Examining the role of the insular cortex in addiction- relevant behaviours by Abhiram Pierre Pushparaj A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy Department of Pharmacology & Toxicology University of Toronto © Copyright by Abhiram Pierre Pushparaj 2016 Examining the role of the insular cortex in addiction-relevant behaviours Abhiram Pierre Pushparaj Doctor of Philosophy Department of Pharmacology University of Toronto 2016 Abstract INTRODUCTION: Recently recognized as a critical brain region underlying the neurocircuitry of addiction, the body of evidence supporting the role of the insular cortex across multiple aspects of addiction is growing rapidly. Understanding the complex role of the insular cortex is crucial to moving forward with the development of therapeutics targeting this brain region for substance-related and addictive disorders. OBJECTIVES: The current series of experiments were conducted to investigate the effects of modulating activity in two insular subregions, through the use of multiple modalities (chemical lesions, pharmacological inactivation or electrical stimulation), on behaviours relevant to addictive disorders, including drug self-administration, reinstatement, and decision-making. METHODS: In the first series of experiments, we investigated the effects of chemical lesions or transient pharmacological inactivation of two distinct subregions of the insular cortex, the rostral agranular (RAIC) vs. caudal granular (CGIC), on the acquisition or performance of choice behaviour in the Rat Gambling Task (rGT). In the next series of experiments, we examined the effects of pharmacological inactivation of the RAIC on nicotine self-administration under a fixed ratio-5 (FR-5) schedule of reinforcement and the reinstatement of nicotine-seeking induced by ii nicotine-associated cues. Finally, we examined the effects of electrical modulation of the insular region on nicotine self-administration under both FR-5 and progressive ratio schedules of reinforcement and the reinstatement of nicotine-seeking by cues and nicotine priming injections. RESULTS: Lesions of the RAIC, but not CGIC, resulted in higher preference for options which produced lower reward amounts with higher reward consistency while inactivation of the RAIC, but not CGIC, shifted previously acquired preferences towards those same low reward/high consistency options. Inactivation of the RAIC attenuated nicotine self-administration and cue- induced reinstatement. Finally, electrical modulation functionally inactivated the insular region, resulting in attenuated nicotine self-administration, under FR-5 and progressive ratios, and of cue- and priming-induced reinstatement. CONCLUSIONS: This work demonstrates involvement of the RAIC, but not CGIC, in cognitive aspects of addictive disorders, such as gambling-relevant behaviours. Yet both regions were demonstrated to be implicated in substance-based addictive disorders. Thus, therapeutics using electrical modulation to target the CGIC may be preferable for substance dependence, with limited cognitive side effects. iii Table of Contents Table of Contents ........................................................................................................................... iv List of Publications ....................................................................................................................... vii List of Abbreviations ..................................................................................................................... ix List of Figures ............................................................................................................................... xii List of Tables ............................................................................................................................... xiii CHAPTER 1. Introduction .............................................................................................. 1 1.1 Substance-related and Addictive Disorders ........................................................................ 1 1.1.1 Addiction: A Major Burden for Canada and the World ......................................... 1 1.1.2 Defining Addiction: Diagnostic Criterion Underlying Addictive Disorders .......... 2 1.1.3 Treating Addiction: A Lack of Highly Efficacious Therapies ................................ 4 1.1.4 Identifying Novel Therapies: The Use of Animal Models ..................................... 5 1.2 The Insular Cortex in the Neurocircuitry of Addiction .................................................... 12 1.2.1 The Cytoarchitecture of the Insular Cortex .......................................................... 12 1.2.2 The Insular Cortex and Interoception ................................................................... 13 1.2.3 Addiction-Relevant Neuronal Receptor Populations within the Insular Cortex ... 15 1.2.4 Damage to the Insular Cortex Disrupts Smoking Addiction ................................ 17 1.2.5 The Insular Cortex within the Neurocircuitry Underlying Addiction .................. 20 1.3 Experimental Rationales ................................................................................................... 51 1.3.1 Rationale for Investigating the Differential Involvement of the Agranular vs Granular Insular Cortex in the Acquisition and Performance of Choice Behavior in a Rodent Gambling Task ................................................................... 52 1.3.2 Rationale for Examining the Involvement of the Rostral Agranular Insular Cortex in Nicotine Self-Administration in Rats .................................................... 54 1.3.3 Rationale for Examining the Effect of Electrically Modulating Insular Cortex Activity on Nicotine Taking and Seeking Behaviour ........................................... 55 iv CHAPTER 2. Differential Involvement of the Agranular vs Granular Insular Cortex in the Acquisition and Performance of Choice Behavior in a Rodent Gambling Task ........... 57 2.1 Abstract ............................................................................................................................. 58 2.2 Introduction ....................................................................................................................... 59 2.3 Materials and Methods ...................................................................................................... 61 2.4 Results ............................................................................................................................... 68 2.5 Discussion ......................................................................................................................... 82 CHAPTER 3. Involvement of the Rostral Agranular Insular Cortex in Nicotine Self-Administration in Rats ...................................................................................................... 88 3.1 Abstract ............................................................................................................................. 89 3.2 Introduction ....................................................................................................................... 90 3.3 Materials and Methods ...................................................................................................... 92 3.4 Results ............................................................................................................................... 99 3.5 Discussion ....................................................................................................................... 105 CHAPTER 4. Electrical Stimulation of the Insular Region Attenuates Nicotine- Taking and Nicotine-Seeking Behaviors ............................................................................... 110 4.1 Abstract ........................................................................................................................... 111 4.2 Introduction ..................................................................................................................... 112 4.3 Materials and Methods .................................................................................................... 114 4.4 Results ............................................................................................................................. 123 4.5 Discussion ....................................................................................................................... 133 CHAPTER 5. General Discussion ............................................................................... 137 5.1 Summary of Critical Findings ......................................................................................... 137 5.1.1 Critical Findings from "Differential Involvement of the Agranular vs. Granular Insular Cortex in the Acquisition and Performance of Choice Behaviour in a Rodent Gambling Task" ............................................................. 138 5.1.2 Critical Findings from "Involvement of the Rostral Agranular Insular Cortex in Nicotine Self-Administration in Rats ............................................................. 140 v 5.1.3 Critical Findings from "Electrical Stimulation of the Insular Region Attenuates Nicotine-Taking and Nicotine-Seeking Behaviors" ......................... 143 5.2 Validity and Limitations with Regards to the Clinical Situation .................................... 145 5.2.1 Animal Models of Behaviour .............................................................................