the group and their accompanying servants set up camp. To provide entertainment, they decide they will appoint a group leader each day that will determine how they will pass their time (24). For each of the ten days they are in the villa, they end up telling stories - both lighthearted and tragic - that convey different moral shortcomings of men, women and clergy alike. They each usually preface each tale with a comment on how to avoid the same wretched and sinful natures of the people in the stories, thus making the collection of tales a type of instruction manual on the devious temptations one should avoid in life. For instance, the third story on the first day tells the tale of Saladen, a sultan from Egypt, and "a rich Jew, Melchizedek by name." Saladen asks Melchizedek to choose which religion is authentic, the "Jewish, the Saracen, or the Christian" (42). Because Melchizedek wisely sidesteps Saladen's wicked trap, he avoids giving an answer that would lose him money and possibly his life. He states that he could not choose among them, just like he could not choose among three children. Saladen only asked this question because he coveted Melchizedek's money. Because his wisdom and prudence inspired Saladen, the sultan borrowed money from the man instead of entrapping him as he had originally intended ( 43-44). Many stories in The Decameron point out the hypocrisy of the Catholic Church of plague-era Europe. In this manner, the stories show that even church leaders are not without sinful temptation. In the sixth story, a money-hungry friar comes after "people with bulging purseswhom he deemed to be lacking in faith" (50-51). At the end of the story, the money-hungry friar

Theme and Narrative Elements in the Short Story Research Paper

Theme and Narrative Elements in the Short Story - Research Paper Example This paper will discuss the literary elements that can be found in the short story entitled A Dark Brown Dog, written by Stephen Crane. This allegory is mainly centered on three characters. According to the writer, the dark brown dog, the father and the child are viewed as the main characters in the story. In most literary publications, the story mainly revolves around the characters. An overview of the story proves that the dark brown dog was mistreated by the family members (Crane, 2008). On the contrary, the dog did not respond violently to the household members even after receiving various forms of torture from the members of the household. In A Dark Brown Dog, the dog was treated inappropriately by the caregivers (Crane, 2008). The child of the household occasionally protected the dog from the harsh beatings. The child usually cried whenever his parents mistreated the dog. In one rare case, the father had struck his son’s head with a large saucepan after he had run toward s him protesting in order to protect the dog. From that point onwards, his parents reconsidered the issue of throwing items to the dog. Finally, the father of the household came home drunk. Afterwards, his son arrived home together with the dog. The father gave the dog a heavy beating and threw the dog outside the house through the window (Crane, 2008). The dog landed in a heap on top of the roof of a shed which was five stories below. The mood at the end of the story assumes that the dog had died after all the exploitation by the family members. In this literary work of fiction, the most prevalent theme is violence. According to the author, the dark brown dog faces violent actions from the members of the household (Collins, 2011). However, the dog was able to cope with the unjust beatings from the family. Further analysis also shows that on some occasions, the child who occasionally protected the dog imposed some form of torture to the dog. The author asserts that earlier on, the c hild thrashed the dog with a stick after he noticed that the dog was pursuing him. After he did this, the dog usually made funny gestures of apology to the child. According to the details contained in the story, the dog usually accepted the beatings from the family members (Collins, 2011). The child occasionally cried after the dog was smashed because the child loved the dog and even treated it like a close friend. The theme of violence in this short allegory has significant effects on the literary works. The child played the role of a close friend and a guardian to the dog (Magill, 1997). There are numerous literal elements that are contained in most short stories. Some of most common elements include the setting, characters, point of view, symbolism and plot. In this short story, the plot and the characters play an eminent role in the advancement of the subject matter of the story (Collins, 2011). As stated earlier, the father, child and the dog are considered the main characters. These characters employ all forms of violence to the helpless dog. In some instances, the dog was usually protected by the child. In addition, these three characters ensured that the plot was alive all through the narration. In conclusion, the plot enables the development of the theme of violence. The author of the story categorically places his ideas in the narration. All these ideas and thoughts are liable for sustaining the main

Ethics, Moral dilemmas, and the Law in Philadelphia, Pennsylvania 2013 Assignment

Ethics, Moral dilemmas, and the Law in Philadelphia, Pennsylvania 2013 - Assignment Example They will always be against the moral ethics of individuals. For instance, a patient suffering from a fatal disease may make a decision to take away his life or fail to disclose his illness to the family with an assumption that it would cause trauma and discomfort (Garber, 2008). It should be noted that, regardless of the current condition of the patient, the nurse still has no right to concede with the patient’s wish to attempt to take his life (Werth, 2013). Siding with the patient will give a clarification that he has accepted the obligation. Instead, the nurse should advice the patient that whatever their condition, it is just temporary and their status will change after some time. Abiding to wrong decisions is considered to be morally wrong. It is considered immoral to accept decisions after knowing that the intended decisions are not right. Morally upright human beings should not be submissive to a thought that goes against human dignity especially if a responsibility is laid in ones hands (Gowans, 1987). It is against the law of the society to accept the concept of an event of what is wrong, like failing to disclose information which is vital to the life of a person. Ethics will, therefore, entail the morals used in identifying what is right and what is wrong. Concerning medical fraternity, the practitioners should have the concept of telling the truth at whatever conditions and

Discussion 6 Essay Example | Topics and Well Written Essays - 250 words - 1

Discussion 6 - Essay Example s the people in IT business and other professionals to know what new opportunities are there, challenges likely to be faced and the trends in the system. In today’s competitive global market people are looking for new opportunities to reduce cost, improve their efficiency in work and gain an edge in the market. Business people are using IT as their tool to realize their aims. The use of disruptive new technologies social networks, content-mobile apps along with proliferation of private and public cloud is making companies to assess the impact this new technologies in their businesses. Business leaders are supposed to understand the use and importance of this new advancement and use them in the improvement of their businesses (Overby, 2013). There is need of listening to the customers in this digital market in order to understand the consume ration of IT. Businesses are filled with large data, a cloud of computing information and mobile services. The argument of Overby that equilibrium of jobs offshore and onshore will be attained is quite correct. Advancement in technology makes works more efficient and creates more job opport unities and development of a

