NORMAL AGING AND COGNITION: SYSTEM-SPECIFIC CHANGES TO G-PROTEIN COUPLED RECEPTOR-MEDIATED SIGNAL TRANSDUCTION WITHIN THE HIPPOCAMPUS BY JOSEPH A. MCQUAIL A Dissertation Submitted to the Graduate Faculty of WAKE FOREST UNIVERSITY GRADUATE SCHOOL OF ARTS AND SCIENCES in Partial Fulfillment of the Requirements for the Degree of DOCTOR OF PHILOSOPHY Neuroscience May 2013 Winston-Salem, North Carolina Approved By: Michelle M. Nicolle, Ph.D., Advisor Examining Committee: David R. Riddle, Ph.D., Chairman Allyn C. Howlett, Ph.D. Mary Lou Voytko, Ph.D. Scott E. Hemby, Ph.D. ACKNOWLEDGEMENTS It was only with the greatest support and consideration from my advisor, Dr. Michelle Nicolle, that I could ever have achieved anything at all as a student of science. Our shared interest in cognitive aging was merely the beginning to what ultimately became my deepest professional relationship. Her guidance and mentorship as a scientist are only exceeded by her advice and concern as a friend. I owe her the greatest debt of thanks for welcoming me into her lab, nurturing my research interests and helping me to become the scientist that I am today. I would also like to thank each of my committee members. Dr. David Riddle was a constant source of encouragement, generous with his advice and a polite devil’s advocate; I was always grateful for his input. Dr. Mary Lou Voytko offered fantastic insights into neurocognitive aging, but also helped me to learn about teaching others; I appreciate that she welcomed me aboard as a teaching assistant so early in my career and continued to help me refine my teaching abilities. Dr. Allyn Howlett helped me transform a small idea inspired by a single article into a fully-fledged proposal and invited me into her lab to make it a reality; her support and enthusiasm for my research never waned. Dr. Scott Hemby made his expertise and resources readily available to address the countless questions and issues that cropped over the duration of my time in the lab; he was a fantastic “lab neighbor” and his generosity enabled so much of my work. Individually and collectively, I am indebted to these great scientists for their direction and consideration. I would like to thank all of the scientists who helped me along the way on my lengthy professional journey. Drs. Linda Werling, Joshua Burk, Pamela Hunt, Michela Gallagher and ii Jessica Mong were all instrumental in shaping the course of my development as an aspiring neuroscientist. They all helped me to arrive at this moment and achieve this great distinction. Most importantly, I must thank my friends and family for their support throughout this experience. Stephanie Willard and Tamara Spence were my closest friends here at Wake Forest and together we shared and endured the challenges of graduate school. My closest friend from high school, Ana Oancea, was constant source of understanding and support as we compared notes our respective graduate education experiences. Finally, it is the love and encouragement of two great women, my mother, Diane McQuail, and my girlfriend, Elizabeth Currin, that made this whole experience and everything that comes after seem truly meaningful. I owe them the greatest “thank you” of all. iii TABLE OF CONTENTS PAGE LIST OF ABBREVIATIONS ……………………………………………………………………..v LIST OF TABLES………………………………………………………………………………..ix LIST OF FIGURES…………………………………………………………………………..…...x ABSTRACT……………………………………………………………………………………...xi CHAPTER I. INTRODUCTION.……………………………………………………………………...1 II. A MODEL OF COGNITIVE AGING IN THE F344 × BROWN NORWAY F1 HYBRID RAT: RESULTS FROM A TRIAL-BASED ANALYSIS..………………….73 Supplementary Text from Neuropharmacology 70:64–73, 2013 III. NEUROINFLAMMATION NOT ASSOCIATED WITH CHOLINERGIC DEGENERATION IN AGED-IMPAIRED BRAIN…………………………………….93 Published in Neurobiology of Aging 32(12):2322.e1–2322.e4, 2011 IV. GABAB RECEPTOR GTP-BINDING IS DECREASED IN THE PREFRONTAL CORTEX BUT NOT THE HIPPOCAMPUS OF AGED RATS…….110 Published in Neurobiology of Aging 33(6):1124.e1–1124.e12, 2012 V. HIPPOCAMPAL Gαq/11 BUT NOT Gαo-COUPLED RECEPTORS ARE ALTERED IN AGING…………………………………………………………………146 Published in Neuropharmcology 70:64–73, 2013 VI. DISCUSSION………………………………………………………………………190 APPENDIX……………………………………………………………...……….……..228 CURRICULUM VITAE………………………………………………............……..