University of Bath PHD The effects of chronic methamphetamine treatment on amine metabolism in rat brain in vivo. Bardsley, Maria E. Award date: 1978 Awarding institution: University of Bath Link to publication Alternative formats If you require this document in an alternative format, please contact: [email protected] General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal ? Take down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Download date: 23. Sep. 2021 THE EFFECTS OF CHRONIC METHAMPHETAMINE TREATMENT ON AMINE METABOLISM IN RAT BRAIN IN VIVO Submitted by Marie E. Bardsley for the degree of Ph.D. of the University of Bath 1978 COPYRIGHT Attention is drawn to the fact that copyright of this thesis rests with its author. This copy of the thesis has been supplied on condition that anyone who consults it is understood to recognise that its copyright rests with its author and that no quotation from the thesis and no information derived from it may be published without the prior written consent of the author. This thesis may be made available for consultation within the University Library and may be photocopied or lent to other libraries for the purposes of consultation MARIE E. BARDSLEY ProQuest Number: U442557 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. uest. ProQuest U442557 Published by ProQuest LLC(2015). Copyright of the Dissertation is held by the Author. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code. Microform Edition © ProQuest LLC. ProQuest LLC 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 48106-1346 TO MY PARENTS The Gods did not reveal, from the beginning. All things to us, but in the course of time Through seeking we may l e a m and know things better. But as for certain truth, no man has known it. Nor shall he know it, neither of the gods Nor yet of all the things of which I speak. For if by chance he were to utter The final truth, he would himself not know it: For all is but a woven web of guesses. Xenophanes ACKNOWLEDGEMENTS I am deeply endebted to my supervisor Professor H.S. Bachelard for his support and patience with a student who persistently tried to run before she could walk. I am very grateful for his philosophical outlook towards my sometimes rather far-fetched and impractical ideas, allowing me to learn by experience. There are a large number of people who have helped me over the past three years. I would especially like to thank John Ridley, Dr. Elizabeth Trifilieff and Dr. Keith Wood for their help with the serotonin and tryptophan assays; Mrs Rosemary Chase and Margaret Grant for showing me how to use the Chromaspek amino-acid analyser, and for running my samples. I would also like to thank Enrico Coen and Kim Gardner for their technical assistance, and Dr. C. Pycock, Dr. P. Emson and Dr. A.C. Cuello for their advice. I would like to acknowledge the help of Jeff Venn in the workshop and Roger Francis in the animal house. Last, but not least, I am grateful to Richard Taylor and Helen Wise for their companionship in the laboratory. CONTENTS Page I ABSTRACT 1 II INTRODUCTION 2 A. Clinical aspects of amphetamine use 2 B. Biochemical effects of amphetamine in vivo and in vitro. 1. Structure-activity relationships 11 2. Storage of catecholamines and other putative neurotransmitters 13 3. Uptake of catecholamines and 5-hydroxy- tryptamine 20 4. Release of catecholamines, 5-hydroxy- tryptamine and acetyl choline 27 C. Biochemical effects of amphetamine on metabolism 41 1. Degradation of the catecholamines 41 2. Synthesis of the catecholamines 46 3. Turnover of the catecholamines 49 4. Degradation of 5-hydroxytryptamine 54 5. Synthesis and turnover of 5-hydroxytryptamine 55 6. Metabolism of acetyl choline 57 D. Effects of amphetamine on the cyclic nucleotides and glycogenolysis 58 E. General metabolic effects of amphetamine 61 administration. F. Summary of the biochemical effects of amphetamine 64 G. 