THE UNIVERSITY OF CHICAGO MECHANISMS UNDERLYING OCD-RELEVANT BEHAVIORS IN MICE A DISSERTATION SUBMITTED TO THE FACULTY OF THE DIVISION OF THE BIOLOGICAL SCIENCES AND THE PRITZKER SCHOOL OF MEDICINE IN CANDIDACY FOR THE DEGREE OF DOCTOR OF PHILOSOPHY COMMITTEE ON NEUROBIOLOGY BY SUMMER LORETTA THOMPSON CHICAGO, ILLINOIS DECEMBER 2017 Copyright © 2017 by Summer Loretta Thompson All Rights Reserved TABLE OF CONTENTS LIST OF FIGURES . vii LIST OF TABLES . viii ACKNOWLEDGMENTS . ix ABSTRACT . xi 1 GENERAL INTRODUCTION . 1 1.1 Obsessive-compulsive disorder .......................... 1 1.1.1 Background ................................ 1 1.1.2 Neural circuitry .............................. 1 1.1.3 Psychiatric genetics and OCD ...................... 2 1.2 Animal models of OCD: a conceptual framework ................ 4 1.2.1 Can an animal have OCD? ........................ 4 1.2.2 Validity .................................. 5 1.2.3 Diagnostic boundaries and endophenotypes . 10 1.2.4 Mechanisms, disease models, and behavioral tests . 12 1.2.5 Utility ................................... 13 1.3 Pharmacological and behavioral rodent models of obsessive-compulsive disorder 18 1.3.1 OCD background ............................. 18 1.3.2 Principles of animal modeling ...................... 20 1.3.3 Pharmacologically induced models of obsessive-compulsive disorder . 24 1.3.4 Environmentally induced models .................... 30 1.3.5 Spontaneous models ........................... 32 1.3.6 Conclusions ................................ 35 2 DISTINCT ROLES FOR β-ARRESTIN2 AND GSK-3 SIGNALING IN 5-HT1BR- MEDIATED PERSEVERATIVE BEHAVIOR AND PREPULSE INHIBITION DEFICITS IN MICE . 37 2.1 Introduction .................................... 37 2.2 Materials and methods .............................. 40 2.2.1 Animals .................................. 40 2.2.2 Drugs ................................... 40 2.2.3 Behavioral testing ............................. 41 2.2.4 Experiments ................................ 42 2.2.5 Statistical analysis ............................ 43 2.3 Results ....................................... 44 2.3.1 GSK-3 inhibition does not affect RU24969-induced open field phenotypes 44 2.3.2 GSK-3 inhibition mitigates RU24969-induced PPI deficits . 45 2.3.3 β-arrestin2 expression affects RU24969-induced open field phenotypes 47 2.3.4 β-arrestin2 expression affects RU24969-induced PPI deficits . 49 iii 2.4 Discussion ..................................... 49 2.5 Supplemental material .............................. 57 2.5.1 Results ................................... 57 3 DOSE-DEPENDENT EFFECTS OF KETAMINE ON 5-HT1BR-INDUCED OCD- LIKE BEHAVIOR IN MICE . 68 3.1 Introduction .................................... 68 3.2 Materials and methods .............................. 70 3.2.1 Animals .................................. 70 3.2.2 Drugs ................................... 70 3.2.3 Open field ................................. 71 3.2.4 Prepulse inhibition ............................ 71 3.2.5 Experiment 1 ............................... 71 3.2.6 Experiment 2 ............................... 72 3.2.7 Experiment 3 ............................... 72 3.2.8 Experiment 4 ............................... 72 3.2.9 Experiment 5 ............................... 72 3.2.10 Statistical analysis ............................ 72 3.3 Results ....................................... 73 3.3.1 Mitigating effects of low-dose ketamine on RU24969-induced OCD-like behavior in the open field ........................ 73 3.3.2 No effect of low-dose ketamine on RU24969-induced PPI deficits . 75 3.3.3 Differential effects of high-dose ketamine on RU24969-induced behav- ioral effects in the open field by time point . 76 3.3.4 Differential effects of high-dose ketamine on RU24969-induced PPI deficits by time point ........................... 79 3.3.5 Acute fluoxetine pretreatment does not mitigate RU24969-induced OCD- like behavior ................................ 79 3.4 Discussion ..................................... 80 3.4.1 Overall ................................... 80 3.4.2 Effects of low-dose ketamine on perseverative hyperlocomotion . 81 3.4.3 Effects of high-dose ketamine on perseverative hyperlocomotion . 81 3.4.4 Effects of ketamine on PPI ........................ 82 3.4.5 Effects of fluoxetine on the 5-HT1BR-induced model . 82 3.4.6 Relation to human trials ......................... 83 3.4.7 5-HT1BR-induced model ......................... 84 3.4.8 Limitations ................................ 85 3.4.9 Conclusions ................................ 85 4 BTBD3 EXPRESSION IN HIPPOCAMPUS MODULATES OCD-RELEVANT BE- HAVIORS . 86 4.1 Introduction .................................... 86 4.2 Materials and methods .............................. 87 4.2.1 Animals .................................. 