…256 iv LIST OF ABBREVIATIONS 2+ 2+ [Ca ]i intracellular Ca concentration ABC avidin biotin complex AC adenylyl cyclase ACh acetylcholine ACh acetylcholine AD Alzheimer's disease AI aged-impaired AMPA 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid AMPAR AMPA receptor ANOVA analysis of variance APV (2R)-amino-5-phosphonovaleric acid ARC Aging Rodent Colony ATP adenosine-5'-triphosphate AU aged-unimpaired AUC area under the curve BAPTA-AM 1,2-bis(o-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid, acetoxymethyl- conjugated CA1 Cornu Ammonis 1 CA2 Cornu Ammonis 2 CA3 Cornu Ammonis 3 cAMP 3'-5'-cyclic adenosine monophosphate CCh carbachol ChAT choline acetyltransferase CICR Ca2+-induced Ca2+ release v CPA cyclopiazonic acid CPM counts per minute DAG diacylglycerol DAPI 40,6-diamidino-2-phenylindole dihydrochloride DG dentate gyrus DHPG (S)-3,5-dihydroxyphenylglycine DNMS delayed non-matching to sample DR delayed recognition EC50 half maximal effective concentration EDTA ethylenediaminetetraacetic acid EGTA ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid EMAX maximum possible effect for agonist ER endoplasmic reticulum F0 initial fluorescence F344 Fisher 344 FBNF1 F344 × Brown Norway F1 hybrid fMRI functional MRI GABA γ-Aminobutyric acid GABAAR GABA A receptor GABABRs GABA B receptor GAD-67 67 kDa isoform of glutamic acid decarboxylase GDP Guanosine diphosphate GPCR G-protein coupled receptor GTP Guanosine-5'-triphosphate GTP-Eu europium-labelled GTP vi GTPγS guanosine-5’-O-(3-thio)triphosphate HC-3 hemicholinium-3 HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid ICS intracellular Ca2+ store IgG immunoglobulin G IP inositol phosphate IP3 inositol 1,4,5-trisphosphate IP3R IP3 receptor LE Long-Evans LTD long term depression LTP long term potentiation mAChR muscarinic acetylcholine receptor MCI mild cognitive impairment mGluR metabotropic glutamate receptor ML molecular layer MRI magnetic resonance imaging MS medial septum nAChR nicotinic acetylcholine receptor NH Hill slope coefficient NHP non-human primate NIA National Institute on Aging NMDA N-Methyl-D-aspartate NMDAR NMDA receptor NPY neuropeptide Y OML outer molecular layer vii PFC prefrontal cortex PI phosphoinositide PIP2 phosphatidylinositol 4,5-bisphosphate PKA protein kinase A PLC phospholipase C RGS regulator of G-protein signaling RMANOVA repeated measures ANOVA RyRs ryanodine receptor SC Schaeffer collatoral sIPSP slow inhibitory post-synaptic potential SLI spatial learning index SPA scintillation proximity assay SR stratum radiation SST somatostatin TBS tris-buffered saline VAChT vesicular acetylcholine transporter VDB vertical diagonal band VGCC voltage gated calcium channel ΔF change in fluorescence viii LIST OF TABLES PAGE CHAPTER I Table 1.1. Classification and Properties of GPCRs in the Hippocampus…………………...72 CHAPTER IV Table 4.1. Parameters of baclofen-stimulated GTP-binding in the hippocampus of young and aged F344 rats…………………...………….……………………144 Table 4.2. Parameters of baclofen-stimulated GTP-binding in the PFC of young and aged F344 rats………………………………..……………………………145 CHAPTER V Table 5.S1. Results of correlation analyses between proximity scores and neurobiological parameters in young and aged rats………………………….…189 ix LIST OF FIGURES PAGE CHAPTER I Figure 1.1. The interaction between chronological or biological aging and cognitive function (“cognitive aging”)……………………………………………………..69 Figure 1.2. Schematic illustration of the hippocampus and adjacent structures…………..…70 Figure 1.3. Key GPCRs in the hippocampus…………………………………………………71 CHAPTER II Figure 2.1. Training trial performance of young and aged rats………………………………89 Figure 2.2. Quadrant-derived measures characterizing probe trial performance in young and aged rats……………………………………………………………...90 Figure 2.3. Savings scores following 24 hour delay imposed in each training block in young, aged-unimpaired and aged-impaired rats……………………………...91 Figure 2.4. Swim speeds of young and aged rats during training trial and probe trial conditions………………………………………………………………………...92 CHAPTER III Figure 3.1. Morris water maze performance in young and aged rats……………..………...105 Figure 3.2. Quantitative measures of cholinergic neurons and activated microglia in young and aged rats within the MS/VDB……………………...……………….106 Figure 3.S1. Bright-field photomicrographs depicting ChAT- and CD68-immunostaining in the MS/VDB of young and aged rats…………………………………..……107 Figure 3.S2. Density of ChAT+ cells thorough rostro-caudal sampling distribution……...…109 CHAPTER IV Figure 4.1. Spatial learning in young and aged rats………………………...………………138 Figure 4.2. Baclofen-stimulated GTP-binding in the hippocampus of young and aged rats…………………………………………………...……………………139 x Figure 4.3. Baclofen-stimulated GTP-binding in the prefrontal cortex of young and aged rats…………………………………………...……………………………140 Figure 4.4. GABABR1 and GABABR2 protein levels in hippocampus of young and aged rats…………………………………………………...……………………141 Figure 4.5. GABABR1 and GABABR2 protein levels in the prefrontal cortex of young and aged rats……………………………………………………………………142 Figure 4.6. Ratios of GABABR1a:GABABR1b across PFC and hippocampus of young and aged rats……………………………………………………………………143 CHAPTER V Figure 5.1. Performance of young