Marketing Anthropology Essay Example for Free

Marketing Anthropology Essay Anthropology and marketing (together with consumer research) were once described as ‘linchpin disciplines in parallel intellectual domains’ (Sherry 1985a: 10). To judge from the prevalent literature, however, this view is not shared by many anthropologists, who tend to look at markets (for example, Carrier 1997) and exchange rather than at marketing per se (Lien 1997 is the obvious exception here). For their part, marketers, always open to new ideas, have over the decades made – albeit eclectic (de Groot 1980:131) – use of the work of anthropologists such as Claude Levi-Strauss and Mary Douglas whose aims in promulgating their ideas on binary oppositions, totemism and grid and group were at the time far removed from the endeavour of marketing both as a discipline and as practice. Can anthropology really be of use to marketing? Can the discipline in effect market itself as an effective potential contributor to solving the problems faced by marketers? There is no reason why not. After all, it is anthropologists who point out that there is more than one market and that these markets, like the Free Market beloved by economists, are all socio-cultural constructions. In this respect, what they have to say about the social costs of markets, as well as about the non-market social institutions upon which markets depends and the social contexts that shape them (cf. Carruthers and Babb 2000:219-222), is extremely pertinent to marketers anxious to come up with definitive answers as to why certain people buy certain products and how to persuade the rest of the world to do so. At the same time, however, there are reasons why anthropology probably cannot be of direct use to marketing. In particular, as we shall see in the following discussion of marketing practices in a Japanese advertising agency, anthropology suffers from the fact that its conclusions are based on long-term immersion in a socio-cultural ‘field’ and that its methodology is frequently unscientific, subjective and imprecise. As part of their persuasive strategy, on the other hand, proponents of marketing need to present their discipline as objective, scientific, speedy and producing the necessary results. How they actually go about obtaining such results, however, and whether they really are as objective and scientific as they claim to their clients, are moot points. This paper focuses, by means of a case study, on how marketing is actually practised in a large advertising agency in Japan and has four main aims. Firstly, it outlines the organisational structure of the agency to show how marketing acts as a social mechanism to back up inter-firm ties based primarily on tenuous personal relationships. Secondly, it reveals how these same interpersonal relations can affect the construction of apparently ‘objective’ marketing strategies. Thirdly, it focuses on the problem of how all marketing campaigns are obliged to shift from ‘scientific’ to ‘artistic’ criteria as statistical data, information and analysis are converted into 1 linguistic and visual images for public consumption. Finally, it will make a few tentative comments on the relations between anthropology and marketing, with a view to developing a comparative theory of advertising as a marketing system, based on the cultural relativity of a specific marketing practice in a Japanese advertising agency (cf. Arnould 1995:110). The Discipline, Organisation and Practice of Marketing The Marketing Division is the engine room of the Japanese advertising agency in which I conducted my research in 1990. At the time, this agency handled more than 600 accounts a year, their value varying from several million to a few thousand dollars. The Marketing Division was almost invariably involved in some way in the ad campaigns, cultural and sporting events, merchandising opportunities, special promotions, POP constructions, and various other activities that the agency carried out on behalf of its clients. Exceptions were those accounts involving media placement or certain kinds of work expressly requested by a client – like, for example, the organisation of a national sales force meeting for a car manufacturer. Even here, however, there was often information that could be usefully relayed back to the Marketing Division (the number and regional distribution of the manufacturer’s sales representatives, as well as possible advance information on new products and/or services to be offered in the coming year). Marketing Discipline As Marianne Lien (1997:11) points out, marketing is both a discipline and a practice. The main aims as a discipline of the Marketing Division were (and, of course, still are): firstly, to acquire as much information as possible from consumers about their clients’ products and services; secondly, to acquire as much information as possible, too, from clients about their own products and services; and, thirdly, to use strategically both kinds of information acquired to develop new accounts. Marketing thus provided those working in the Marketing Division with the dispassionate data that account executives needed in their personal networking with (potential) clients whom they cajoled, persuaded, impressed and pleaded with to part with (more) money. Marketing Organisation In order to achieve the three overall objectives outlined above, the agency established a certain set of organisational features to enable marketing practice to take place. Firstly, the Marketing Division, which consisted of almost 90 members, was structured into three separate, but interlocking, sub-divisions. These consisted of Computer Systems; Market Development and Merchandising; and Marketing. The last was itself sub-divided into three departments, each of which was broken down into three or four sections. 1 Each section consisted of from six to a dozen members, led by a Section Leader, under whom they worked in teams of two to three on an account. These teams were not fixed. Thus one member, A, might work with another, B, under the Section Leader (SL) on a contact lens advertising campaign, but find herself assigned to worked with C under SL on an airline company’s business class service account, and with D under SL on a computer manufacturer’s consumer survey. In this respect, the daily life of members of the Marketing Note that, unlike the Marketing department in Viking foods discussed by Lien. Department was similar to that of product managers described by Lien (1997:69), being characterised by ‘frequent shifts from one activity to another, a wide network of communications, and a considerable amount of time spent in meetings or talking on the telephone’. Secondly, tasks (or accounts) were allocated formally through the hierarchical divisional structure – by departments first, then by sections – according to their existing responsibilities and perceived suitability for the job in hand. Each SL then distributed these tasks to individual members on the basis of their current overall workloads. At the same time, however, there was an informal allocation of accounts involving individuals. Each SL or DL could take on a job directly from account executives handling particular accounts on behalf of their clients. Here, prior experiences and personal contacts were important influences on AEs’ decisions as to whether to go through formal or informal channels of recruitment. The account executive in charge of the NFC contact lens campaign described in my book (Moeran 1996), for example, went directly to a particular SL in the Marketing Department because of some smart work that the latter had done for the AE on a different account some months previously. Mutual respect had been established and the contact lens campaign provided both parties with an opportunity to assess and, in the event, positively validate their working relationship. There were certain organisational advantages to the ways in which accounts were distributed in the manner described here. Firstly, by freely permitting interpersonal relations between account executives and marketers, the Agency ensured that there was competitiveness at each structural level of department and section. Such competition was felt to be healthy for the Agency as a whole, and to encourage its continued growth. Secondly, by assigning individual members of each section in the Marketing Department to working in different combinations of people on different tasks, the Agency ensured that each member of the Marketing Department received training in a wide variety of marketing problems and was obliged to interact fully with fellow section members, thereby promoting a sense of cooperation, cohesion and mutual understanding. This in itself meant that each section developed the broadest possible shared knowledge of marketing issues, because of the knowledge gained by individual members and the interaction among them. Marketing Practice Accounts were won by the Agency primarily through the liaison work conducted with a (potential) client by an account executive (who might be a very senior manager or junior ‘salesman’ recruited only a few years earlier). Once an agreement was made between Agency and client – and such an agreement might be limited to the Agency’s participation in a competitive presentation, the outcome of which might lead to an account being established – the AE concerned would put together an account team. An account team consists of the AE in charge (possibly with assistants); the Marketing Team (generally of 2 persons under a Marketing Director [MD], but sometimes much larger, depending on the size of the account and the work to be done); the Creative Team (consisting of Creative Director [CD], Copywriter, and Art Director [AD] as a minimum, but usually including two ADs – one for print-, the other for TV-related work); and Media Planner/Buyer(s). The job of the account team is to carry out successfully the task set by the client, and to this end meets initially for an orientation meeting in which the issues and problems relayed by the client to the AE are explained and discussed to all members. 2 Prior to this, however, the AE provides the marketing team with all the information and data that he has been able to extract from the client (a lot of it highly confidential to the company concerned). The marketing team, therefore, tends to come prepared and to have certain quite specific questions regarding the nature of the statistics provided, the target market, retail outlets, and so on. If it has done its homework properly – which is not always the case, given the number of different accounts on which the team’s members are working and the pressure of work that they are under – the marketing team may well have several pertinent suggestions for further research. It is on the basis of these discussions that the AE then asks the MD to carry out such research as is thought necessary for the matter in hand. In the meantime, the creative team is asked to mull over the issues generally and to think of possible ways of coping ‘creatively’ (that is, linguistically and visually) with the client’s marketing problems. Back in the Marketing Department, the MD will tell his subordinates to carry out specific tasks, such as a consumer survey to find out who precisely makes use of a particular product and why. This kind of task is fairly mechanical in its general form, since the Agency does this sort of work for dozens of clients every year, but has to be tailored to the present client’s particular situation, needs and expectations. The MD will therefore discuss his subordinate’s proposal, make some suggestions to ensure that all points are overed (and that may well include some additional questions to elicit further information from the target audience that has taken on importance during their discussion), and then give them permission to have the work carried out. All surveys of this kind are subcontracted by the Agency to marketing firms and research organisations of one sort or another. This means that the marketing team’s members are rarely involved in direct face-to-face contact or interaction with the consumers of the products that they wish to advertise,3 except when small ‘focus group’ interviews take place (usually in one of the Agency’s buildings). The informal nature of such groups, the different kinds of insights that they can yield, and the need to spot and pursue particular comments mean that members of the marketing team should be present to listen to and, as warranted, direct the discussion so that the Agency’s particular objectives are achieved. In general, however, the only evidence of consumers in the Agency is indirect, through reports, statistics, figures, data analyses and other information that, paradoxically, are always seen to be insufficient or ‘incomplete’ (cf. Lien 1997:112). Once the results of the survey are returned, the marketers enter them into their computers (since all such information is stored and can be used to generate comparative data for other accounts as and when required). They can make use of particular programmes to sort and analyse such data, but ultimately they need to be able to present their results in readily comprehensible form to other members of the account team. Here again, the MD tends to ensure that the information presented at the next meeting is to the point and properly hierarchised in terms of importance. This leads to the marketing team’s putting forward things like: a positioning statement, slogan, purchasing decision The Media Planners do not usually participate in these early meetings since their task is primarily to provide information of suitable media, and slots therein, for the finished campaign to be placed in. 3 A similar point is made by Lien (1997. 11) in her study of Viking Foods. Focus Groups usually consist of about half a dozen people who represent by age, gender, socio-economic grouping and so on the type of target audience being addressed, and who have agreed to talk about (their attitudes towards) a particular product or product range – usually in exchange for some gift or money. Interviews are carried out in a small meeting room (that may have a one-way mirror to enable outside observation) and tend to last between one and two hours. 4 2 4 odel (high/low involvement; think/feel product relationship), product message concept, and creative frame. One of the main objectives of this initial – and, if properly done, only – round of research is to discover the balance between what are terms product, user and end benefits, since it is these factors that determine the way in which an ad campaign should be presented and, therefore, how the creative team should visualise the marketing problems analysed and ensuing suggestions from the marketing team. It is here that we come to the crux of marketing as practised in an advertising agency (whether in Japan or elsewhere). Creative people tend to be suspicious of marketing people and vice-versa. This is primarily because marketers believe that they work rationally and that the creative frames that they produce are founded on objective data and analyses. Creative people, on the other hand, believe that their work should be ‘inspired’, and that such inspiration can take the place at the expense of the data and analyses provided for their consideration. As a result, when it comes to producing creative work for an ad campaign, copywriters and creative directors tend not to pay strict attention to what the marketing team has told them. For example, attracted by the idea of a particular celebrity or filming location, they may come up with ideas that in no way meet the pragmatic demands of a particular ad campaign that may require emphasis on product benefits that are irrelevant to the chosen location or celebrity suggested for endorsement. This does not always happen, of course. A good and professional creative team – and such teams are not infrequent – will follow the marketing team’s instructions. In such cases, their success is based on a creative interpretation of the data and analyses provided. Agency-Client Interaction If there is some indecision and argument among different elements of the account team – and it is the presiding account executive’s job to ensure that marketers and creatives do not come to blows over their disagreements – they almost invariably band together when meeting and presenting their plans to the client. Such meetings can take place several, even more than a dozen, times during the course of an account team’s preparations for an ad campaign. At most of them the MD will be present, until such time as it is clear that the client has accepted the Agency’s campaign strategy and the creative team has to fine-tune the objectives outlined therein. Very often, therefore, the marketing team will not stay on a particular account long enough to learn of its finished result, although a good AE will keep his MD abreast of creative developments and show him the (near) finalised campaign prior to the client’s final approval. But marketers do not get involved in the production side of a campaign (studio photography, television commercial filming, and so on) – unless one of those concerned knows what is going on when, happens to be nearby at the time, and drops in to see how things are going. In other words, the marketing team’s job is to see a project through until accepted by the client. It will then dissolve and its members will be assigned to new accounts. Advertising Campaigns: A Case Study To illustrate in more detail particular examples of marketing practice in the Agency, let me cite as a case study the preparation of contact lens campaign in Japan. This example is illuminating because it reveals a number of typical problems faced by an advertising agency in the formulation and execution of campaigns on behalf of its clients. These include the interface between marketing and creative people within an agency and the interpretation of marketing analysis and data; the 5 transposition of marketing analysis into ‘creative’ (i. e. linguistic, visual and design) ideas; the interface between agency and client in the ‘selling’ of a campaign proposal; and the problems of having to appeal to more than one ‘consumer’ target. When the Nihon Fibre Corporation asked the Agency to prepare an advertising campaign for its new Ikon Breath O2 oxygen-passing GCL hard contact lenses in early 1990, it provided a considerable amount of product information with which to help and guide those concerned. This information included the following facts: firstly, with a differential coefficient (DK factor) of 150, Ikon Breath O2 had the highest rate of oxygen permeation of all lenses currently manufactured and marketed in Japan. As a result, secondly, Ikon Breath O2 was the first lens authorized for continuous wear by Japan’s Ministry of Health. Thirdly, the lens was particularly flexible, dirt and water resistant, durable, and of extremely high quality. The client asked the Agency to confirm that the targeted market consisted of young people and to create a campaign that would help NFC capture initially a minimum three per cent share of the market, rising to ten per cent over three years. The Agency immediately formed an account team, consisting of eight members all told. Their first step was to arrange for the marketing team to carry out its own consumer research before proceeding further. A detailed survey – of 500 men and women – was worked out in consultation with the account executive and the client, and was executed by a market research company subcontracted by the Agency. Results confirmed that the targeted audience for the Ikon Breath O2 advertising campaign should be young people, but particularly young women, between the ages of 18 and 27 years, since it was they who were most likely to wear contact lenses. At the same time, however, the survey also revealed that there was little brand loyalty among contact lens wearers so that, with effective advertising, it should be possible to persuade users to shift from their current brand to Ikon Breath O2 lenses. It also showed that young women were not overly concerned with price provided that lenses were safe and comfortable to wear, which meant that Ikon Breath O2’s comparatively high price in itself should not prove a major obstacle to brand switching or sales. On a less positive note, however, the account team also discovered that users were primarily concerned with comfort and were not interested in the technology that went into the manufacture of contact lenses (thereby obviating the apparent advantage of Ikon Breath O2’s high DK factor of which NFC was so proud); and that, because almost all contact lens users consulted medical specialists prior to purchase, the advertising campaign would have to address a second audience consisting mainly of middle-aged men. All in all, therefore, Ikon Breath O2 lenses had an advantage in being of superb quality, approved by medical experts and recognized, together with other GCL lenses, as being the safest for one’s eyes. Its disadvantages were that NFC had no ‘name’ in the contact lens market and that users knew very little about GCL lenses or contact lenses in general. This meant that the advertising campaign had to be backed up by point of purchase sales promotion (in the form of a brochure) to ensure that the product survived. Moreover, it was clear that Ikon Breath O2’s technical advantage (the DK 150 factor) would not last long because rival companies would soon be able to make lenses with a differential coefficient that surpassed that developed by NFC. 5 On this occasion, because the advertising budget was comparatively small, the media buyer was not brought in until later stages in the campaign’s preparations. The AE in charge of the NFC account interacted individually with the media buyer and presented the latter’s suggestions to the account team as a whole. 