TITLE AND INDEX TRACE AMINE-ASSOCIATED RECEPTOR 1 REGULATION OF METHAMPHETAMINE INTAKE AND AVERSION By JOHN H. HARKNESS A DISSERTATION Presented to the Department of Behavioral Neuroscience and the Oregon Health & Science University School of Medicine in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy September 2015 TABLE OF CONTENTS LIST OF FIGURES v LIST OF TABLES vii LIST OF ABBREVIATIONS viii ACKNOWLEDGEMENTS xiii ABSTRACT xv CHAPTER 1: General Introduction 1 History of Methamphetamine Use 1 Methamphetamine as a Drug of Abuse 1 Impact on Health and Communities 4 Current Therapies for Methamphetamine Abuse 5 Pharmacology of Methamphetamine 8 Pharmacokinetics 9 Absorption and Distribution 9 Metabolism and Excretion 11 Pharmacodynamics 12 Actions at Monoamine transporters 12 Trace Amine-Associated Receptor 1 (TAAR1) 13 Factors Mediating Methamphetamine Addiction 18 Adaptations to the Basal Ganglia Reward Pathway 18 Positive and Negative Subjective Effects of 20 Methamphetamine i Physiological Effects 22 Stress Axis Activation 22 Thermal Effects 24 Genetic Influences in Drug Use Behavior 25 Forward and Reverse Genetic Approaches 26 Quantitative Traits 28 QTL Mapping Populations 31 Crosses Derived from Inbred Strains 31 Selective Bred Lines 34 The High and Low MA Drinking (MADR) 39 Selected Lines Methods to Study Drug Effects in Animals 41 Two-Bottle Choice Drinking 42 Self-Administration 45 Conditioned Place Preference and Place 46 Aversion Conditioned Taste Aversion 50 QTL Mapping and Gene Expression Analysis in the MADR 51 Lines Taar1 Knockout Mice 53 Experimental Goals and Hypotheses 55 ii CHAPTER 2: Trace Amine-Associated Receptor 1 Regulation of 58 Methamphetamine Intake and Related Traits CHAPTER 3: Trace Amine-Associated Receptor 1 and Stress 94 Axis Regulation of Methamphetamine Intake CHAPTER 4: General Discussion 139 Goals and Hypotheses 139 Main Findings 140 Responses to Methamphetamine in MADR Mice 145 Methamphetamine Consumption 153 Taste Perception in Methamphetamine Consumption 154 Cognitive, Impulsivity, and Anxiety-like Traits in 156 Methamphetamine Consumption Methamphetamine Reward 159 Methamphetamine Stimulation 163 Methamphetamine Aversion 165 TAAR1-Associated Methamphetamine Effects 167 Thermal Effects 167 Stress Axis Response to Methamphetamine 169 MADR Line Responses to Cocaine and Ethanol 170 Gene Expression Differences in the MADR Lines 172 iii Monoamine and TAAR1 Systems in 176 Methamphetamine Intake NET and SERT in Aversive Effects of 178 Methamphetamine Considerations of the Genetic Models 179 Summary and Conclusions 182 Future Directions 186 REFERENCES 189 iv LIST OF FIGURES Figure 1.1: Proposed mechanism of action for TAAR1 regulation of 16 monoamine activity following MA exposure. Figure 2.1: A quantitative trait locus (QTL) on mouse chromosome 64 10 has a major effect on MA drinking in MADR mice. Figure 2.2: Schematic transmembrane topology of mouse TAAR1 75 and frequency of B6- and D2-like Taar1 alleles in MALDR and MAHDR mice. Figure 2.3: MA consumption and sensitivity to MA-induced CTA 77 differs by Taar1 genotype Figure 2.4: MA- and ethanol-induced hypothermia differs by Taar1 81 genotype. Figure 2.5: The effect of MA on body temperature differs in 84 replicate 3 MAHDR and MALDR mice. Figure 2.6: Function and expression of B6-like and D2-like 86 isoforms of TAAR1 in Taar1- transfected cells. Figure 3.1: Plasma ACTH and CORT difference values from saline 110 baseline after administration of several doses of MA. Figure 3.2: Plasma CORT difference values from saline baseline 112 60 minutes after administration of 2 mg/kg MA in MADR and TAAR1 transgenic mice. v Figure 3.3: MA consumption over days by 116 MAHDR mice offered MA in solution with corticosterone. Figure 3.4: MA preference vs. water over days by MAHDR mice 118 offered MA in solution with CORT. Figure 3.5: Plasma CORT levels 8 h after consuming water, CORT, MA or MA+CORT in MAHDR mice. 120 Figure 3.6: NISX-induced CTA in MAHDR and MALDR mice. 124 Figure 3.7: FLUX-induced CTA between the MADR lines. 126 Figure 3.8: Repeated treatment with NISX prior to MA-induced CTA in the MADR lines. 128 Figure 3.9: Repeated treatment with FLUX prior to MA-induced CTA between the MADR lines. 