Defense Date

2010

Document Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Pharmacology & Toxicology

First Advisor

Laura Sim-Selley

Abstract

CB1 receptors (CB1R) mediate the psychoactive and therapeutic effects of cannabinoids including ∆9-tetrahydrocannabinol (THC), the main psychoactive constituent in marijuana. However, therapeutic use is limited by side effects and tolerance and dependence with chronic administration. Tolerance to cannabinoid-mediated effects is associated with CB1R adaptations, including desensitization (receptor-G-protein uncoupling) and downregulation (receptor degradation). The objectives of this thesis are to investigate the regional-specificity in CB1R function and regulation. Previous studies have investigated CB1Rs in a subset of regions involved in cannabinoid effects, but an inclusive regional comparison of the relative efficacies of different classes of cannabinoids to activate G-proteins has not been conducted. A novel unbiased whole-brain analysis was developed based on Statistical Parametric Mapping (SPM) for 3D-reconstructed mouse brain images derived from agonist-stimulated [35S]GTPgS autoradiography, which has not been described before. SPM demonstrated regional differences in the relative efficacies of cannabinoid agonists methanandamide (M-AEA), CP55,940 (CP), and WIN55,212-2 (WIN) in mouse brains. To assess potential contribution of novel sites, CB1R knockout (KO) mice were used. SPM analysis revealed that WIN, but not CP or M-AEA, stimulated [35S]GTPgS binding in regions that partially overlapped with the expression of CB1Rs. We then examined the role of the regulatory protein Beta-arrestin-2 (βarr2) in CB1R adaptations to chronic THC treatment. Deletion of βarr2 reduced CB1R desensitization/downregulation in the cerebellum, caudal periaqueductal gray (PAG), and spinal cord. However in hippocampus, amygdala and rostral PAG, similar desensitization was present in both genotypes. Interestingly, enhanced desensitization was found in the hypothalamus and cortex in βarr2 KO animals. Intra-regional differences in the magnitude of desensitization were noted in the caudal hippocampus, where βarr2 KO animals exhibited greater desensitization compared to WT. Regional differences in βarr2-mediated CB1R adaptation were associated with differential effects on tolerance, where THC-mediated antinociception, but not catalepsy or hypothermia, was attenuated in βarr2 KO mice. Overall, studies using SPM revealed intra- and inter-regional specificity in the function and regulation of CB1Rs and underscores an advantage of using a whole-brain unbiased approach. Understanding the regulation of CB1R signaling within different anatomical contexts represents an important fundamental prerequisite in the therapeutic exploitation of the cannabinoid system.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

September 2010

coronal_FigS2 B-1.mov (380 kB)
reconstruction video _coronal

sagittal_FigS2 B-2.mov (1355 kB)
reconstruction video _sagittal

transverse_FigS2 B-3.mov (1183 kB)
reconstruction video _transverse

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