6 As a result of intense discussions following this survey, the account team moved slowly towards what it thought should be as the campaign’s overall ‘tone and manner’. Ideally, advertisements should be information-oriented: the campaign needed to put across a number of points about the special product benefits that differentiated it from similar lenses on the market (in particular, its flexibility and high rate of oxygen-permeation). Practically, however – as the marketing team had to emphasize time and time again – the campaign needed to stress the functional and emotional benefits that users would obtain from wearing Ikon Breath O2 lenses (for example, continuous wear, safety, release from anxiety and so on). This meant that the advertising itself should be emotional (and information left to the promotional brochure) and stress the end benefits to consumers, rather than the lenses’ product benefits. Because the marketing team had concluded that the product’s end benefits should be stressed, copywriter and art director opted for user imagery rather than product characteristics when thinking of ideas for copy and visuals. However, they were thwarted in their endeavours by a number of problems. Firstly, advertising industry self-policing regulations prohibited the use of certain words and images (for example, the notion of ‘safety’, plus a visual of someone asleep while wearing contact lenses), and insisted on the inclusion in all advertising of a warning that the Ikon Breath O2 lens was a medical product that should be purchased through a medical specialist. This constriction meant that the creative team’s could not use the idea of ‘continuous wear’ because, even though so certified by Japan’s Ministry of Health, opticians and doctors were generally of the opinion that Ikon Breath O2 lenses were bound to affect individual wearers in different ways. NFC was terrified of antagonizing the medical world which would often be recommending its product, so the product manager concerned refused to permit the use of any word or visual connected with ‘continuous wear’. Thus, to the account team’s collective dismay, the product’s end benefit to consumers could not be effectively advertised. Secondly, precisely because Ikon Breath O2 lenses had to be recommended by medical specialists, NFC’s advertising campaign needed to address the latter as well as young women users. In other words, the campaign’s tone and manner had to appeal to two totally different segments of the market, while at the same time satisfying those employed in the client company. This caused the creative team immense difficulties, especially because – thirdly – the product manager of NFC’s contact lens manufacturing division was convinced that the high differential coefficient set Ikon Breath O2 lenses apart from all other contact lenses on the market and would appeal to members of the medical profession. So he insisted on emphasizing what he saw as the unique technological qualities of the product. In other words, not only did he relegate young women who were expected to buy the product to secondary importance; he ignored the marketing team’s recommendation that user benefit be stressed. Instead, for a long time he insisted on the creative team’s focussing on product benefit, even though the DK factor was only a marginal and temporary advantage to NFC. As a result of these two sets of disagreements, the copywriter came up with two different key ideas. The first was based on the product’s characteristics, and thus supported the manufacturer’s (but went against his own marketing team’s) product benefit point of view, with the phrase ‘corneal physiology’ (kakumaku seiri). The second also stressed a feature of the product, but managed to emphasize the user benefits that young women could gain from wearing lenses that were both ‘hard’ and ‘soft’ (yawarakai). The former headline was the only way to break brand parity and make Ikon Breath O2 temporarily distinct from all other lenses on the market (the product manager liked the distinction; the marketing team disliked the temporary nature of that distinction). At this stage in the negotiations, the account executive in charge felt obliged to tow an obsequious line, but needed to appease his marketing team and ensure that the creative team came up with something else if at all possible, since 7 corneal physiology gave Ikon Breath O2 lenses only a temporary advantage. As a result, the copywriter introduced the word ‘serious’ (majime) into discussions – on the grounds that NFC was a ‘serious’ (majime) manufacturer (it was, after all, a well-known and respected Japanese corporation) which had developed a product that, by a process of assimilation, could also be regarded as ‘serious’; moreover, by a further rubbing-off process, as the marketing team agreed, such ‘seriousness’ could be attributed to users who decided to buy and wear Ikon Breath O2 lenses. In this way, both the distinction between product benefit and user benefit might be overcome. The copywriter’s last idea was the one that broke the deadlock (and it was at certain moments an extremely tense deadlock) between the account team as a whole and members of NFC’s contact lens manufacturing division. After a series of meetings in which copywriter and designer desperately tried to convince the client that the idea of softness and hardness was not a product characteristic, but an image designed to support the benefits to consumers wearing Ikon Breath O2 lenses, the product manager accepted the account team’s proposals in principle, provided that ‘serious’ was used as a back-up selling point. Soft hard’ (yawaraka hard) was adopted as the key headline phrase for the campaign as a whole. It can be seen that the marketing team’s analysis of how NFC should successfully enter the contact lens market met two stumbling blocks during the early stages of preparation for the advertising campaign. The first was within the account team itself, where the copywriter in particular tended to opt for the manufacturer’s approach by emphasising the product benefit of Ikon Breath O2. The second was when the Agency’s account team had to persuade the client to accept its analysis and campaign proposal. But the next major problem facing the account team was how to convert this linguistic rendering of market analysis into visual terms. What sort of visual image would adequately fulfil the marketing aims of the campaign and make the campaign as a whole – including television commercial and promotional materials – readily recognizable to the targeted audience? It was almost immediately accepted by the account team that the safest way to achieve this important aim was to use a celebrity or personality (talent in Japanese) to endorse the product. Here there was little argument, because it is generally recognized in the advertising industry that celebrity endorsement is an excellent and readily appreciated linkage device in multi-media campaigns of the kind requested by NFC. Moreover, since television commercials in Japan are more often than not only fifteen seconds long and therefore cannot include any detailed product information, personalities have proved to be attention grabbers in an image-dominated medium and to have a useful, short-term effect on sales because of their popularity in other parts of the entertainment industry. At the same time, not all personalities come across equally well in the rather differing media of television and magazines or newspapers, so that the account team felt obliged to look for someone who was more than a mere pop idol and who could act. It was here that those concerned encountered the most difficulty. The presence of a famous personality was crucial since s/he would be able to attract public attention to a new product and hopefully draw people into retail outlets to buy Ikon Breath O2 lenses. It was agreed right from the start that the personality should be a young woman, in the same age group as the targeted audience, and Japanese. (After all, a ‘blue eyed foreigner’ endorsing Ikon Breath O2 contact lenses would hardly be appropriate for brown-eyed Japanese. ) Just who this woman should be, however, proved problematic. Tennis players (who could indulge in both ‘hard’ activities and ‘soft’ romance) were discarded early on because the professional season was already in full swing at the time the campaign was being prepared. Classical musicians, while romantic and thus ‘soft’, were not seen to be ‘hard’ enough, while the idea of using a Japanese ‘talent’, Miyazawa Rie (everyone on the account team’s favourite at the time), was reluctantly rejected because, even though photographs of her in the nude were at the time causing a 8 minor sensation among Japanese men interested in soft-porn, she was rather inappropriate for a medical product like a contact lens which was aimed at young women. Any personality chosen had to show certain distinct qualities. One of these was a ‘presence’ (sonzaikan) that would attract people’s attention on the page or screen. Another was ‘topicality’ (wadaisei) that stemmed from her professional activities. A third was ‘future potential’ (nobisei), meaning that the celebrity had not yet peaked in her career, but would attract further widespread media attention and so, it was hoped, indirectly promote Ikon Breath O2 lenses and NFC. Most importantly, however, she had to suit the product. In the early stages of the campaign’s preparations, the creative team found itself in a slight quandary. They wanted to choose a celebrity whose personality fitted the ‘soft-hard’ and ‘serious’ ideas and who would then anchor a particular image to Ikon Breath O2 lenses, although it proved difficult to find someone who would fit the product and appeal to all those concerned. Eventually, the woman chosen was an actress, Sekine Miho, who epitomized the kind of modern woman that the creative team was seeking, but who was also about to star in a national television (NHK) drama series that autumn – a series in which she played a starring role as a ‘soft’, romantic character. Although popularity in itself can act as a straightjacket when it comes to celebrity endorsement of a product, in this case it was judged – correctly, it transpired – that Sekine had enough ‘depth’ (fukasa) to bring a special image to Ikon Breath O2 lenses. Once the celebrity had been decided on, the creative team was able to fix the tone and manner, expression and style of the advertising campaign as a whole. Sekine was a ‘high class’ (or ‘one rank up’ in Japanese-English parlance) celebrity who matched NFC’s image of itself as a ‘high class’ (ichiryu) company and who was made to reflect that sense of eliteness in deportment and clothing. At the same time, NFC was a ‘serious’ manufacturer and wanted a serious, rather than frivolous, personality who could then be photographed in soft-focus, serious poses to suit the serious medical product being advertised. This seriousness was expressed further by means of ery slightly tinted black and white photographs which, to the art director’s – but, not initially, the product manager’s – eye made Sekine look even ‘softer’ in appearance and so match the campaign’s headline of yawaraka hard. This softness was further reinforced by the heart-shaped lens cut at the bottom of every print ad, and on the front of the brochure, which the art director m ade green rather than blue – partly to differentiate the Ikon Breath O2 campaign from all other contact lens campaigns run at that time, and partly to appeal to the fad for ‘ecological’ colours then-current among young women in particular. This case study shows that there is an extremely complex relationship linking marketing and creative aspects of any advertising campaign. In this case, market research showed that Ikon Breath O2 lenses were special because of the safety that derived from their technical quality, but that consumers themselves were not interested in technical matters since their major concern was with comfort. Hence the need to focus the advertising campaign on user benefit. Yet the client insisted on stressing product benefit – a stance made more difficult for the creative team because it could not legally use the only real consumer benefit available to it (continuous wear), and so had to find something that would appeal to both manufacturer and direct and indirect ‘consumers’ of the lens in question. In the end, the ideas of ‘soft hard’ and ‘serious’ were adopted as compromise positions for both client and agency, as well as for creative and marketing teams. Concluding Comments Let us in conclusion try to follow two separate lines of thought. One of these is, as promised, the relationship between marketing and anthropology; the other that between advertising and marketing. 9 Although convergence between anthropology, marketing and consumer research may be growing, the evidence suggested by the case study in this paper is that huge differences still exist. Marketing people in the advertising agency in which I studied may be interested in anthropology; they may even have dipped into the work of anthropologists here and there. But their view of the discipline tends to be rather old-fashioned, and they certainly do not have time to go in for the kind of intensive, detail ethnographic nquiry of consumers that anthropologists might encourage. If anthropologists are to make a useful contribution to marketing, therefore, they need to present their material and analyses succinctly and in readily digestible form, since marketing people hate things that are overcomplicated. It is, perhaps, for this rather than any other reason that someone like Mary Douglas (Douglas and Isherwood 1979) has been so favourably received. In the end, marketing people aim to be positivist, science-like (rather than scientific, as such), and rationalist in their ad campaigns. They aspire to measure and predict on the basis of observer categories, if only because this is the simplest way to sell a campaign to a client. In this respect, they are closer to the kind of sociology and anthropology advocated in the 1940s and 50s (which would explain their adoption of Talcott Parsons’s theory of action, for example), than to the present-day ‘interpretive’ trends in the discipline, and thus favour in their practices an outmoded – and among most anthropologists themselves, discredited – form of discourse. So, ‘if anthropologists are kings of the castle, it is a castle most other people have never heard of’ (Chapman and Buckley 1997: 234). As Malcolm Chapman and Peter Buckley wryly observe, we need perhaps to spend some time entirely outside social anthropology in order to be convinced of the truth of this fact. Secondly, as part of this positivist, science-like approach, marketers in the Japanese advertising agency tended to make clear-cut categories that would be easily understood by both their colleagues in other divisions in the Agency and by their clients. These categories tended to present the consumer world as a series of binary oppositions (between individual and group, modern and traditional, idealist and materialist, and so on [cf. Lien 1997: 202-8]) that they then presented as matrix or quadripartite structures (the Agency’s Purchase Decision Model, for example, was structured in terms of think/feel and high/low involvement axes). In this respect, their work could be said to exhibit a basic form of structuralism. One of these oppositions was that made between product benefit and user benefit (with its variant end benefit). As this case study has shown, this is a distinction that lies at the heart of all advertising and needs to be teased out if we are successfully to decode particular advertisements in a manner that goes beyond the work of Barthes (1977), Williamson (1978), Goffman (1979) and others. Thirdly, one of the factors anchoring marketing to the kind of structured thinking characteristic of modernist disciplines, perhaps, is that the creation of meaning in commodities is inextricably bound up with the establishment of a sense of difference between one object and all others of its class. After all, the three tasks of advertising are: to stand out from the surrounding competition to attract people’s attention; to communicate (both rationally and emotionally) what it is intended to communicate; and to predispose people to buy or keep on buying what is advertised. The sole preoccupation of those engaged in the Ikon Breath 02 campaign was to create what they referred to as the ‘parity break’: to set NFC’s contact lenses apart from all other contact lenses on sale in Japan, and from all other products on the market. At the same time, the idea of parity break extended to the style in which the campaign was to be presented (tinted monochrome photo, green logo, and so on). In this respect, the structure of meaning in advertising is akin to that found in the syntagmatic and paradigmatic axes of structural linguistics where particular choices of words and phrases are influenced by the overall structure and availability of meanings in the language in which a speaker is communicating. That the work of LeviStrauss should be known to most marketers, therefore, is hardly surprising. Marketing practice is in many respects an application of the principles of structural anthropology to the selling of products. 10 Fourthly, although those working in marketing and consumer research take it as given that there is one-way flow of activity stemming from the manufacturer and targeted at the end consumer, in fact, as this case study shows, advertising – as well as the marketing that an advertising agency conducts on behalf of a client – always addresses at least two audiences. One of these is, of course, the group of targeted consumers (even though they are somewhat removed from the direct experience of marketers in their work). In this particular case, to complicate the issue further, there were two groups of consumers, since the campaign had to address both young women and middle-aged male opticians. Another audience is the client. As we have seen, the assumed or proven dis/likes of both consumers and advertising client affect the final meaning of the products advertised, and the client in particular had to be satisfied with the Agency’s campaign approach before consumer ‘needs’ could be addressed. At the same time, we should recognise that a third audience exists among different members of the account team within the Agency itself, since each of the three separate parties involved in account servicing, marketing and creative work needed to be satisfied by the arguments of the other two. In this respect, perhaps, we should note that marketing people have spent a lot of time over the decades making use of insights developed in learning behaviour, personality theory and psychoanalysis which they then apply to individual consumers. In the process, however, they have tended to overlook the forms of social organisation of which these individuals are a part (cf. de Groot 1980:44). Yet it is precisely the ways in which individual consumers interact that is crucial to an understanding of consumption and thus of how marketing should address its targeted audience: how networks function, for example, reveals a lot about the vital role of word-of-mouth in marketing successes and failures; how status groups operate and on what grounds can tell marketers a lot about the motivations and practices of their targeted audience. Anthropologists should be able to help by providing sociological analyses of these and other mechanisms pertinent to the marketing endeavour. In particular, their extensive work on ritual and symbolism should be of use in foreign, ‘third world’ markets. Fifthly, most products are made to be sold. As a result, different manufacturers have in mind different kinds of sales strategies, target audiences, and marketing methods that have somehow to be translated into persuasive linguistic and visual images – not only in advertising, but also in packaging and product design. For the most part, producers of the commodities in question find themselves obliged to call on the specialized services of copywriters and art designers who are seen to be more in tune with the consumers than are they themselves. This is how advertising agencies market themselves. But within any agency, the creation of advertising involves an ever-present tension between sales and marketing people, on the one hand, and creative staff, on the other; between the not necessarily compatible demands for the dissemination of product and other market information, on the one hand, and for linguistic and visual images that will attract consumers’ attention and push them into retail outlets to make purchases, on the other. This is not always taken into account by those currently writing about advertising. More interestingly, perhaps, the opposition that is perceived to exist between data and statistical analysis, on the one hand, and the creation of images, on the other, parallels that seen to pertain between a social science like economics or marketing and a more humanities-like discipline such as anthropology. Perhaps the role for an anthropology of marketing is to bridge this great divide.