129 Figure 4.1: Venn diagram of MA-related traits presented in Chapters 2 and 3, organized by mouse line. 184 vi LIST OF TABLES 142 Table 4.1: Summary of Current Results Table 4.2: Summary of Previous Findings in MADR Lines and 146 Progenitor Strains vii LIST OF ABBREVIATIONS +/+ – wildtype genotype CeA – central amygdala +/- – heterozygous genotype cM – centimorgan -/- – knockout genotype Cmax – maximum (peak) plasma 5-HT – serotonin concentration 5-HT1A – serotonin receptor 1A CNS – central nervous system 5-HT1B – serotonin receptor 1B CORT – corticosterone ACTH – adrenocorticotropin CPA – conditioned place aversion hormone CPP – conditioned place preference ANOVA – analysis of variance CRF – corticotropin-releasing factor Ar – androgen receptor gene CS+ – conditioned positive stimulus B6 – C57BL6/J CS- – conditioned negative stimulus B6D2F2 – F2 generation of C57BL/6J and DBA/2J cross CTA - conditioned taste aversion BCA – bicinchoninic acid D1 – receptors bp – base pair D2 – receptors BXD – C57BL/6J and DBA/2J D3 – receptors recombinant inbred line(s) D2 – DBA/2J cAMP –3'-5'-cyclic adenosine monophosphate DA – dopamine Casp8 – caspase 8, apoptosis- DAT – dopamine transporter related cysteine peptidase gene viii d-isomer – dextrorotatory isomer Fos – FOS gene dl – deciliter GABA – γ-aminobutyric acid DMSO – dimethyl sulfoxide GFP – green fluorescent protein EC50 – half maximal effective GLU – glutamate concentration Grm4 – metabotropic glutamate EIA – enzyme Immunoassay receptor 4 gene ELISA – enzyme-linked H2 – heritability immunosorbent assay HIV – human immunodeficiency ES – embryonic stem virus Esr1 – estrogen receptor 1 gene Hivep2 – human immunodeficiency virus type 1 enhancer binding Esr2 – estrogen receptor 2 gene protein 2 gene EtOH – ethanol h – hour EPPTB – N-(3-Ethoxy-phenyl)-4- HMACT - high methamphetamine pyrrolidin-1-yl-3-trifluoromethyl- activation benzamide HPA – hypothalamic-pituitary- adrenal axis eQTL – expression quantitative trait locus Hsce70 – immunoglobulin heavy chain binding protein 70 gene F – female Hsp40 – heat shock protein 40 gene F2 – second filial generation offspring Htr3 – serotonin receptor 3 gene FCS – fetal calf serum Il6 – interleukin 6 gene FLUX – fluoxetine i.p. – intraperitoneal ix i.v. – intravenous MALSENS – methamphetamine low sensitization mouse line KCl – potassium chloride Map3k5 – mitogen-activated protein kg – kilogram kinase kinase kinase 5 gene KO – knockout Mapk3 – ERK-1 MAP kinase gene l – liter Mb – mega base LC – locus coeruleus MDMA – 3,4-methylenedioxy- methamphetamine LiCl – lithium chloride µg – microgram l-isomer – levorotatory isomer mg – milligram LMACT – low methamphetamine activation mg/kg – milligram per kilogram LOD – log of the odds mGlur1 – metabotropic glutamate receptor 1 gene M – male Mpdz – multiple PDZ domain protein MA – methamphetamine gene MADR – methamphetamine drinking mPFC – medial prefrontal cortex mouse lines NAcc – nucleus accumbens MAHDR – methamphetamine high drinking mouse line NaCl – sodium chloride MAHSENS – methamphetamine NE - norepinephrine high sensitization mouse line NET – norepinephrine transporter MALDR – methamphetamine low drinking mouse line NIDA – National Institute on Drug Abuse x NISX – nisoxetine QTG – quantitative trait gene nM – nanomolar QTL – quantitative trait locus NFκB – nuclear factor of kappa light Rela – NFκB subunit p65 gene polypeptide Rep – replicate Nfkb1 – nuclear factor of kappa light polypeptide gene enhancer in B- RI – recombinant inbred cells 1 gene RIPA – radioimmunoprecipitation Nfkb2 – nuclear factor of kappa light assay buffer polypeptide gene enhancer in B- cells 2 gene SA – self-administration NTS – nucleus of the solitary tract SEM – standard error of the mean Oprm1 – µ opioid receptor gene SERT – serotonin transporter PCR – polymerase chain reaction Slc6a2 – NET gene PFC – prefrontal cortex Slc6a4 – SERT gene pg – picogram SNP – single nucleotide polymorphism PKA – protein kinase A Τ – circadian free-running period PKC – protein kinase C T0AM – thyronamine POMC – proopiomelanocortin T1AM – 3-iodothyronamine PVN –paraventricular nucleus of the hypothalamus Taar1 – gene encoding trace amine- associated receptor 1 qPCR – quantitative polymerase chain reaction xi TAAR1 – trace amine-associated receptor 1 UNODC – United Nations Office on Drugs and Crime US – unconditioned stimulus VAPORHCS – Veterans Affairs Portland Health Care System VMAT2 – vesicular monoamine transporter 2 VMB – ventral midbrain WHO – World Health Organization WT – wildtype UCP – uncoupling protein UCP1 – uncoupling protein 1 UCP3 – uncoupling protein xii ACKNOWLEDGMENTS I am grateful to have been surrounded by many brilliant and inquisitive minds throughout my journey to this point. These people instilled in me a sense of curiosity and discovery that makes me hungry to explore and push boundaries. I attribute many of my successes to their support and influence. My mom, dad, and brother encouraged me without reservation, even when the path was hard or uncertain. They taught me to investigate and challenge the world around me, and pushed me to jump from the highest rock, but only after checking the depth of the water. Together, my family has shown me unconditional love and support for which I am forever grateful. I would also