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Characterization of Hypothalamic Neuropeptides in Mammalian Reproduction

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Characterization of Hypothalamic Neuropeptides in Mammalian Reproduction University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Doctoral Dissertations Graduate School 12-2015 CHARACTERIZATION OF HYPOTHALAMIC NEUROPEPTIDES IN MAMMALIAN REPRODUCTION Valeria Tanco University of Tennessee - Knoxville, [email protected] Follow this and additional works at: https://trace.tennessee.edu/utk_graddiss Part of the Veterinary Medicine Commons Recommended Citation Tanco, Valeria, "CHARACTERIZATION OF HYPOTHALAMIC NEUROPEPTIDES IN MAMMALIAN REPRODUCTION. " PhD diss., University of Tennessee, 2015. https://trace.tennessee.edu/utk_graddiss/3614 This Dissertation is brought to you for free and open access by the Graduate School at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Doctoral Dissertations by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. To the Graduate Council: I am submitting herewith a dissertation written by Valeria Tanco entitled "CHARACTERIZATION OF HYPOTHALAMIC NEUROPEPTIDES IN MAMMALIAN REPRODUCTION." I have examined the final electronic copy of this dissertation for form and content and recommend that it be accepted in partial fulfillment of the equirr ements for the degree of Doctor of Philosophy, with a major in Comparative and Experimental Medicine. Brian K Whitlock, Major Professor We have read this dissertation and recommend its acceptance: Cheryl Kojima, Joe Bartges, Lanett Edwards Accepted for the Council: Carolyn R. Hodges Vice Provost and Dean of the Graduate School (Original signatures are on file with official studentecor r ds.) CHARACTERIZATION OF HYPOTHALAMIC NEUROPEPTIDES IN MAMMALIAN REPRODUCTION A Dissertation Presented for the Doctor of Philosophy Degree The University of Tennessee, Knoxville Valeria Tanco December 2015 “ANYONE WHO CONDUCTS AN ARGUMENT BY APPEALING TO AUTHORITY IS NOT USING INTELLIGENCE; HE IS JUST USING MEMORY.” - LEONARDO DA VINCI ii DEDICATION This thesis is dedicated to the two most important people in my life, my wife Sophy for her unwavering love and support; and to our daughter Emilia, for changing the essence of my being for the better…every day. I would also like to dedicate this thesis to my parents for teaching me perseverance and what the unconditional love of a parent to a child looks like. To you, thank you. I couldn’t have made it this far without you. iii ACKNOWLEDGEMENTS First, I would like to acknowledge my mentor, Dr Brian Whitlock for believing in me and graciously accepting me as his student three years ago. Second, I would like to thank my committee members Drs Joe Bartges, Cheryl Kojima and Lannett Edwards for their knowledge and support. I would also like to thank the University of Tennessee and the College of Veterinary Medicine for their financial support through this program and the chance to finish my studies as I worked in the clinic. Last but not least, I’d like to thank the people without whom I wouldn’t have been able to carry out my projects or had as much fun during each one as I did: Charlie, Tate, Lisa, Tammy and my summer students for their endless nights at the farm and Becca for all her help carrying out our LH assays. To each and every one of you, thank you. iv ABSTRACT Understanding of regulation of reproduction at the level of the brain changed dramatically with the discovery of reproductive neuropeptides. To date there have been no studies to characterize the distribution of reproductive neuropeptides in the bovine hypothalamus at different stages of the estrus cycle or to determine the physiological effects of peripheral administration of Gonadotropin inhibiting hormone (GnIH) in intact female cattle and ovariohysterectomized bitches. The goal of the first study was to determine distribution and connectivity of kisspeptin, dynorphin, and GnIH in the hypothalami of sexually mature female cattle during the estrous cycle. To this end, hypothalami of female cattle were collected during periestrus and diestrus. The neuroanatomical distribution, synaptic connectivity, and response to different circulating progesterone concentrations suggest these neuropeptides play a pivotal role in the regulation of reproduction in cyclic cattle. Another goal was to test the hypothesis that IV administration of GnIH would decrease serum LH concentrations in post-pubertal heifers. Two studies were carried out to this end. The objective of the first study was to determine whether IV GnIH administration would decrease basal serum LH concentrations. The objective of the second study was to determine whether continuous IV administration of GnIH during the expected time of the LH surge had an effect on serum LH concentrations and ovulation. Results suggest that exogenous administration of GnIH decreases basal LH concentrations but is not able to suppress the surge release of LH or ovulation in post-pubertal heifers. Finally, the objective of the third study was to determine the effects of IV administration of GnIH on serum LH concentrations in ovariohysterectomized bitches as a potential alternative to surgical sterilization. Results suggest that IV administration of GnIH is not able to suppress serum LH concentrations in the ovariohysterectomized bitch. v In summary, expression of kisspeptin, dynorphin, and GnIH changes with progesterone concentrations in sexually mature female cattle. In addition, exogenous administration of GnIH affects basal plasma concentrations of LH but not LH surge characteristics in sexually mature female cattle. It also appears that exogenous administration of GnIH does not affect plasma LH concentrations in ovariohysterectomized adult bitches. vi TABLE OF CONTENTS CHAPTER 1 GENERAL INTRODUCTION .......................................................................................... 1 CHAPTER 2 LITERATURE REVIEW .................................................................................................. 5 2.1 HYPOTHALAMIC-PITUITARY ANATOMY ........................................................................................ 6 2.2 HYPOTHALAMIC-PITUITARY PHYSIOLOGY ................................................................................... 8 2.2.1 REGULATION OF THE REPRODUCTIVE CYCLE BY THE HYPOTHALAMIC-PITUITARY-GONADAL AXIS (HPG AXIS) .............................................................................................................................. 8 Bovine estrus cycle .................................................................................................................... 9 Canine estrus cycle .................................................................................................................. 11 2.3 HYPOTHALAMIC NEUROPEPTIDES ............................................................................................... 15 2.3.1 GONADOTROPIN INHIBITING HORMONE ............................................................................... 16 2.3.1.1 Discovery..................................................................................................................... 16 2.3.1.2 Cells, fibers and receptor localization ......................................................................... 17 2.3.1.3 Regulation of reproduction by GnIH ........................................................................... 19 Role of GnIH on gonadotropin secretion .............................................................................. 20 Role of GnIH in seasonality.................................................................................................. 23 2.3.1.4 Other physiological roles of GnIH .............................................................................. 25 Role of GnIH in metabolic state related effects on reproduction ......................................... 25 Role of GnIH in stress related effects on reproduction ......................................................... 27 2.3.2 OTHER HYPOTHALAMIC NEUROPEPTIDES ............................................................................. 29 2.3.2.1 Kisspeptin .................................................................................................................... 29 2.3.2.2 Neurokinin B ............................................................................................................... 34 vii 2.3.2.3 Dynorphin .................................................................................................................... 35 2.4 NEUROPEPTIDES AND GNRH PULSE CREATING MECHANISM HYPOTHESIS ................................ 37 2.5 APPENDIX ....................................................................................................................................... 40 CHAPTER 3 OBJECTIVES .................................................................................................................... 41 CHAPTER 4 DISTRIBUTION AND REGULATION OF GONADOTROPIN-RELEASING HORMONE, KISSPEPTIN, GONADOTROPIN INHIBITING HORMONE AND DYNORPHIN IN THE BOVINE HYPOTHALAMUS .................................................................................................. 43 4.1 ABSTRACT ...................................................................................................................................... 44 4.2 INTRODUCTION .............................................................................................................................. 44 4.3 MATERIAL AND METHODS ............................................................................................................ 47 4.3.1 ETHICS
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