Virtue Ethics Essay Example for Free

Virtue Ethics Essay In the field of bioethics, Aristotles ethics does not have the same practical force as action-based theories, especially when the dilemma in question involves deciding whether or not to do some particular action (pulling the plug on a patient). In any case, there are two ways we can apply the character based approach to bioethics. First, the specific healthcare provider (nurse, doctor, etc.) could continually ask themselves What kind of character do I want to have as a healthcare provider, and what actions are consistent with this character?. A healthcare provider can ask themselves whether a certain action reflects the kind of character they want to be associated with. On a more general scale, the healthcare industry itself could ask itself What kind of character do we want associated with the healthcare industry and how would this particular action contribute or take away from that character? Secondly, for Aristotle a virtuous action is a mean between two extremes. For example, bravery is the middle-point between cowardice and foolhardiness. However, it is not always clear whether every action is a mean of this way. How would pulling the plug on a patient fit in Aristotles golden mean analysis? I think the safest way to apply Aristotles theory would be to emphasize character, rather than get lost in the thickets of golden mean analysis. Using Aristotelian ethics to resolve particular cases is always difficult, because the question of what ought I to do? is always secondary to the question what ought I to become?. After all, Aristotle himself admits that we should not expect precision in ethical matters.

Preparation For Providing Effective And Efficient Mentorship Nursing Essay

Preparation For Providing Effective And Efficient Mentorship Nursing Essay In preparation for this assignment searches have been conducted using the following databases: Cinahl, Pubmed and Medline search terms Learning theories, Learning styles, Mentorship and Practice Education Facilitator were used singularly or in combination. Manual searching of relevant nursing journals and publications have been performed. Articles published in English in the last 10 years were reviewed. This assignment will examine the issues and processes that need to be considered whilst facilitating and supporting a student to meet their learning outcomes in a practice placement, there will be discussion regarding the assessment strategies available as a mentor, which contribute to the overall assessment of students. Furthermore learning styles and theories will be discussed, along with the clinical setting as an environment to facilitate the learning needs required to achieve success in completing the practice placement learning outcomes. A brief overview of the clinical setting will be provided to capture the ambiance of the learning environment. The setting for this location is an Elective Orthopaedic Unit within the Northwest and in conforming to the Nursing and Midwifery Council (NMC (2008); any names used will be pseudonyms to protect identification, confidentiality and dignity will be maintained at all times. NMC (2008) states that a mentor is a registered nurse who has completed the approved mentorship programme and meets the criteria set out in NMC (2008) standards for mentors, practice teachers and teachers. Whereas Gopee (2008) argues the term mentor is frequently used to mean trusted friend, guide and advisor. There is also identified the eight mandatory standards that must be achieved to become a mentor, the author will be referring to these standards to support learning and development in practice. To ensure the student who will be named as Lucy for the purpose of this assignment has a positive learning experience it is paramount that learning environment is supportive for Lucy and for those who work alongside her. An orientation pack and welcoming letter are provided with useful information and an introduction to the allocated mentor is provided when ever possible on attendance the first shift. Kenworthy and Nicklin (2000) support the view that the more comfortable and welcome a student feels within the working environment, the more likely it is that effective learning will take place which is a key factor in successful learning. Pearcy and Elliot (2004) found that students who have had negative experiences with their mentor and placements, tend to leave the course before qualifying however Beskine (2009) advocates that starting off with a positive approach promotes the students experience as well as the quality of the placement. Adhering to the mandatory standards set by the NMC (2008) Standards to Support Learning and Assessment in Practice; and in order to establish the stage of Lucys training and her learning needs and style a mutually agreed time and location is identified for the initial meeting to take place. This meeting provides the opportunity to co-ordinate off duty and to identify another team member who will be available to support Lucys learning needs in the absence of her mentor. Once Lucys learning style had been established, then negotiating the best way forward to facilitate the learning experiences relevant to the placement can take place This provides the opportunities for Lucy to meet all of the learning outcomes and needs. Direction towards self-management of learning should be encouraged to help promote professional development, and to instil the importance of responsibility and accountability. At the half way point Lucy and her mentor will have a formative assessment, evaluation of her progress this will determine any action plans or adjustments necessary to enable Lucy to meet her learning outcomes. NMC standards for mentors (2008) state that a mentor must contribute to evaluation of student learning and assessment experiences, suggesting aspects for change resulting from such evaluation. A summative assessment would usually take place when completing student documents whilst focusing on the whole of the placement. In a study by Hart and Rotem (1994) it is indicated that the qualities of a mentor are a vital factor in providing and achieving a positive and valuable learning environment; moreover the study found that stressful events for nursing students during clinical practice have shown the initial clinical experience was the most anxiety producing part of clinical experience Cahill (1996) acknowledges that the single most crucial factor in creating a positive learning environment is the relationship between staff and the student nurse; it is also discussed that a common problem that arises for a student is that they are regularly unable to work with their mentor. Many nurses and nursing students in the past found themselves to be autonomous practitioners, learning their skills within the situations they found themselves in without adequate guidance or support (Bray Nettleton, 2008). Whereas Higgins et al (2009) suggest the success of any nursing student in any program in any clinical setting is complex, enhancing the complicated nature of education and the perception of competencies whether student or mentor. This is an important concept to be sensitive to, as the process of moving forward with the knowledge and skills needs to be supported and nurtured in order to facilitate the standard of care that is safe, accountable, and competent. Honey and Mumford (1992) Identified If mentors are to teach students effectively they must be aware of different learning styles and the importance of them; furthermore Honey and Mumford explore four different styles of learning and is a variation on Kolbs learning cycle, which was originally adapted from Kolb and Fry (1975). It is thought whenever possible, it is essential to allow the students individual style of learning determine the choice of teaching. Within Kolbs model there are four different styles of learning they describe, activist, theorist, pragmatist, and reflector. They also propose that although many people can be a mixture of the four styles, they primarily have a preference for only one. However Clark (2007) suggests that labelling such as activist versus reflector has no scientific justification. Downie and Basford (2003) believe, that to identify the learning outcomes at the beginning of the placement is of great benefit to the student, knowing the stage of training that the student is currently at helps to support a student effectively within the clinical setting; moreover they suggest that most students prefer the traditional learning method described as pedagogy which is where the mentor or teacher has full responsibility for what the student learns. Andragogy is described by Knowles (1990) as adult learning where the student is motivated and applies life experiences to learning, therefore this suggests the use of andragogy teaching method would be beneficial to Lucy as she is very motivated in learning outside if the learning outcomes. When establishing Lucys learning style, formal teaching sessions were introduced to demonstrate how theory is applied in practice. A taxonomy frequently used in nursing is the framework by Benner (1984) in which there are five levels; novice, beginner, competent, proficient and expert. According to Benners stages of clinical competence, which shows the journey of a novice to an expert then Lucy appears to be an advanced beginner as signs of understanding NMC The code: standards of conduct, performance and ethics for nurses and midwifes have been identified along with previous experience in other clinical areas. As required By NMC (2008) Standards to Support Learning and Assessment in Practice along with fostering professional growth, the mentor encourages Lucy to reflect on her practice and empowered Lucy to be autonomous within the limitations of the supervised role, whilst being fair and accurate in the assessment of the student. Validity is the most crucial aspect of any assessment; it shows the extent to which the assessment measures and what it is expected to measure. A valid assessment is one that assesses learning outcome; it is set out to assess and not to target other outcomes that may not have been learnt adequately at that point. Reliability is also a key factor to be considered in assessments, the term reliability is used to indicate the consistency of which an assessment measures and what it is designed to measure. However Goding (1997) suggests that reliability and validity are inappropriate indicators of nursing practice and that they reduce practice to a list of objectives, which holds very little relation to the intricacy of practice. The NMC (2006) advocate that assessment is said to be reliable, if it gives similar results when used on different occasions, and with different assessors. It is suggested by Hand (2006) that continuous assessment allows the student to achieve their target through progressive goal setting. Whereas Clifford (1994) explains continual assessment of practice, implies the students performance is monitored on a daily basis during clinical practice, which would be done on a day-to-day basis. Regular meetings between Lucy and her mentor allow discussions with mutual respect to evaluate the progress she is or may not be making, enabling action plans to be implemented if necessary. According to NMC (2008) the mentor should provide the student with constructive feedback and assist them in identifying future learning needs and actions. Employing evidence-based practice, observation and communication equips the mentor with some of the necessary tools to identify the strengths and weaknesses and provide feedback; remaining objective when assessing the student to meet the required outcomes also means being objective when completing student documentation. It could be tempting when assessing Lucy to overeat her performance causing the halo effect, although by underrating her performance could cause the horn effect; this could happen if there is a clash of personalities between a student and a mentor. Krech et al (1962) document that human tendencies can influence a manager or mentors point of view causing stereotyping and the halo and horn effect Research by Watson Harris (1999) that examines support of students in practice placement established that some student nurses are being allowed to pass clinical assessments without having demonstrated sufficient competence; the study revealed that some Practioners did not feel it was their responsibility to fail students. Moreover Duffy (2003) reports that it is inevitable that some students will not be able to meet the required level of practice and it is essential that mentors do not avoid the difficult issue of having to fail these students. Duffy (2003) identified that weak students tend to have poor interpersonal skills, are disinterested in practice learning and are frequently late, and that they also lack personal insight and the awareness of professional boundaries. Duffy goes on to recommend that early intervention is essential if mentors or the clinical team notice an issue or concern about students, time must be made to discuss the issues. There may be other factors affecting performance and these mitigating circumstances may need to be considered, however performance must meet the required standard. The NMC (2008) Standards to support learning and assessment in practice state the mentor is accountable and responsible for assessing the total performance of the student including attitude, skills and behaviour. There are fundamental individuals that can offer support and guidance to the mentor. The mentor can access support in assessing a student by way of the links to the Practice Education Facilitator (PEF), whose multidimensional role includes; according to Salvoni (2001) helping to support and enhance clinical practice and teach student nurses practical skills within the academic setting, in recognition of the theory-practice relationship. In addition to this Rowan Barber (2000) and Richards et al (2001) support the view that the PEF furthers staff in the development of their teaching and assessment skills. The PEF, University Link Lecturer and The Academic Adviser are pivotal in supporting and guiding both the mentor and the student if there have been concerns expressed in a students progress within the placement. There is a potential that a mentor could be mentoring a student who is not able to achieve within the placement, if a mentor has concerns it is essential that this concerns be acted upon in the appropriate and in a timely manner. Duffy (2003) recognised mentors do not always identify and deal with problems early enough in students placement, they find it difficult to fail a student, in order to prepare a mentor for their role and the responsibilities in failing the student it is vital that they are supported and backed up by the PEF. The NMC (2008) states mentors should manage failing students to enable them to enhance their performance and capabilities for safe and effective practice. The student should also understand their failure and the implications of this for their future. NMC (2006) states students must evaluate their placement as part of the educational audit process. Also as the named mentor you are responsible for making the final assessment and are accountable for passing or failing the student. It is inescapable that some students will not be able to achieve the required level of competence whilst in the practice placement and it is essential that mentors do not evade the difficult issue of having to fail a student. In conclusion this assignment has examined a plethora of evidence, research, reviews and studies to elicit discussions and reflection surrounding what issues and processes have to be considered whilst supporting a student in a practice placement along with comparing the range of assessment strategies accessible to mentors which, contributes to the overall assessment of students. Wilkinson (1999) argues liasing between clinicians and educationalist enhances the quality of assessment and promotes a practice based learning culture. Learning styles and learning theories have been investigated. It is important to assess each student as an individual in the context of their learning needs to strengthen the integration of theory and practice within the clinical setting likewise it is imperative that the mentor has an understanding of the learning outcomes. The mentor was able to pass Lucy in her practice placement and maintained professional boundaries throughout the experience.