87 iv 4.2.2 Drugs ................................... 88 4.2.3 Open field ................................. 88 4.2.4 Barbering ................................. 89 4.2.5 Wheel-running .............................. 89 4.2.6 Dig test .................................. 89 4.2.7 Marble-burying .............................. 89 4.2.8 Nest building ............................... 90 4.2.9 Cognitive testing ............................. 90 4.2.10 Viral-mediated knockdown of Btbd3 expression . 93 4.2.11 Dendritic morphology .......................... 94 4.2.12 Statistical analysis ............................ 95 4.3 Results ....................................... 96 4.3.1 Btbd3 KO mice have deficits in compulsive-like behavior . 96 4.3.2 Btbd3 KO mice have behavioral deficits in measures of exploration . 98 4.3.3 Barbering behavior is preventable by chronic OCD-effective, but not -ineffective treatment in Btbd3 WT and HT but not KO mice . 100 4.3.4 Btbd3 knockdown in hippocampus during early development recapit- ulates KO deficits in adulthood . 102 4.3.5 Btbd3 KO mice have altered dendritic morphology in select brain regions104 4.4 Discussion ..................................... 106 4.4.1 Summary of findings . 106 4.4.2 Btbd3 knockout leads to compulsive-like behavioral deficits . 107 4.4.3 Btbd3 knockout causes deficits in exploratory behavior . 109 4.4.4 Barbering is selectively responsive to OCD-effective treatment . 109 4.4.5 Btbd3 knockdown in hippocampus recapitulates several behavioral deficits of the global knockout . 110 4.4.6 Behavioral effects of neonatal hippocampal Btbd3 knockdown are not primarily attributable to the dorsal hippocampus . 112 4.4.7 Behavioral effects of hippocampal Btbd3 knockdown are not confined to expression loss during the neonatal developmental period . 113 4.4.8 Behavioral effects of hippocampal Btbd3 knockdown are not attributable to cell-autonomous effects on dendritic morphology . 113 4.4.9 Relationship of behavioral effects of Btbd3 to OCD . 114 4.4.10 Conclusions ................................ 115 4.5 Supplemental material .............................. 116 4.5.1 Methods .................................. 116 4.5.2 Results ................................... 121 5 DISCUSSION . 133 5.1 Overview ...................................... 133 5.1.1 Chapter 2 - Summary . 134 5.1.2 Chapter 2 - Limitations and future directions . 134 5.1.3 Chapter 3 - Summary . 136 v 5.1.4 Chapter 3 - Limitations and future directions . 137 5.1.5 Chapter 4 - Summary . 137 5.1.6 Chapter 4 - Limitations and future directions . 138 5.2 Conclusions .................................... 140 REFERENCES . 141 vi LIST OF FIGURES 2.1 GSK-3 inhibition in open field test ......................... 44 2.2 GSK-3 inhibition mitigated low dose, but not high dose . 46 2.3 Arrb2 genotype affects RU24969-induced open field test activity changes . 47 2.4 Arrb2 genotype affects RU24969-induced changes in prepulse inhibition . 48 2.5 Representative paths mice took in the open field in Experiments 4 (A-C) and 3 (D-F) ......................................... 51 2.6 GSK-3 inhibition did not affect RU24969-induced changes in additional measures of activity in the open field test ........................... 58 2.7 GSK-3 inhibition with a second GSK-3 inhibitor did not affect high dose RU24969- induced behavior in the open field or prepulse inhibition. 59 2.8 No effect of delayed GSK-3 inhibition on low dose RU24969-induced behavior in the open field ..................................... 60 2.9 Arrb2 genotype affected RU24969-induced changes in additional measures of ac- tivity in the open field test .............................. 64 2.10 Arrb2 genotype affects RU24969-induced changes in open field behavior regard- less of baseline activity level ............................. 66 3.1 Low-dose ketamine pretreatment mitigated RU24969-induced locomotor perse- veration ........................................ 74 3.2 High-dose ketamine pretreatment acutely exacerbated RU24969-induced locomo- tor perseveration ................................... 77 3.3 Acute fluoxetine pretreatment did not mitigate RU24969-induced locomotor per- severation or prepulse inhibition deficits ...................... 80 4.1 Btbd3 knockout mice deficits in exploration and goal-directed behavior . 97 4.2 Barbering behavior selectively prevented by fluoxetine treatment . 101 4.3 Hippocampal Btbd3 knockdown mice recapitulate some behavioral deficits . 103 4.4 Btbd3 expression modulates dendritic morphology in key brain regions . 105 4.5 Behavioral deficits in Btbd3 knockout mice are selective . 122 4.6 Behavioral deficits in Btbd3 knockout mice not due to gross impairments . 124 4.7 Effects of Btbd3 expression on responsiveness of barbering to fluoxetine . 126 4.8 Cre virus successfully knocks down