- Email: [email protected]
Benefits of ethological analysis of behaviour
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the affinity of the receptor for the G protein rather than the proportion of the receptors distributed between the R and R~ states. The change in the affinity of the receptor would then bring about a change in the rate of receptor - G protein interaction. Of the two models, I prefer to view a G protein-coupled receptor as a 'flexible' protein that can take up a number of different conformations depending upon the 'shape' of the
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ligand that is bound and, that these different conformations have different affinities for G proteins, which can be greater or less than that of the receptor with no ligand bound. However, the choice of model comes down to how the function of the receptor in the membrane of a cell is viewed, which brings us back to the question 'What are the molecular mechanisms of agonism at G proteincoupled receptors?'
Benefits of ethological analysis of behaviour Cost:benefit ratio of an ethological analysis of behaviour in the elevated maze model of anxiety We read with interest the recent article in TiPS by Dawson and Tricklebank on use of the elevated plus maze in the search for novel anxiolytic agents 1. It was concluded that the elevated plus maze test 'is prone to false positives and false negatives, particularly when the drug in question alters locomotor activity' and 'on this basis, it is difficult to justify its use as anything other than a preliminary screen as a prelude to testingin more robust animal models of anxiety'. Furthermore, while it is acknowledged by Dawson and Tricklebank that the recent use of risk assessment analysis and a 'zero' rather than a 'plus' design have helped to refine the maze model, itis asserted that these 'potential improvements have yet to be combined with the more sensitive visual tracking system to determine whether they have real advantages over the current procedure'. In addition, Dawson and Tricklebank suggest that 'whatever the outcome of such studies, scoring risk assessment behaviours is time consuming and tedious and detracts from the simplicity of the test, which is considered to be one of the main advantages of the elevated plus maze'. Considering the last point first, the majority of authors to report elevated plus maze studies observe and record number and duration of open- and
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TiPS -
August 1995 (Vol. 16)
closed-arm entries as their principal dependent measure. To incorporate an ethological analysis using risk assessment we find it necessary to use an additional two keys on a computer keypad to score head dips and stretch attend postures2. This analysis is no more time consuming and tedious than the traditional observation and recording methods used in most laboratories. While it is acknowledged that the observational approach is more time consuming and tedious than the sensitive video-tracking approach advocated by Dawson and Tricklebank, there is little advantage in using such an automated method to measure rodent behaviour if it is impossible to decide what the results mean3. Interestingly, in the concluding sentence of their article, Dawson and Tricklebank acknowledge that 'it is only by incorporating more detailed behavioural analysis that its (the elevated plus maze) validity can be properly explored'. Problems of data interpretation caused by reliance on automatic measurement of rodent locomotion such as photocells or video-tracking are common and are well documented3. It is our view that the small additional cost of observing rodent behaviour on an elevated maze is outweighed by the benefit afforded by overcoming the potential confound
B. 6ardner Biological Laboratory, University of Kent, Canterbury, UK CT2 7NJ. References
1 Left,P. (1995) Trends Pharmacol. Sci. 16, 89-97 2 De Lean,A., Stadel,J. M. and Lefkowitz, R.J. (1980)J. Biol. Chem. 255,7108-7117 3 Costa,T.,Ogino,Y.,Munson,P.J.,Onaran, H. O. and Rodbard,D. (1992)Mol. Pharmacol. 41, 549-560 4 Bourne, H. R., Sanders, D. A. and McCormick,F. (1990)Nature 348,125-132
of alterations in locomotor activity. Thus, our studies using ethological analysis of rodent behaviour on an elevated zero maze have demonstrated a dissociation between drug effects on locomotion and anxiety2,4. For example, the 5-HTac receptor agonist m-chlorophenylpiperazine (mCPP) decreased entries into and time spent in open quadrants of the elevated zero maze, an effect which, in previous studies using the elevated plus maze, was confounded by a decrease in locomotion. However, our observation that mCPP increased the incidence of stretch attend postures at closed-open quadrant boundaries indicated that the anxiogenic-like profile of the drug could not be explained by locomotor suppression2. Turning to the profile of stimulant drugs, interpretation of the effects of amphetamine and cocaine on anxiety in humans is often confounded by variations in test environment, rating scales and individual subjects ~7. These data suggest that interpretation of the anxiolytic-like profile of amphetamine in rats on an elevated plus or zero maze as a false positive is over-simplistic. Finally, it is our view that it is naive and potentially misleading to rely on any single animal model of anxiety to predict clinical efficacy in humans. In our laboratory we routinely use a battery of rodent anxiety models incorporating both ethological and conflict procedures in an attempt to predict anxiolytic efficacy in patients. Our data suggest that the elevated zero maze used in conjunction with
L ethological analysis continues to justify its place in such a battery of tests. C. T. Dourish, S. S. Grewal, J. K. Shepherd, K. J. Stanhope, D. J. Bill and A. Fletcher Department of Neuropharmacology, Wyeth Research (UK), Taplow, Maidenhead, UK SL60PH.
