Alpha-2 Adrenergic and Opioid Spinal Analgesic Synergy: Utility and Cellular Mechanisms A DISSERTATION SUBMITTED TO THE FACULTY OF THE GRADUATE SCHOOL OF THE UNIVERSITY OF MINNESOTA BY

i Acknowledgements I extend my deep thanks to the following individuals for their help, support and professional development. Kelley Kitto for unmatched technical skills and direct contributions to this work. Dr. Carrie Wade for understanding and humor throughout graduate school. Dan Schuster for friendship and assistance. H. Oanh Nguyen for always lending a hand. The Thesis Committee: Dr. George Wilcox for endless ideas, thorough review and interest in my research. Dr. Carolyn Fairbanks for always being helpful in maintaining progress and encouraging words. Dr. Lucy Vulchanova for insight and her love of science. Dr. Esam El-Fakahany for career development wisdom and numerous letters of support. Dr. Al Beitz for being gracious enough to join my committee. Abstract Agonists acting at spinal α 2-adrenergic receptors (α 2 AR) and opioid receptors (OR) produce analgesia through common intracellular signaling systems primarily mediated through inhibitory G proteins. Furthermore, co-activation of spinal α 2 AR and OR produces antinociceptive synergy. Synergistic analgesic interactions are important, as conventional opioid therapy is limited clinically due to the development of adverse side effects such as tolerance, dependence, abuse liability and opioid-induced hyperalgesia. Agonist combinations that interact synergistically may bypass these unwanted side effects by allowing decreases of analgesic dose and increasing the therapeutic index. Synergy between analgesic compounds has been shown experimentally and utilized clinically, yet the underlying cellular mechanisms mediating this phenomenon remain relatively unexplored. Elucidating the mechanisms underlying analgesic synergy may have broad clinical implications and may lead to the discovery of novel drug targets for pain management. The goal of this study was therefore to determine the cellular mechanisms mediating the synergistic interaction between agonists acting at two anatomically co-localized G protein-coupled receptors (GPCRs) in the spinal cord. In the first phase of these studies, we evaluated the ability of the selective delta-opioid receptor (DOP) agonist deltorphin II (DELT), the α 2 AR agonist clonidine (CLON) or their combination to inhibit nociceptive responses from mice iv in the tail flick test. We then examined the possible underlying signaling mechanisms involved through co-administration of inhibitors known to affect the above-mentioned receptor pair. Second, we looked at the ability of the DOP-selective agonist DELT, the α 2 AR agonist CLON, or their combination to inhibit calcitonin gene-related peptide (CGRP) release from spinal cord slices and spinal cord synaptosomes. Third, we determined the specific signaling mediator involved in α 2 AR/DOP analgesic synergy using genetically manipulated mice. …