Polar Bear :: essays research papers

SPECIES- Ursus Maritimus, Polar Bear, lord of the Arctic. Lives in an area of five million square miles of snow and ice. From Siberia to Alaska and across Canada, Greenland and the Islands north of Norway, he is the master of all living things except man. It lives in the brutal cold, ice, and snow. The temperature can plunge down frequently to -40 degrees and sometimes even lower but that does not bother the polar bear because of its color-less skin and layer of insulation fat. Its range extending around the northern polar region. Necessities of Life-The polar bear eats mostly seals which he has to hunt. His trick is to wait by a breathing in the ice and when a seal comes up by that breathing hole, he grabs it so fast it knocks it unconscious and then he eats it. Other pray is a walrus calf or a musk ox stuck in snow, birds, eggs, fish, and dead whales. And sometimes in the summer it eats berries and grass. The polar bear has no water to drink so it only eats the skin and blubber, avoiding the meat. So by eating the blubber and leaving the meat the male bear is helping keep his body in balance with the surrounding environment. The bear would usually stay in a den or bury it self in the snow to avoid the suns ultra violent rays. Usually the pregnant female polar bear stays in the den.Food Chain-The polar bear finds its way on the top of its food chain. No predator on earth approaches the bear in size. The polar bear towers over everything else in his food chain. The only thing that could stop him is a human with a gun.Super Amazing Facts-The polar bear is well adapted to its special live style. The hairs of its thick coat are hollow, giving extra insulation against the bitter arctic cold. It also has an extra layer of insulating fat under its skin and the hairs on the polar bear are color less, when the sun hits them they look white. Polar bears are twice as big as lions or tigers, a typical adult male weighs half a ton and is five feet high at the shoulder. His paws are a fool wide. When he stands up straight, he can look an elephant in the eye.Explain- My species was endangered bad back in 1965 with no more than 5,000 left in the world.

The friar - An Analysis Essay -- Essays Papers

The friar - An Analysis Character Analysis The Friar-- Humble Shepherd or Crafty Wolf? Chaucer was known for his ironic descriptions of various sojourners in the Canterbury Tales . The description given to the Friar in the "General Prologue" does not stray from Chaucer’s trademark. The Friar is described as a "limitour" [begs on the behalf of the poor], yet we see that he is a bachelor on a love hunt, a crooked businessman and does complete his duties as a Friar. The Friar knows many beautiful women, many affluent men, and rarely associates with the class of people he should live among. The Friar’s duties were to live among the poor, to beg on their behalf and to give his earnings to aid their struggle for livelihood. However, Chaucer allows the reader to see the true character of the Friar. He knows: â€Å"so muche of daliance and fair language..† (Norton 211). This no doubt is a way to woo women with sweet words and a crafty tongue. This strategy is also in lines 265-266: â€Å"somwhat he lipsed for his wantounesse to make his English sweete upon his tongue... † This was repeated in lines 215-217: â€Å"Ful wel biloved and familer was he..... with the worthy wommen of the town-" A Friar’s duties was not supposed to flirt with the women of the town but to beg for poor. The Friar, using what money he has earned â€Å"his tipet was ay farsed ful of knives and pinnes for to yiven faire wives..† (233-234). This states that he buys gifts for women as well. The Friar, as it turns out, is not begging for money to appease his goal to fee d the poor, but rather is wooing women to appease his flesh! The Friar is not just a ladies' man under the guise of a humanitarian, he is also a crooked businessman. He uses his position in the church to get... ...on, E. Talbot, et al. The Norton Anthology of English Literature. Ed. M.H. Abrams. New York. W.W. Norton & Company,  © 1993 http://www.freeimages.com Provided by The Free Site. Online. Internet. 14 Dec 1998. http://www.freeimages.com http://www.barrysclipart.com Gowen, Barry. Online. Internet. 14 Dec 1998. http://barrysclipart.com http://www.mhare.com/mhare/FriarFolk.htm March Hare © 1998. Online. Internet. 15 Dec 1998. http://www.mhare.com/mhare/FriarFolk.htm http://wally.rit.edu/cary/19th_century/16.html Millersville Univesity. Online. Internet. 15 Dec 1998. http://wally.rit.edu/cary/19th_century/16.html http://icg.harvard.edu/~chaucer/ Harvard University. Online. Internet. 15 Dec 1998. http://icg.harvard.edu/~chaucer/ Deluxe Ellesmere manuscript of the Canterbury Tales, housed in the Huntington Library, in San Marino, California. 21 Dec 1998.

Class Conflict in Britain :: Politics Political

Class Conflict in Britain "Class conflict has gradually been diluted by growing affluence." "The history of all hitherto existing societies is the history of class struggle." This famous opening line from Marx Communist Manifesto refers to the struggle between the labouring, working classes and the bourgeoisie owners of the means of production. The proletariat are exploited by the capitalists for profit and are therefore forced to live in poverty and dire conditions. Marx predicted that eventually the proletariat would overthrow this capitalist system and replace it with a system which is often referred to as Communist - whereby the workers have control. Today, whenever the words 'class' or 'class conflict' are mentioned people usually turn to Marx definition and picture the poor worker fighting for better pay, better living and working conditions. The typical class conflict is typified as workers versus the owners, or bourgeoisie. In Britain this struggle did not develop in the way that Marx predicted - there has never been a genuine proletariat revolutionary threat. In its place has been a tradition of reformist socialism with the Labour Party and the Trades Unions being the main campaigners. In Britain the traditional class conflict is often depicted as Labour Party versus Conservative Party. The Labour Party have fought for workers rights and have been supported at elections by the working class, whereas the Conservatives have drawn most of their support from the middle classes. It is argued that today this traditional class conflict, depicted in no better fashion than the Miners' Strike of 1984, has been diluted by growing affluence. In otherwords the working class have become economically better off. They were given the right to buy council houses, to own shares and have, it is argued, become more middle class. The working class today have a lot more to lose in a fierce class struggle and are therefore happy to uphold the system. The huge decline in the traditional industries, such as coal, has coincided with a rise in the size of the non-manual, service industry - the sphere in which the 'middle classes' tend to be employed. In 1964 50% of the workforce were employed in the manual sector, compared to 36% in 1992. These figures coincide with a 15% rise in the non-manual, 'petty bourgeoisie' jobs. Whilst there may be some truth in this 'embourgeoisement' theory, there is also no doubting the fact that it is an exaggerated view. To say that 'we are all middle class' (Blair 1998) is an absurdity. Class conflict may have been subdued but not only because of growing affluence. The capitalists have managed to silence what was once a

High School and Republic High Books Essay

Ms. P Andy Lau Argumentative Essay Censorship is the government is control the media, censorship can used to control or protect people. Some groups feel that books with objectionable material should be censored for many different reasons,such as sexual content,against a religion ,bad language,racism violence. I believe that books shouldn’t be censored because we have the right to read it,and it depends on your own personality. Therefore I intend to prove that the banning of books in schools is completely unjustified. In Fahrenheit 451,There is a old women in the story,she is willing to die to not leave her books. It shows that books are so important to us, and our society that she was willing to die rather than give them up,also the character Guy Montag is curious to read those books,she quitted his job for those books. In the article â€Å"You have insulted me â€Å" by kurt Vonnegut. He is angry because his books got burned by the school board. †That’s because people speak coarsely in real life†this shows people speak bad word in real life,so why can’t books have bad words. Both the article supports my opinion that book shouldn’t be banged from high school. †Two books pulled form Republic school library shelves†this article is two of the three Republic High books singled out in a public complaint last year will now removed from the school curriculum and library. Those three books are â€Å"Speak†by Laurie Halse Anderson. Kurt Vonnegut’s†Slaughterhouse Five†and Sarh Ockler’s â€Å"Twenty Boy Summer†. And they didn’t banned â€Å"Speak†because it didn’t describe sex so much,and it has a good message. Also they just banned those books in classroom,students can still read it in library and independence reading . The second article is talking about parents and school districts have debated what books are appropriate for a school library collection and what books should be banned. ACLU didn’t ban books because they say its against the First Amendment . also they said†You clearly can’t remove a book because you disagree with the ideas in them†,this shows books shouldn’t get banned . We have to see different face of books ,bad way and good way. I believe that books should not be censored because books can send us good messages and our imaginary . It also depends on your own personality,if we have a evil mind ,we could only see the evil thing in the books,if we are normal human,we suppose to able to learn something form books,so I am sure that banning of books in schools is completely unjustified.