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References
1 Dawson, G. R. and Tricklebank, M. D. (1995)Trends Pharrnacol. Sci.16, 33-36 2 Shepherd,J. K., Grewal,S. S., Fletcher,A., Bill, D. J. and Dourish, C. T. (1944) Psychopharmacology 116,56-64 3 Dourish,C.T.(1987)in ExperimentalPsychopharmacology(Greenshaw,A.J.andDourish, C. T.,eds),pp. 153-21l, HumanaPress
Dawson and Tricklebank reply Ethological analysis may not be the answer to the problems of the elevated plus maze It is encouraging that other laboratories involved in assessing the anxiolytic potential of compounds share our concerns regarding the use of the elevated plus maze, and realize that it is necessary to use it only in conjunction with other tests. Dourish and colleagues also suggest that by using ethological measures of behaviour some of the limitations of the elevated plus maze can be overcome. However, in the same paper in which the authors reported the anxiogenic effects of mCPP (Ref. 1), they also reported that although the standard anxiolytic, chlordiazepoxide, dosedependently increased the time on the open arm, it did not dose-dependently affect the ethological measures of head dips and stretch attend postures. Moreover, they also reported that the putative anxiolytic, ondansetron (a 5-HT 3receptor antagonist), dose-dependently affects time on the open arms and stretch attend postures, but not head dips. Taken together these results suggest that it may not be prudent to rely on one or two ethological measures of behaviour and that the approach
proposed by Dourish and colleagues may require further refinement. Nevertheless, it is heartening that these authors have replicated our finding that the psychostimulant, amphetamine, and in addition cocaine, have anxiolytic-like effects in the elevated maze 2. Moreover, we await with interest the publication of these results as they may show whether an ethological analysis in combination with a zero maze can distinguish between stimulant and anxiolytic-like effects of psychoactive compounds. It is, of course, apparent that the interpretation of anxiety states in animals relies heavily on knowledge of the subjective effects of various standard compounds in humans. Cocaine and amphetamine are clearly stimulants in both species but, as Dourish and colleagues point out, the effects of these compounds on anxiety levels in the clinic is difficult to discern, although neither would be used as a first-line medicine for anxious patients. Similarly, mCPP should not be viewed solely as an anxiogenic agent, since while it induces the symptoms of panic in
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4 Grewal, S. S., Bill, D. ]., Fletcher, A. and Dourish, C. T. Br. ]. Pharmacol. (in press) 5 Rickels, K., Hesbacher, P. and Fisher, E. (1976) PsychoI. Med. 6, 62,3-630 6 Johanson, C. E. and De Wit, H. (1989) NIDA Res. Monogr. Serv. 92, 171-210 7 Mathews, R. J. and Wilson, W. H. (1989) Neuropsychobiolow 21, 117-123
patients suffering panic attacks, it also induces migraine in migraineurs. Which, if any, of these effects is responsible for the changes in behaviour in the plus or zero maze is not an easy question to answer, either with or without the benefits of ethological analysis. Finally, as Dourish and co-workers point out, problems of data interpretation caused by reliance on automatic measurement are well documented. It was with this in mind that we put considerable effort into ensuring that the correlation between the data determined by the visual tracking system and that scored by a human observer was > 0.95 (Ref. 13), thus ensuring its validity before we began the series of experiments reported in our recent article in TIPS. G. R. Dawson and M. D. Tricklebank Merck Sharp & Dohme Research Laboratories, Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, UK CM202QR
References
1 Shepherd,J. K., Grewal,S. S., Fletcher,A., Bill,D. J. and Dourish,C. T. (I994)Ps,vchopharmacology 116,56-64 2 Grewal, S. S., Bill, D. J,, Fletcher, A. and Dourish, C. T. Br. I. Pharmacol. (in press) 3 Dawson,G. R., Crawford,S. P., Stanhope, K.J., Iversen,S. D. and Tricklebank,M D. (1994) PsychopharmacolG,y 113,570-572
Debate The Debate section is an occasional feature that allows discussion of highly topical issues in all areas of pharmacology.
Suggestions for topics for the Debate section are welcomed and may be addressed directly to the Editor, either in writing or by telephone. Priority is given to topics of broad pharmacological interest.