'Catecholamines in the brain as mediators of the amphetamine psychosis' 66 H. Metabolism and the metabolites of amphetamine 68 1. Uptake into and release from adrenergically innervated tissues of amphetamine itself 68 2. Storage of amphetamine in the body 70 3. Metabolism of amphetamine 71 J. Aim of the research 79 III & IV MATERIALS, METHODS AND RESULTS 83 III ADMINISTRATION OF METHAMPHETAMINE IN THE DRINKING WATER 84 A. Treatment of animals and behavioural monitoring 84 1. Materials and Methods 84 2. Results 85 B. Choice of brain regions 91 D. Fluorometric determination of noradrenaline, dopamine, serotonin and histamine 103 1. Materials and Methods 103 2. Results 115 E . Comments on the route of administration of methamphetamine and the methodology used to measure amine levels 121 IV ADMINISTRATION OF METHAMPHETAMINE BY. INTRA-PERITONEAL INJECTION 125 A. Treatment of animals 125 1. Materials and Methods 125 2. Results 126 B. Regional dissection of the brain 138 C. Estimation of tyrosine hydroxylase activity 143 1. Materials and Methods 143 2. Results: Regional tyrosine hydroxylase activity in experimental animal groups 155 D. Radioenzymatic measurement of noradrenaline dopamine and their metabolites 164 1. Materials and Methods 164 2. Results: Regional amine levels in experimental animal groups 188 E. Determination of brain tryptophan levels 193 1. Materials and Methods 193 2. Results: Regional tryptophan levels in the experimental animals groups 195 F. Measurement of brain serotonin levels 207 1. Materials and Methods 207 2, Results 212 G. Determination of amino acid levels in brain regions 219 1. Materials and Methods 219 2. Results 221 H. Determination of total and free tryptophan levels, phenylalanine and tyrosine levels in plasma 231 1. Materials and Methods 231 2. Results 233 V DISCUSSION 237 A. Catecholamine metabolism 237 1. Levels of phenylalanine and tyrosine in the plasma and in the brain 237 2. Tyrosine hydroxylase activity . 238 3. Levels of catecholamines and their metabolites in brain regions 241 4. Conclusions 244 B. Serotonin metabolism 246 1. Levels of tryptophan in theplasma and in the brain 246 2. Brain serotonin 252 C. Amino acid uptake 253 D. Radioenzymatic assays 254 E. Behavioural correlates 258 F. General Comments 266 1. Controls 266 2. Enzymic adaptation 267 3. Tyrosine hydroxulase 268 4. Alterations on the "blood-brain barrier" on amphetamine treatment 269 G. Conclusions 270 H. 1 - 6 Future Work 272 REFERENCES 274 — I— I. ABSTRACT Rats were chronically treated with methamphetamine over a seven week period during which time their weights, food consumption and drinking rates were recorded. The effects of chronic treatment with, and withdrawal from, methamphetamine on amine metabolism in selected brain regions was investigated. Three groups of experimental animals were studied; chronically treated rats, rats which had been withdrawn from the drug for 36 h and controls. The parameters chosen to reflect changes in catecholamine metabolism in the brain were: tyrosine hydroxylase activity and the levels of noradrenaline, dopamine and their non-O-methylated meta­ bolites and of phenylalanine and tyrosine. The metabolism of serotonin was studied by measuring the levels of "free" tryptophan in the plasma,and the levels of tryptophan and serotonin in the brain. Initially the methamphetamine was administered in the drinking water at doses of 5, 10, 20 and 40 mg/kg/24 h increasing in a stepwise manner over a period of 20 days. This dose was found to produce toxic reactions in the rats and it was found that it was not possible to control the intake of the drug. In subsequent experiments the rats were given 5, 10 and 15 mg/kg/24 h i.p. increasing in a stepwise manner over a period of 15 days. The first set of injected animals did not develop tolerance to the anorexic effects of methamphetamine and the chronically treated and withdrawn animals showed changes in tyrosine hydroxylase activity and in the levels of noradrenaline and dopamine. -la- The three other sets of injected animals did develop tolerance to the anorexic effects of methamphetamine. Tyrosine hydroxylase and the levels of noradrenaline, dopamine, their non-O-methylated metabolites and of tryptophan and serotonin did not change in these chronically-treated or withdrawn animals, but changes were seen in the levels of "free" tryptophan in the plasma and the levels of most amino acids in the brain. —2— II. INTRODUCTION A. CLINICAL ASPECTS OF AMPHETAMINE USE Historically the study of amphetamines began in 1887 when Edeleano pre­ pared the volatile^amine phenylisopropylamine (amphetamine). Barger and Dale (1910) found that it was related to adrenaline in its pharma­ cological actions (see Connell, 1970). In 1927 Allés synthesized amphetamine in the course of a search for a substance to replace ephe- drine, which was difficult to obtain from its natural sources.