Titration Journal

E r J. Biochem. 40,177-185 (1973) u. intra prison boothular Tit rotterion of cyclicalalalalalalalalalalalalalalalalalalalalal adenylic blistering Bound to Receptor Proteins and coefficient of correlation with cyclic- angstrom Levels in the Surviving betray hitch Lien DO KHAC,Simone HARBON Hubert J. CLAUSER and lnstitut de Biochimie, Universit6 de Paris-Sud, Orsay (Received April 9/July 17, 1973) take a commissions on the watch from bottom checks incubated with or with egress theophylline and/or epinephrin suck in been tested for their tot cyclic adenylic pungent content and for their competency to bind exogenicly added cyclic antiophthalmic factor.Less cyclic 3H amp do- nonhing be stunnedflow inthe forces subsequently theophylline and/or epinephrin treatment indicating that the turn out in cyclic type A take aim was accompanied by a n gain in the quantity of cyclic antiophthalmic factor hold back intracellularly to the cyclic angstrom-d ependent protein kinases. maximum cyclic adenosine monophosphate fertilization capacities, as cargonful by total cyclic deoxyadenosine monophosphate re-sentencings, were stock-still akin in each(preno houral) cases. Accu pukee estimations of intracellular cover of cyclic angstrom have been gibe with the level of cyclic group A in the create from raw stuff the reception seems to obey fair strength kinetics, a n obvious intracellular K d for cyclic adenosine monophosphate has been evaluated as 330 nM.The findings be logical approximately(prenomoal) with a real relief in the intracellular spinal column unalterable as comp ard to that metrical in vitro (28 nM) or with the fact that the cyclic al-Qaida in the cell whitethorn non ein truth last(predicate) be available for the kinase protein receptors. They besides no minute of arcate that the system depict whitethorn indicate useful for poring over whatever achievable intracellular control beyon d the pace of cyclic angstrom syn thesis.Regulation of cellular metabolous process by adenosine 3 5-monophosphate (cyclic adenylic virulent) I, its mediation by performer of complex protein kinases 2,3 and the mechanism of the activation of these enzymes 461 have been well attested inside the past years in the eukaryotic cell. Activation has been demonstrated to pass along correspond to equality (1) through a n interaction of cyclic adenylic acid with the regulatory sub unit of measurement (R) of the enzyme, leading to a disassociation of this subunit from the catalytic subunit (C) which is thus activated. RC cyclic international ampere + R cyclic adenosine monophosphate C . (1) + + moreover completely satisfactory correlations in the midst of the levels of intracellular cyclic adenine and its ultimate metabolic effects have been in more cases difficult to welcome. Striking examples for this situation argon to be bring in the results of Craig et al. 7 in rat plo sive, of Stull and Mayer 8 in rabbit gaunt muscle concerning the mandate of phosphorylase activation, of Schaeffer et al. 9 and Miller et al. lo concerning regulation of glycogen metabolism in suprarenal glandectomized rats, and of Harbon and Clauser Ill This fly the coop is dedicated to Professor E. Lederer for his 65 th anniversary. Abbreviations. cyclic international ampere adenosine 3 5-monophosphate. in the rat uterus stimulated by prostaglandin El or E,. I n all these cases, cyclic angstrom unit levels may be elevated without eliciting the expected metabolic responses. dickens hypotheses have been formulated to rationalize these obvious discrepancies, both a come down in the activation of the enzymes mediating cyclic amp action inwardly the cell, or a compartmentalization of the intracellular nucleotide. and so it seems indispensable to measure straight the full stop to which the basic step of the activation sequence (Equation 1) devises the manifest intrac ellular cyclic vitamin A stringencys.This competency be achieved by establishing in sacrosanct cells or create from raw stuffs, correlations in the midst of the levels of intracellular cyclic international ampere chthonic well delimitate physiological conditions, the expiration to which it is adjoin to the limited receptor protein and the goal to which the complex protein kinases are in the restless evince. Satisfactory correlations between cyclic angstrom unit levels and protein kinase activation have been recently establish in various weaves by Corbin et al. I21 and Soderling et al. 13.The chip in work was to investigate if correlations could in like manner be obtained between intracellular cyclic angstrom levels and the measurements of intracellular cyclic adenosine monophosphate indentured to receptor protein (R cyclic axerophthol) in the live on rat stopover incubated with or without theophylline and adrenaline. The results inform demonstrate that E r J. Biochem. 40 (1973) u. 178 Intracellular Titration of cyclic deoxyadenosine monophosphate-Receptor Protein top precise titrations of endogenetic cyclic axerophthol keep back versus cyclic antiophthalmic factor lay in the intact waver may be obtained.An discernible Kd value for the intracehlar cyclic vitamin A cover song is observed which differs wide from the K d of the said(prenominal) stuffing established in vitro 14-161. This rule may prove to be useful for flying fielding the modification of cyclic vitamin A backbone down the stairs conditions where the formation and breakdown of cyclic international ampere does non seem to be stirred. A earlier report of these results has been explicit 17. MATERIALS AND METHODS Cylic group A was obtained from P L Biochemicals Inc. , theophylline and Tris from Merck (Darmstadt), Na,adenosine triphosphate 4 H,O, L- adrenaline bitartrate from Calbiochem.Cellulose ester membrane filters (HA 0. 45 pm, 24 mm) were ac quired from Millipore Corp. entirely reagents apply were fruits of Prolabo (reagent grade). Cyclic 3H angstrom unit was a product of New England Nuclear Inc. , specific occupation 24 Ci/ mmol. Animals were Wistar rats weighing more or less cc to 300 g and fasted 24 h before the experiments. Tissue homogenizations were performed with an revolutionary Turrax homogenizer. The reaction mixture for the fecundation hindrance contained in a final mess of 250 p1, 20 mM TrisHC1 caramel pH 7. 5, 10 mM MgCI,, 6. 7 mM theophylline and cyclic 3H angstrom unit a t various ingresss as indicated.The reaction was initiated by the addition of a n aliquot of occlusion rips equivalent to 70- 150 pg protein. manner B. I n this case, cyclic 3H adenylic acidwas added to the homogenizing mediocre a t saturating concentrations up to 0. 2 p M a t 0 C, centrifugation was carried out immediately and cyclic 3H adenine marge measured right away on the suck out. Cyclic 3H adenine qualify to t he proteins, chthonian either condition, was determined later polar incubation times at 0 C the reaction mixtures were and then diluted to 3 m l with frigidity airplane pilot (20mM TrisHC1, 10mM MgCl,, pH 7. 5) and passed through cellulose ethanoate Millipore filters (0. 45 pm).The filters were process with 25ml of the akin buffer, dehydrated and counted in i 0 ml sparkle fluid, in a Packard Tri-Carb liquid sheen spectrometer. Results were expressed as pmol cyclic international ampere saltation/mg protein the concentration of endogenetic untagged cyclic AMP has been ever so taken into account for the estimation of the specific bodily function of cyclic 3HAMP march in the incubation medium. brooding Procedures The animals were killed by decapitation. The full points were speedily removed, freed from connective tissue, cut to small pieces, pooled and divide into equal parts. 200-250 mg tissue were preincubated in 2. ml Krebs-Ringer-bicarbonate buffer pH 7. 4, bobble phase (95O/, O,, 5O//, CO,) for 30 min a t 37 C, in the absence or heraldic bearing of 10 mM theophylline. broodings were then performed in the absence or social movement of adrenaline (5 pM) for change periods of time. rootage of the Tissue Standard spine Assays for Cyclic A M P Two regularitys have been deviced to extract the tissue and cypher the stuffing of exogenic cyclic 3HAMP to the extracted proteins, both slightly modified from the manner defined by Walton and Garren 15. Method A . The tissue was equalise a t 0 C in 3 ml of one of the following solutions 20 mM TrisHCl buffer pH 7. or 20 mM atomic number 11 acetate pH 7. 5 or 4 mM EDTA pH 6. 0. Theophylline (10 mM) was always register in the various homogenizing media in value to minimize whatever degradation of cyclio AMP by phosphodiesterase present in stoppage extracts. A root centrifugation was carried out for 5 min a t 3000 x g , followed by a joint ohm one a t 50000 x g for 30min. The supe rnatants will be referred to as Tris extract, acetate extract and EDTA extract. Assay for Cyclic-AMP Levels For cyclic AMP assay, the tissue was equal in 3 ml cold 7 trichloroacetic acid and centrifuged for 30 min a t 50000 xg. afterward addition of 0. 1 ml N HC1, the supernatants were extracted 7-8 times with twice their account book of cold ether and evaporated to dryness. make out levels of cyclic AMP in the tissue trichloroacetic acid extract were determined agree to Gilman victimization a protein b a s e and the heatstable inhibitor prepared from rabbit worn muscle 161. I n some instances, cyclic AMP content was alike evaluated in the Tris and acetate extracts. Proteins were precipitated by trichloroacetic acid and extracts processed as draw preceding(prenominal). Proteins in the extracts were determined according to Lowry et al. 18 using bovid serum albumin as a standard. RESULTS AXD DISCUSSION Total Cyclic-AMP Levels in wood pussy Diaphragm. solvents of epinephrine and Theophylline In order to study the cyclic AMP cover song business leader of rat diaphragm proteins and its viable rnodification down the stairs the influence of epinephrine, it seemed unavoidable to test the first effect of the catecholamine, to wit the rise in the tissue cyclic AMP level on a lower floor our observational conditions. Em. J. Biochem. 40 (1973) L. Do Khac, S. Harbon, and H. J. Clauser remand 1. Total cyclic A H P levels in trichloroacetic acid extracts of rat diaphragm.Effect of epinephrine and theophylline R a t diaphragms (200-250 mg) were preincubated for 30 rnin a t 37 C in 2. 5 ml Krebs-Ringer-bicarbonate buffer (0, 95/0-C0, 50/0) in the absence or presence of 10mM theophylline, Incubation was then performed for 5 rnin with or without 5 pM epinephrine. The tissue was then homogenized in 7O/, trichloroacetic acid for cyclic AMP assay as described under Methods. Levels of cyclic AMP were expressed as pmol cyclic AMP/ carbonmg wet tissue and as pmol c yclic AMP/mg soluble protein (as figured by the Lowry procedure in the Tris extract. determine are means f S. E. M. of 5 assorted experiments Incubation condit,ions Total cyclic AMP TheoDhvlline EDineDhrine pmo1/ coke mg pmol/mg wet tissue soluble protein 41 f 8. 0 20. 5 f 4. 7 104 & 1. 1 52 & 0. 47 93 f 4. 5 46 & 2 350 f 21 clxx f 10. 7 179 control board 3. distribution of cyclic 3HAMP- ski fertilisationfractions i n opposite hom. ogenutes from rat diaplwagms incubated with or without epinephrine Preincubation and incubation conditions as described in Table 2. Tissues were homogenized in 3 ml 20mM TrisHCI, p H 7. 5, 4 mM EDTA or 20 mM sodium acetate pH 7. and centrifuged for 5 rnin at 3000 x g, the supernatants were centrifuged once more at 500OOxg for 30 min yielding extract 1 and crack 1. The sediment of the first centrifugation was resuspended in 1. 5 ml of the tally buffer and centrifuged at 500OOxg for 30 min giving extract 2 and blastoff 2. bond activity for cyclic rSHAMP was measured in each fraction as described in the text under method A and was expressed as pmol cyclic AMP bound/l00 mg wet tissue Fraction Cyclic AMP bound in EDTA ethanoate Tris extract extract, extract, 5 yM noepinoepino epinephrine nephrine nephrine + + + + Lhrine pmo1/ speed of light mg wet tissue Extract 1 Extract 2 nip 1 Pellet 2 15. 70 1. 47 0. 76 1. 49 14. 90 1. 54 0. 83 1. 50 15. 30 1. 35 0. 80 1. 10 9. 40 0. 80 0. 44 0. 39 Table 2. Cyclic A M P levels in different extracts obtained from epinephrine- treat and untreated rat diaphragms Preincubation with 10 mM theophylline and incubation conditions in the absence or presence of 5 pM epinephrine as in Table 1. Diaphragms were homogenized in troika different solutions cold 7O/, trichloroacetic acid, Tris-HC1 pH 7. 5 or acetate p H 7. 5 as described under methods.Centrifugation was carried out for 30 rnin at 50000 x g. Soluble Tris extract, acetate extract and their similar sediments were deproteinized by 7 o/o tr ichloroacetic acid before cyclic AMP assay Incubation with epinephrine None 5wM Total cyclic AMP in Trichloroacetic 20 mM acetate acid extract pellet 57 280 20 mM Tris extract pellet 48 218 9. 5 26 extract pellet 45 242 pmo1/ snow mg wet tissue 8. 5 8. 