TiPS - August 1095 (Vol. 16)
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the affinity of the receptor for the G protein rather than the proportion of the receptors distributed between the R and R~ states. The change in the affinity of the receptor would then bring about a change in the rate of receptor - G protein interaction. Of the two models, I prefer to view a G protein-coupled receptor as a 'flexible' protein that can take up a number of different conformations depending upon the 'shape' of the
R
S
ligand that is bound and, that these different conformations have different affinities for G proteins, which can be greater or less than that of the receptor with no ligand bound. However, the choice of model comes down to how the function of the receptor in the membrane of a cell is viewed, which brings us back to the question 'What are the molecular mechanisms of agonism at G proteincoupled receptors?'
Benefits of ethological analysis of behaviour Cost:benefit ratio of an ethological analysis of behaviour in the elevated maze model of anxiety We read with interest the recent article in TiPS by Dawson and Tricklebank on use of the elevated plus maze in the search for novel anxiolytic agents 1. It was concluded that the elevated plus maze test 'is prone to false positives and false negatives, particularly when the drug in question alters locomotor activity' and 'on this basis, it is difficult to justify its use as anything other than a preliminary screen as a prelude to testingin more robust animal models of anxiety'. Furthermore, while it is acknowledged by Dawson and Tricklebank that the recent use of risk assessment analysis and a 'zero' rather than a 'plus' design have helped to refine the maze model, itis asserted that these 'potential improvements have yet to be combined with the more sensitive visual tracking system to determine whether they have real advantages over the current procedure'. In addition, Dawson and Tricklebank suggest that 'whatever the outcome of such studies, scoring risk assessment behaviours is time consuming and tedious and detracts from the simplicity of the test, which is considered to be one of the main advantages of the elevated plus maze'. Considering the last point first, the majority of authors to report elevated plus maze studies observe and record number and duration of open- and
2
6 0
TiPS -
August 1995 (Vol. 16)
closed-arm entries as their principal dependent measure. To incorporate an ethological analysis using risk assessment we find it necessary to use an additional two keys on a computer keypad to score head dips and stretch attend postures2. This analysis is no more time consuming and tedious than the traditional observation and recording methods used in most laboratories. While it is acknowledged that the observational approach is more time consuming and tedious than the sensitive video-tracking approach advocated by Dawson and Tricklebank, there is little advantage in using such an automated method to measure rodent behaviour if it is impossible to decide what the results mean3. Interestingly, in the concluding sentence of their article, Dawson and Tricklebank acknowledge that 'it is only by incorporating more detailed behavioural analysis that its (the elevated plus maze) validity can be properly explored'. Problems of data interpretation caused by reliance on automatic measurement of rodent locomotion such as photocells or video-tracking are common and are well documented3. It is our view that the small additional cost of observing rodent behaviour on an elevated maze is outweighed by the benefit afforded by overcoming the potential confound
B. 6ardner Biological Laboratory, University of Kent, Canterbury, UK CT2 7NJ. References
1 Left,P. (1995) Trends Pharmacol. Sci. 16, 89-97 2 De Lean,A., Stadel,J. M. and Lefkowitz, R.J. (1980)J. Biol. Chem. 255,7108-7117 3 Costa,T.,Ogino,Y.,Munson,P.J.,Onaran, H. O. and Rodbard,D. (1992)Mol. Pharmacol. 41, 549-560 4 Bourne, H. R., Sanders, D. A. and McCormick,F. (1990)Nature 348,125-132
of alterations in locomotor activity. Thus, our studies using ethological analysis of rodent behaviour on an elevated zero maze have demonstrated a dissociation between drug effects on locomotion and anxiety2,4. For example, the 5-HTac receptor agonist m-chlorophenylpiperazine (mCPP) decreased entries into and time spent in open quadrants of the elevated zero maze, an effect which, in previous studies using the elevated plus maze, was confounded by a decrease in locomotion. However, our observation that mCPP increased the incidence of stretch attend postures at closed-open quadrant boundaries indicated that the anxiogenic-like profile of the drug could not be explained by locomotor suppression2. Turning to the profile of stimulant drugs, interpretation of the effects of amphetamine and cocaine on anxiety in humans is often confounded by variations in test environment, rating scales and individual subjects ~7. These data suggest that interpretation of the anxiolytic-like profile of amphetamine in rats on an elevated plus or zero maze as a false positive is over-simplistic. Finally, it is our view that it is naive and potentially misleading to rely on any single animal model of anxiety to predict clinical efficacy in humans. In our laboratory we routinely use a battery of rodent anxiety models incorporating both ethological and conflict procedures in an attempt to predict anxiolytic efficacy in patients. Our data suggest that the elevated zero maze used in conjunction with
L ethological analysis continues to justify its place in such a battery of tests. C. T. Dourish, S. S. Grewal, J. K. Shepherd, K. J. Stanhope, D. J. Bill and A. Fletcher Department of Neuropharmacology, Wyeth Research (UK), Taplow, Maidenhead, UK SL60PH.
