Subcellular & Subsynaptic Localization of Group IMetabotropic Glutamate Receptors in the Nucleus Accumbens Public

Abstract

Changes in glutamate neurotransmission in the nucleus accumbens (NAc) have emerged as an important neuroadaptive mechanism in response to cocaine. There is evidence that group I metabotropic glutamate receptors (mGluR1 and mGluR5) in the NAc play an important role in the neurochemical and pathophysiological mechanisms that underlie addiction to psychostimulants. In order to understand the substrate by which group I mGluRs mediate their effects, we undertook a detailed analysis, using immunocytochemical electron microscopy (EM), of the ultrastructural localization of mGluR1a and mGluR5 in the NAc of normal and cocaine-treated animals.

We first examined the subcellular and subsynaptic localization of mGluR1a and mGluR5 in the shell and core of the NAc in normal monkey and rat. In both species, group I mGluRs are mainly postsynaptic in dendrites and spines, with rare presynaptic labeling in unmyelinated axons. At the subsynaptic level, these elements had a significantly larger proportion of plasma membrane-bound mGluR5 in rats compared to monkeys. Conversely, mGluR1a displayed the same pattern of labeling in the two species.

Next, using a combination of anterograde tract-tracing and EM immunocytochemistry, we found that limbic prefrontal cortical and midline thalamic terminals contacted mGluR1a- and mGluR5-immunoreactive dendrites and spines in both the shell and core, but to varying degrees. In addition, mGluR5 was found to be more frequently expressed perisynaptically, and closer extrasynaptically, at both types of synapses.

Finally, we looked at group I mGluR distribution in the accumbens of rats treated either acutely or chronically with cocaine. Surprisingly, only minor changes were seen in the subsynaptic localization of mGluR1a following acute and chronic cocaine exposures, while none were seen for mGluR5. In contrast, a significant reduction of plasma membrane-bound mGluR1a and mGluR5 was induced by local administration of the group I mGluR agonist, DHPG.

Overall, this work provides a detailed map of the localization of the group I mGluRs, including their relationship to two glutamatergic afferents, and how these receptors may be trafficked in response to cocaine or agonist stimulation, providing a foundation for the interpretation of future functional studies of these receptors in the NAc.

Table of Contents

Chapter 1: Introduction................................................................... 1

Preface................................................................................. ......... 2 1.1 Rationale behind this project............................................................ 3 1.2 The Nucleus Accumbens............................................................... 4 1.2.1 General information about the nucleus accumbens and its organization 4 1.2.2 Major Afferents and Efferents of the Nucleus Accumbens.................. 7 1.2.3 General Functions of the Nucleus Accumbens.............................. 10 1.2.4 Cocaine and the Nucleus Accumbens.......................................... 12 1.3 Glutamate Receptors..................................................................... 15 1.3.1 Ionotropic Glutamate Receptors................................................. 15 1.3.2 Overview of Metabotropic Glutamate Receptors.............................. 16 1.3.3 Group I mGluRs.................................................................. 17 1.3.4 Plasticity of GPCRs and the Group I mGluRs................................. 21 1.3.5 Group I mGluRs as Therapeutic Drug Targets................................. 24 1.3.6 Group I mGluRs' function and localization in other brain regions.......... 26 1.3.7 Functional Colocalization of mGluR1 and mGluR5.......................... 27 1.3.8 Group I mGluRs Localization and Function in the Nucleus Accumbens... 28 1.3.9 Group I mGluRs Role in Glia in the Nucleus Accumbens................... 30 1.3.10 Group I mGluRs and Cocaine................................................... 31 1.4 Summary of Research.................................................................... 32 1.4.1 Specific Aim 1......................................................................... 33 1.4.2 Specific Aim 2..................................................................... 33 1.4.3 Specific Aim 3..................................................................... 35 Chapter 2: Specific Aim 1: To compare the subcellular and subsynaptic localization of mGluR1a and mGluR5 in the shell and core of the nucleus accumbens in normal rats and monkeys........................................................................... 37 2.1 Abstract.................................................................................... 38 2.2 Introduction................................................................................ 39 2.3 Materials & Methods..................................................................... 41 2.3.1 Animals and tissue preparation................................................... 41 2.3.2 Primary Antibodies................................................................ 41 2.3.3 Single Immunoperoxidase labeling for light microscopy..................... 44 2.3.4 Single Immunoperoxidase labeling for electron microscopy................ 45 2.3.5 Single Preembedding Immunogold labeling for electron microscopy...... 46 2.3.6 Double Preembedding Immunoperoxidase labeling for electron microscopy colocalization.............................................................................. 47 2.3.7 Double Preembedding Immunoperoxidase and Immunogold labeling for electron microscopy colocalization of mGluR1a or mGluR5 with Neuronal Markers.................................................................................... 47 2.3.8 Analysis of Material............................................................... 48 2.3.8.1 Antibody Penetration in Tissue............................................ 48 2.3.8.2 Single immunoperoxidase labeling for group I mGluRs............... 48 2.3.8.3 Single immunogold labeling for group I mGluRs...................... 49

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Molecular & Systems Pharmacology
Degree	PhD
Submission	Dissertation
Language	English
Research Field	Biology, Neuroscience
Mot-clé	Nucleus Accumbens Immunocytochemistry Electron Microscopy mGluRs
Committee Chair / Thesis Advisor	Smith, Yoland, Emory University
Committee Members	Hall, Randy A, Emory University Howell, Leonard, Emory University Kuhar, Michael J, Emory University

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