3 As verbalisen in Table 1, epinephrine (5 pM) in the absence of theophylline increases (by a factor of 2. 5) the total cyclic AMP content of rat diaphragm extracted by trichloroacetic acid.Theophylline alone (10 mM) had a stimulating effect, repeat when both compounds were used together, the rise in cyclic AMP levels was 8- t o 9-fold, reaching 350pmol cyclic AMP/100 mg wet tissue. When cyclic AMP was assayed in either acetate or Tris extracts after deproteinization with trichloroacetic acid the determine obtained were identical t o those found when the diaphragms were directly extracted with trichloroacetic acid hence virtually no(prenominal) of the cyclic nucleotide in these extracts was associatcd with membrane-bound fract ions (Table 2). Eur. J. Biochem. 0 (1973) Location of Cyclic AMP-Binding Fractions Table 3 shows the distribution of cyclic AMP cover song activity in various fractions of three rat diaphragm homogenates measured by method A in all cases more than goo/, of this activity was recovered in the 50000 x g supernatant, virtually no cyclic AMP covering occurred in the pellets. Preincubation of the diaphragm with epinephrine did non characterise the percentage distribution of the hot nucleotide between the supernatants and the pellets, hence ensuant experiments have been performed on the soluble extracts.On the former(a) hand, in the case of epinephrine-treated diaphragms, less exogenous labelled cyclic AMP ( somewhat 50-60/0) was bound to the various fractions, indicating a decrease in the rachis capacity of the extract as compared to the untreated diaphragm. Dilution by endogenic cyclic AMP cannot explain the effect of epinephrine, since wages was made for this parameter (see Methods) the phenomenon was consistently ordered and will be get on substantiated and discussed below.The covering fire capacities of the various extracts for cyclic E3HAMP have likewise been verified in the absence of any free endogenetic cyclic AMP after removal of the latter by filtration through Sephadex G 50 (1x 37 cm) columns, previously equilibrated with 20 mM Tris-HC1 buffer, pH 7. 5 a t 4 C. I n these experiments, the detail of which w l not be reported in i l the present manuscript, the effect of epinephrine was still observed, when dorsum was measured on the main protein outpouring emerging with the void volume of the columns. When the department of corrections outlined in the clxxx Intracellular Titration of Cyclic AMP-Receptor Protein Binding Z A 0. 51 / 0 20 40 60 duration ( m i n ) l / f r e e cyclic AMP (nM-) l / f r e e cyclic A M P (nM-) Fig. 1. The time wurse and cyclic-AMP-concentration dependence of cyclic A M P fertilisation in rat-diaphragm extrac ts (method A ) . (A) Diaphragms were incubated for 30 min in the presence of 10 mM theophylline and extracted with Tris HCI buffer (method A). Cyclic AMP fertilization was estimated in the presence of various concentrations of cyclic E3HAMP 20nM ( 0 0 ) 60nM ( ) 0 0 SO& (A-A) 100 nM ( ) -. , a t 0 C. The react,ion mixtures contained in a final volume of 2. 5 ml, 20 mM Tris-HC1 buffer, pH 7. , 10 mM MgCI,, 6. 5 mM theophylline. The reaction was initiated by the addition of 930 pg protein. At the indicated times, aliquots were pipetted, immediately diluted with cold 30 mM Tris-HC1buffer pH 7. 5,lO mM MgCl, and passed on the Millipore filters. Filters were washed with the equivalent buffer, dried and counted. Binding activity is expressed as pmol cyclic AMP bound/mg protein. (B) Data obtained from quasi(prenominal) experiments where covering fire for cyclic AMP was performed a t 0 C, for 1 h, in the presence of cyclic aHIAMP ranging from 12 nM to 110 &I. Double-reciprocal plot, according to Klotz 25 Fig. 2.Cyclic-AMP- soaking up dependence of cyclic A M P bond in rat-diaphragm extracts (method B ) . Binding assays were carried out as described under method B. unhomogeneous concentrations of cyclic 3HAMP ranging from 12nM to 200 nM were added directly to the homogenizing medium for preparing extracts from epinephrine treated (A-A) and untreated (0-0) rat diaphragms. Aliquot,s of the extracts were filtered through Millipore filters, dried and counted. Double-reciprocal plot, according to Klotz 25 present paper were use to these mental images, the results were essentially identical to those obtained with the unfiltered extracts.Specificity. Kinetics and Concentration Dependence of Exogenous Cyclic-AMP Binding in the Extracts Specificity of cyclic AMP natural covering has been assessed by dilution experiments of cyclic 3HAMP (100 nM) with unlabelled nucleotides (adenine, AMP, ATP, cyclic AMP) a t molar concentrations equalling up t o 100 times cyclic 3HAM P concentrations. I n no case, still with unlabelled cyclic AMP, the beat of money of hot material bound to proteins by either method A or B was significantly reduced (the details of these experiments are not reported).When various concentrations of cyclic 3HAMP were added to diaphragm extracts (after homogenization and centrifugation) and the concealment reaction (method A) carried out for different incubation times at 0 C (Fig. I), it appears that saturation was obtained at a concentration of 80 nM for the cyclic nucleotide which essentially coincides with previously published data 14-161 and that screening proportionality was reached a t p H 7. 5 and 0 C after less than 60 min incubation. It has also been verified that with the protein concentration used (70-150 pg in 250 pl) adhere of cyclic AMP was directly proportional to the amount of added proteins.From a reciprocal plot of cyclic AMP bond versus cyclic AMP concentration (inset of Fig. I), an unmixed Kd of 33 nM can be calculated. When similar experiments were performed by adding various concentrations of cyclic 3HAMP into the homogenizing medium (method B) and using diaphragms which have been incubated in the presence and absence of epinephrine, the double-reciprocal plots of Fig. 2 were obtained. The unmistakable Kd set calculated with this method (45 nM) are in the same range as with method A.I n addition this figure shows that epinephrine treatment of the diaphragms does not modify this Kd but decreases the amount of exogenous cyclic AMP which can be bound to the extract proteins. By comparing exogenous cyclic AMP binding value obtained with methods A and B, it appears (Table 4) that when cyclic 3HAMPwas added to the Eur. J. Biochem. 40 (1973) L. Do Khac, S. Harbon, and H. J. Clauser Table 4. Comparison of exogenous binding of cyclic SIIAMP to diaphragm extracts by method A or method B. Rat diaphragms were incubated with theophylline in the absence or presancc of 5 p M epinephrine.Ex tracts in Tris-HC1 were prepared as described under method A for subsequent binding of cyclic 3HAMP (100 nM), 1 h, a t 0 C. A second serial of extracts were prepared in the same way but in the prescnce of 100 nM cyclic 3HABIP in the homogenizing medium (method R) binding of cyclic 3HAMP was measured in a n aliquot immediately after centrifugation at 0 C ( virtually 1 h after the end of incubation). Values are expressed as pmol bound cyclic AMP/mg protein. Numerals deep down brackets indicate number of experiments Method Cvclic A P bound with M 5 pM epinephrine no epinephrine pmol/mg protein 4 f 0. 22 (9) 4. 80 5 0. 2 (5) 181 6 t e . ? 4 Q Q E A B 2 f 0. 13 (9) 3 f 0. 19 (5) 0 I I I 30 60 90 * Time (rnin) homogenization medium (extract B) prouder binding determine were obtained both with epinephrine-treated and untreated diaphragms, than with method A. This demonstrates that some additional binding of endogenetic cyclic AMP occurred during the homogenization and fractionation p rocedures, which tends to decrease the amount of un set-aside(p) binding sites available for exogenous cyclic 3HAMP. so method B has been currently used to measure exogenous cyclic AMP binding, since the values obtained with this method seem to think over intracellular conditions more accurately.Fig. 3. Time human body of cyclic 3HAMP binding in extracts from rat diaphragms incubated in the absence or presence of theophylline orland epinephrine. Half rat diaphragms were preincubated in the absence (m, A ) or in the presence ( 0 , 0 ) of 10 m31 theophylline for 30 min at 37 C. Epinephrine (5 pM) was added ( A , 0 )and incubation continued for 5min. Tissue was homogenized in 1. 5 ml Tris-HC1 buffer containing 200 nnf cyclic 3HAMP and centrifuged at 5000xg for 10 min at 0 C.Binding of cyclic 3HAMP was measured in aliquots of the supernatant at the times indicated, through Millipore filtration, t = 0 corresponds to the onset of the extraction. Results are expressed as pmol cyclic AMP bound/ mg protein (without correction for cyclic AMP deepen) Effect of Theophylline and Epinephrine Treatment on the Binding of Exogenous Cyclic 3HAMP by Diaphragm Extracts Fig. 3 shows the results of a common experiment in which diaphragms have been incubated in the absence or presence of theophylline and epinephrine. Homogenization has been performed according to method B, the centrifugation time of the homogenate kept to a inimum (10 min), and the binding capacity for cyclic 3HAMP determined a t different times. As may have been expected, this cyclic 3HAMP binding (which measures the residual binding capacities of the extracts) was, in the course of the whole titration period, in return related t o the amount of endogenetic cyclic AMP present in the relevant extracts (see Table 1). Hence the agents which increase the intracellular cyclic AMP level appear to decrease the amount of binding sites available for exogenous cyclic 3HAMP, probably through an increase of endogenic cy clic AMP binding to the receptors.I n order to titrate endogenous binding of cyclic AMP accurately, experiments were designed to estiEm. J. Biochem. 40 (1973) mate the total binding capacities of the extracts through complete alternate of endogenously bound cyclic AMP with cyclic 3HAMP, and also to estimate the actual amount of exchange occurring in the extracts between endogenous bound unlabelled cyclic AMP and exogenous cyclic 3HAMP during the titration period. A precise knowledge of these dickens parameters is required for the determination of the binding sites occupied by endogenous cyclic AMP at the moment where the tissues are homogenized.Cyclic-AM P Exchange and Determination of Maximal Binding Capacities Total cyclic AMP exchange has been measured under the conditions defined by Wilchek et al. 19 for parotid gland and skeletal muscle extracts from both treated and untreated diaphragms were f i s t incubated at 0 C with cyclic 3HAMP (100 nM) under binding conditions of met hod A and then allowed t o exchange with 1 pM unlabelled cyclic AMP at 20 C in the presence of 100p. M ATP and 10mM MgCl,. Fig. 4 shows that almost complete exchange of the bound labelled nucleotide occurred at heart 30 min, 182Intracellular Titration of Cyclic AMP-Receptor Protein Binding 0 10 20 30 40 50 60 Time (min) 70 80 90 Fig. 4. Exchange of bound cyclic SHIAMP. Extracts were prepared from epinephrine-treated ( + o ) and untreated (0-0) rat diaphragms. Binding of cyclic 3HAMP was carried out a t 0 C in a volume of 2. 5 ml with 500 pg proteins, and 100 nM cyclic r3HAMP in Tris-HC1 buffer, MgCl, and theophylline a t the concentrations described for the standard binding assay. After 1-h incubation, 1 pM unlabelled cyclic AMP and 100 pM ATP were added and the mixture allowed to stand at 20C.At the different times indicated in the figure, aliquots corresponding t o 50 pg protein were pipetted, promptly diluted with 20 mM Tris-HC1 buffer, 2. 5 mM MgC1, p H 7 5 and filtered throug h Millipore filters. The filters . were washed with the same buffer, dried and counted. Results are expressed as pmol/mg protein 0 30 60 90 120 Time (rnin) 180 240 Total binding capacities of the proteins could thus be measured by incubating the extracts first with 100 nM unlabelled cyclic AMP a t 0 C and carrying on the exchange reaction in the presence of 1 pM cyclic I13HAMP at 20 C for 1-2 h the values obtained fairishd 8. -9. 5 pmol cyclic 3HAMP/mg soluble protein, both with epinephrine-treated and untreated diaphragms. These results were corroborate by direct assay of bound cyclic AMP the extracts have been richly modify with unlabelled 1pM cyclic AMP and filtered as described. After washing the Millipore filters, bound cyclic AMP was extracted by cold 7 O/, trichloroacetic acid and the cyclic nucleotide was directly assayed according to Gilman 16. The average value was 9. 8 f 0. 4 pmol cyclic AMP bound per mg protein, which is of the same order of magnitude as the amoun t of bound cyclic 3HAMP calculated above.Previously published data are in close agreement with these values. Walton and GarFen 15 reported supreme binding capacities of 9. 