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References
1 Dawson, G. R. and Tricklebank, M. D. (1995)Trends Pharrnacol. Sci.16, 33-36 2 Shepherd,J. K., Grewal,S. S., Fletcher,A., Bill, D. J. and Dourish, C. T. (1944) Psychopharmacology 116,56-64 3 Dourish,C.T.(1987)in ExperimentalPsychopharmacology(Greenshaw,A.J.andDourish, C. T.,eds),pp. 153-21l, HumanaPress
Dawson and Tricklebank reply Ethological analysis may not be the answer to the problems of the elevated plus maze It is encouraging that other laboratories involved in assessing the anxiolytic potential of compounds share our concerns regarding the use of the elevated plus maze, and realize that it is necessary to use it only in conjunction with other tests. Dourish and colleagues also suggest that by using ethological measures of behaviour some of the limitations of the elevated plus maze can be overcome. However, in the same paper in which the authors reported the anxiogenic effects of mCPP (Ref. 1), they also reported that although the standard anxiolytic, chlordiazepoxide, dosedependently increased the time on the open arm, it did not dose-dependently affect the ethological measures of head dips and stretch attend postures. Moreover, they also reported that the putative anxiolytic, ondansetron (a 5-HT 3receptor antagonist), dose-dependently affects time on the open arms and stretch attend postures, but not head dips. Taken together these results suggest that it may not be prudent to rely on one or two ethological measures of behaviour and that the approach
proposed by Dourish and colleagues may require further refinement. Nevertheless, it is heartening that these authors have replicated our finding that the psychostimulant, amphetamine, and in addition cocaine, have anxiolytic-like effects in the elevated maze 2. Moreover, we await with interest the publication of these results as they may show whether an ethological analysis in combination with a zero maze can distinguish between stimulant and anxiolytic-like effects of psychoactive compounds. It is, of course, apparent that the interpretation of anxiety states in animals relies heavily on knowledge of the subjective effects of various standard compounds in humans. Cocaine and amphetamine are clearly stimulants in both species but, as Dourish and colleagues point out, the effects of these compounds on anxiety levels in the clinic is difficult to discern, although neither would be used as a first-line medicine for anxious patients. Similarly, mCPP should not be viewed solely as an anxiogenic agent, since while it induces the symptoms of panic in
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4 Grewal, S. S., Bill, D. ]., Fletcher, A. and Dourish, C. T. Br. ]. Pharmacol. (in press) 5 Rickels, K., Hesbacher, P. and Fisher, E. (1976) PsychoI. Med. 6, 62,3-630 6 Johanson, C. E. and De Wit, H. (1989) NIDA Res. Monogr. Serv. 92, 171-210 7 Mathews, R. J. and Wilson, W. H. (1989) Neuropsychobiolow 21, 117-123
patients suffering panic attacks, it also induces migraine in migraineurs. Which, if any, of these effects is responsible for the changes in behaviour in the plus or zero maze is not an easy question to answer, either with or without the benefits of ethological analysis. Finally, as Dourish and co-workers point out, problems of data interpretation caused by reliance on automatic measurement are well documented. It was with this in mind that we put considerable effort into ensuring that the correlation between the data determined by the visual tracking system and that scored by a human observer was > 0.95 (Ref. 13), thus ensuring its validity before we began the series of experiments reported in our recent article in TIPS. G. R. Dawson and M. D. Tricklebank Merck Sharp & Dohme Research Laboratories, Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, UK CM202QR
References
1 Shepherd,J. K., Grewal,S. S., Fletcher,A., Bill,D. J. and Dourish,C. T. (I994)Ps,vchopharmacology 116,56-64 2 Grewal, S. S., Bill, D. J,, Fletcher, A. and Dourish, C. T. Br. I. Pharmacol. (in press) 3 Dawson,G. R., Crawford,S. P., Stanhope, K.J., Iversen,S. D. and Tricklebank,M D. (1994) PsychopharmacolG,y 113,570-572
Debate The Debate section is an occasional feature that allows discussion of highly topical issues in all areas of pharmacology.
Suggestions for topics for the Debate section are welcomed and may be addressed directly to the Editor, either in writing or by telephone. Priority is given to topics of broad pharmacological interest.
TiPS - August 1095 (Vol. 16)
2 6 1