Virginia Commonwealth University VCU Scholars Compass Theses and Dissertations Graduate School 2013 Behavioral and genomic characterization of scheduled ethanol deprivation Jonathan Warner Virginia Commonwealth University Follow this and additional works at: https://scholarscompass.vcu.edu/etd Part of the Medical Pharmacology Commons © The Author Downloaded from https://scholarscompass.vcu.edu/etd/3264 This Dissertation is brought to you for free and open access by the Graduate School at VCU Scholars Compass. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of VCU Scholars Compass. For more information, please contact [email protected]. Behavioral and genomic characterization of scheduled ethanol deprivation A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University by Jonathan Andrew Warner Bachelor of Science, Virginia Commonwealth University, 2007 Director: Michael F. Miles, M.D., Ph.D. Professor, Department of Pharmacology and Toxicology Virginia Commonwealth University Richmond, Virginia November 8th, 2013 II Acknowledgments I would like to sincerely thank my advisor, Michael Miles, for his heroic patience and invaluable guidance throughout my graduate career. I learned an incredible amount during my time under his tutelage, and I say without reservation that I could not have had a better mentor. I feel very lucky to have found my way to Dr. Miles’ laboratory. I would also like to thank my graduate committee for their suggestions and input along the way, and for sitting through my presentations. I would like to thank my mother, father, and sister for all of their support and assistance. Without them this time would have been far more difficult, and I might not have made it this far. I would also like to thank my friends, who have kept me sane. I express my sincere gratitude to Dr. Yan Zhang and Dr. Yunyun Yuan, who gave me the opportunity to study their compound, NAQ. Finally, I would also like to thank the past and present members of the Miles Laboratory, all of whom have contributed in some way to the success of my project. III Table of Contents Acknowledgments........................................................................................................................... ii List of Tables .................................................................................................................................. v List of Figures ............................................................................................................................... vii Abstract ......................................................................................................................................... vii Chapter 1. Introduction ................................................................................................................... 1 Chapter 2. Background and significance ........................................................................................ 8 Alcoholism and alcohol use disorders ......................................................................................... 8 Neurobiology and treatment of alcoholism in humans ............................................................. 10 Animal models of excessive alcohol consumption ................................................................... 13 Chronic intermittent ethanol vapor (CIE) ................................................................................. 14 Intermittent alcohol access (IAA) ............................................................................................. 16 Influence of age, genetics, and environment on IAA ................................................................ 19 Neurobiology of Intermittent Alcohol Access .......................................................................... 24 Chapter 3. Genomic analysis of the alcohol deprivation effect identifies alterations in mesocorticolimbic gene expression functioning in neuroplasticity .............................................. 39 Introduction ............................................................................................................................... 39 Materials and Methods .............................................................................................................. 41 Results ....................................................................................................................................... 45 Discussion ................................................................................................................................. 67 Chapter 4. Intermittent alcohol access elevates ethanol intake in C57BL/6J and C57BL/6NCrl mice ............................................................................................................................................... 82 Introduction ............................................................................................................................... 82 Materials and Methods .............................................................................................................. 86 Results ....................................................................................................................................... 90 Discussion ............................................................................................................................... 114 Chapter 5. Nucleus accumbens transcriptional regulation in the intermittent alcohol access model: differential expression and weighted gene co-expression networks ............................... 120 IV Introduction ............................................................................................................................. 120 Materials and Methods ............................................................................................................ 124 Results ..................................................................................................................................... 131 Discussion ............................................................................................................................... 155 Chapter 6. Mu-opioid receptor-selective antagonist NAQ selectively reduces high concentration alcohol consumption and delays the effects of intermittent alcohol access ................................ 170 Introduction ............................................................................................................................. 170 Materials and Methods ............................................................................................................ 174 Results ..................................................................................................................................... 177 Discussion ............................................................................................................................... 187 Chapter 7. General discussion, limitations, and future directions ............................................. 194 Alcohol deprivation effect studies........................................................................................... 194 Intermittent alcohol access ...................................................................................................... 197 Limitations .............................................................................................................................. 207 Transcriptional regulation mechanisms in NAc during drinking and scheduled deprivation . 215 Other likely nucleus accumbens adaptations of note for future study .................................... 222 List of References ............................................................................................................................ i Appendix 1. Supplementary Materials Location ..................................................................... xxxix Vita ................................................................................................................................................. xl V List of Tables Table 3.1. Top 25 results in NAc from transcription factor binding site analysis performed with oPossum (http://burgundy.cmmt.ubc.ca/oPOSSUM/) in promoter regions +/- 2000 base-pairs from transcription start site for genes regulated by ethanol deprivation in nucleus accumbens, prefrontal cortex, and ventral midbrain of C57BL/6J mice. ......................................................... 60 Table 3.2. Top 20 candidate genes for deprivation-induced elevated ethanol consumption within and across brain regions based on ranking with ToppNet (http://toppgene.cchmc.org/) using 5 gene lists related to ethanol consumption as training sets ............................................................ 65 Table 5.1. Differential gene expression determined by multi-class LIMMA. ............................ 132 Table 5.2. Transcription factor binding site analysis in genes differentially expressed between IAA and CA nucleus accumbens. ............................................................................................... 135 Table 5.3. Significantly over-represented transcription factors with factor, binding protein, or direct mediator of action regulated across groups (LIMMA, F < .05 and/or adjusted p-value < .05) .............................................................................................................................................