8 pmol/mg protein for adrenal extracts, whereas Gilman l6 found a total binding of 12pmol/mg protein in muscle extracts. The values for maximum cyclic AMP binding are truly low as compared t o the total endogenous cyclic AMP present in the extract (46 pmol/mg protein with the theophylline-treated diaphragm and clxx pmol/mg protein with the epinephrine theophylline-treated diaphragm).It must be added that the binding proteins, saturated with cyclic AMP or not, were almost completely bear on the Millipore filters, and that endogenous cyclic AMP, not Fig. 5. T i m e course of cyclic A M P exchange under binding (0 C) and exchange (20 C conditions. Extracts were prepared from epinephrine treated (0,A ) and untreated ( 0 , A) r a t diaphragms. Binding of cyclic AMP was performed as described in Fig. 2 in the presen ce of 100 nM cyclic AMP for 60 min at 0 C. A t the end of the binding reaction 1 pM cyclic 3HAMP was added t. the different extracts, in the absence (A, A ) or presence ( 0 , 0 ) of l00p. M ATP. The reaction mixtures were maintained a t 0 C for 2 h and then at 20 C (arrow) for 2 more hours. At the different times indicated on the figure, aliquots corresponding t o 70 pg protein were pipetted and treated as in Fig. 4. Results are expressed as cyclic rHAMP bound in pmol/mg protein. bound to these fractions, was quantitatively recovered in the Millipore filtrates after trichloroacetic acid extraction. The consummation t o which this free cyclic AMP may or not be bound to other proteins is presently not known.Cyclic-AMP Exchange under Binding Conditions The extent of cyclic AMP exchange under binding conditions (0 C, 1 h, 100 nM cyclic AMP) must be controlled if corrections for simultaneous exchange have to be applied t o binding data extracts of rat diaphragms treated with theophyllin e and theophylline epinephrine were first saturated with 1 O O n M unlabelled cyclic AMP (binding conditions) and then exchanged with 1 pM cyclic 3HAMP but a t 0 C. After 2 h, the temperature was brocaded to 20 C and completion ofthe exchange measured after 1-2 h further incubation.Fig. 5 shows that a t 0 C, within 1h incubation time, which are the conditions described above for the binding assay, about 200/, of total sites were exchangeable. Under these conditions, ATP and Mg ions slightly increase the exchange velocity. I n addition, this figure confirms that a t 20 C total exchange capacities were identical for epinephrine-treated and untreated diaphragms hence initial + + Em. J. Biochem. 40 (1973) L. Do Khac, S. Harbon, and H. J. Clauser 183 Table 5. relationship between intracellular cyclic A M P levels and cyclic AM P binding in extracts from diaphragm incubated under various conditions Diaphragms were incubated with or without 10 mM theophylline for 30 min at 37 C, 5 pM epin ephrine was added where indicated and incubation continued for varying times. From each incubation, half a diaphragm was extracted by trichloroacetic acid for cyclic AMP estimation. The other half was homogenized with Tris-HC1buffer lOOnM cyclic 3HAMP(method B) for exogenous cyclic AMP binding after 1 h a t 0 C maximal binding capacities were determined in the same extracts a t 20 C in the presence of 1 pM cyclic 3HAMP under conditions described for cyclic A P exchange.R. esults are expressed as pmol cyclic AMP/mg M protein. endogenic binding values were calculated as the difference between maximal binding capacities ( A )and exogenousbinding ( B ) and corrected for the 200/, exchange + Incubation conditions Theophylline 10 mM Epinephrine 5t*M Time Cyclic AMP Total level Maximal binding Exogenous capacity binding (a) (b) Endogenous binding (a-b) corrected min pmol/mg protein + + + + + + 0 2 10 30 5 5 20. 5 52 43 38 46 clxx f 4. 7 & 0. 47 f2 f 10. 7 9. 6 f 0. 9 9. 4 f 0. 1 9. 2 0 9. 40 8. 9 5 0. 73 8. 9 & 0. 85 5. 35 f0. 40 4. 50 f 0. 133 4. 40 4. 70 4. 46 f 0. 20 2. 7 f0. 224 5. 31 6. 13 6 5. 5 5. 53 7. 77 differences in residual binding capacities reflect variations in the degree of saturation of the receptor proteins by endogenous cyclic AMP, rather than modifications of their maximal binding capacity. 1 Titration o Endogenous Cyclic-AMP Binding in Rat f Diaphragm. set up of Theophylline and Epinephrine Since total binding capacities of the receptor proteins in the extracts and the amount of exogenous cyclic 3HAMP bound by these extracts after homogenization may be estimated, it appears thinkable to calculate endogenous cyclic AMP bound in the intact harmoniums, correcting for a 2001, exchange during the titration period.Table 5 summarizes the results of a series of experiments where diaphragms have been incubated under conditions which modify endogenous levels of cyclic AMP in every case, half of the diaphragm was extracted with cold trichloroaceti c acid (see Methods) for the assay of intracellular cyclic AMP levels the second half was extracted according to method B for the estimation of exogenous cyclic 3HAMP binding and of total cyclic AMP binding capacities. The endogenous cyclic AMP bound was calculated from the latter observational data.This table definitely establishes that the average values obtained for the intracellular binding of endogenous cyclic AMP in the intact organ seem to correlate with its cyclic AMP levels. A reciprocal plot of intracellular binding versus intracellular cyclic AMP concentrations (Fig. 6) shows that this correlation fits unproblematic saturation kinetics very accurately. I n the unstimulated diaphragm (no theophylline nor epinephrine added to the incubation medium) about 50/, of the available binding sites are occupied by endogenous cyclic AMP this Eur. J. Biochem. 40 (1973) -0. 002 I 0. 002 l/Free cyclic AMP (nM-) 0 0. 004 . Fig. 6. Reciprocal plot of intracellular cyclic A M P levels an d cyclic A M P binding in rat-diaphragm extracts. Data arc obtained from experiments performed as described in Table 5 and replotted according t o the Klotz equation. The interpose on the y axis yields a n estimate of the number of binding sites and the x intercept provides a n estimation of the intracellular seeming dissociation constant. Statistical analysis of the data were performed according to Cleland 26 using a Wang electronic computing machine alue increases to almost goo/,, when the diaphragms have been fully stimulated with both theophylline and epinephrine. Various treatments with one of the agonists alone cause endogenous bindings ranging between these 2 extreme values. The spare Kd value for intracellular binding according to this plot was estimated to 330 nM f 50, as compared to the apparent Kd (33-45 nM) when binding was assayed in the extracts (Fig. l and 2). Hence a difference of about one order of magnitude appears to obtain between the Kd values calculated wi thin the cell and the 84 Intracellular Titration of Cyclic AMP-Receptor Protein Binding same constant measured with diaphragm homogenates. The double-reciprocal plot may also be used to calculate the intracellular maximal binding capacities, from its intercept with the set up axis. A value of 8. 9 pmol/mg protein was found which coincides with the values measured in the extracts by total cyclic 3HAMP exchange. This dissonance between the intracellular Kd and the Kd measured in vitro in a variety of tissue extracts including diaphragm may a t first sight seem surprising.It has however repeatedly been pointed out that cyclic AMP concentration even in the unstimulated cell was far in excess of the concentration which should result in almost maximal stimulation of protein kinases and compartmentalization of the nucleotide within the cell has usually been postulated to explain this contradiction in terms 8,9,20. The present work shows that despite these high intracellular concentration s of cyclic AMP, protein kinases could indeed not be fully activated, since under the same conditions, the receptor proteins appear not to be fully saturated with cyclic AMP. think RemarksAs might have been expected from Equation (1) (if this reaction truly reflects intracellular conditions) a rise in cyclic AMP should be mateed by an increase in the amount of cyclic AMP bound to receptor protein in the cell. The results reported show this indeed to be the case in the isolated rat diaphragm when this tissue is stimulated by various agents which increase the level of cyclic AMP the amount of protein receptors endogenously saturated by cyclic AMP (R cyclic AMP) rises, as indicated in our experiments by a decrease in their ability to bind exogenously added cyclic 3HAMP after tissue extraction.Maximal binding capacities for cyclic AMP do not seem to be affected under any circumstance. A parallel approach t o the study of this problem has been undertaken by Corbin et al. 12 and Soderli ng et al. 13 who investigated in fatty tissue under various stimulatory conditions, the state of activation of the catalytic subunit (C) by assaying the cyclic AMP dependence of the protein kinase in tissues extracts. These authors demonstrated that under well-defined xperimental conditions, in that location was a quantitative relationship between the intracellular level of cyclic AMP and the amount of the active C unit which could be separated from the complex protein kinase RC. even so in their experiments high concentrations of NaCl had to be added to the extracts, since in its absence R and C tended to reassociate almost immediately, indicating that cyclic AMP is no monthlong bound to its receptor protein (R). The situation in various other tissue xtracts has been found to be analogous, except with skeletal muscle, where preliminary results obtained by the authors led them to suggest that the protein kinase subunits do not readily reassociate. This seems also to be the case for the diaphragm, since under the conditions of the present work, it has been possible to titrate for R * cyclic AMP in the in the buff extracts even in the absence of high salt concentrations acccurate estimations of intracelM a r binding of cyclic AMP have been obtained and correlated with the absolute amounts of the nucleotide present in the stimulated and unstimulated cell.The binding seems t o obey simple saturation kinetics but the apparent Kd of this binding is about10 times higher as compared with the crude extracts. These results may be explained by cyclic AMP compartmentalization within the cell in this case, however, the simple saturation kinetics would indicate that the various pools of the cyclic nucleotide attain equilibrium very rapidly.Or else, if cyclic AMP within the cell is not compartmentalized, and if the reaction described by Equation (1) may be applied, without any modification, to intracellular equilibria, a decrease in the apparent Kd could be merely a c onsequence of the dilution (about 10-fold) of the protein components during extraction of the tissue, while cyclic AMP concentrations are maintained by the addition of exogenous cyclic 3HAMP.However these two hypotheses are certainly oversimplified, since they do not take into account factors like the intracellular concentration of the heat-stable kinase inhibitor 21,22, ATP or Mg2+ 19,23, which are known to affect cyclic AMP binding either in crude extracts or with purified protein kinase preparations. It seems impossible to decide at present which of these interpretations is most likely to reflect true intracellular conditions. It is noteworthy that the apparent Kd estimated is close to the intracehlar cyclic AMP concentration of the nstimulated tissue, a fact which should account for maximal sensitivity of the regulatory mechanisms under physiological conditions. Hormonal controls at the level of cyclic AMP-receptor protein interaction have hitherto never been described the data reported above provide a suitable means for investigating such problems. The authors are very much indebted to Mrs Ginette Delarbre for her excellent skillful assistance and to Mrs Marie-ThBrBse Crosnier for preparing the manuscript. The present work has been performed give thanks to two official grants of the C. N. R. S. Paris, France ERA No 33 and ATP No 429. 914), to a grant obtained from the D. G. R. S. T. (No 72. 7. 0135), to a generous contribution of the Fondation pour la Recherche Mf? dicale Franpise and to a participation of the CEA (Saclay, France) in the purchase of radioactive compounds. The work has been performed as a partial fulfillment of a thesis (Doctorat Bs-Sciences) submitted by L. D. -K. Eur. J. Biochem. 40 (1973) L. Do Khac, S. Harbon, and H. J. Clauser REFERENCES 1. Robison, G. A. , Butcher, R. W. & Sutherland, E. W. (1968) Ann. Rev. Biochem. 37, 149-174. 2.Walsh, D. A. , Perkins, J. P. & Krebs, E. G. (1968) J. Biol. Chem. 243, 3763-3765. 3. Kuo, J. F. & Gr eengard, P. (1969) Proc. Nut. Acad. Xci. U . S. A. 64, 1349-1355. 4. Reimann, E. M. , Brostrom, C. O. , Corbin, J. D. , King, C. A. & Krebs, E. G. (1971) Biochem. Biophys. Res. Commun. 42, 187-194. 5. Tao, M, Salas, M. L. & Lipmann, F. (1970) Proc. Nut. Acad. Sci. U . S. A. 67, 408-414. 6. Gill, G. N. & Garren, L. D. (1970)Biochem. Biophys. Res. Commun. 39, 335-343. 7. Craig, J. W. , Rall, T. W. & Larner, J. 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