ABSTRACT Title of Document: CHARACTERIZATION AND CONTROL OF AGGRESSION AND REPRODUCTION IN THE MALE CLOUDED LEOPARD (NEOFELIS NEBULOSA) Heather Burton DeCaluwe, Doctor of Philosophy, 2012 Directed By: Professor Mary Ann Ottinger, Department of Animal and Avian Sciences Clouded leopards are a striking and elusive cat species whose secretive nature has made it difficult to gather information on population statistics and behavior in the wild, where the population is in decline. While captive populations are intended as a hedge against extinction, breeding clouded leopards ex situ has been a challenge, primarily due to extreme male aggression toward females. Despite the importance of aggression in this species, there has as yet been no systematic study characterizing the basis of aggressive episodes. Two mechanisms seem to underlie the aggressive behavior in clouded leopards: degree of anxiety and circulating testosterone levels. Three studies were conducted to characterize mechanisms modulating aggression in male clouded leopards. In Study 1, sixteen adult male clouded leopards were categorized as ‘anxious’ or ‘calm’ using a keeper questionnaire and fecal endocrine (androgen and glucocorticoid) profiles; these measures were correlated with behavior rates and frequencies before, during, and after a series of behavioral reaction tests aimed at assessing an individual’s response to stress-inducing situations. In Study 2, the behavioral and endocrine responses to the same tests were compared in the same clouded leopards following three treatments: 1) an anxiety- reducing psychotropic drug (clomipramine, n = 4); 2) a gonadotropin releasing hormone agonist (deslorelin, n = 5), or 3) no treatment (n = 4). In Study 3, the long- term effects of the drug treatments on spermatogenesis and hormone concentrations were compared in clouded leopards (n = 2/treatment) and domestic cats (n = 5/treatment), a model for non-domestic felid reproduction. Studies revealed important findings about the basis of aggressive behavior in male clouded leopards. First, two of the behavioral reaction tests – ‘mirror image stimulation’ and ‘unfamiliar people’ – were effective tools for evaluating temperament and eliciting a behavioral response. Second, treatment with both clomipramine and deslorelin reduced anxious and aggressive behaviors (e.g. ‘tail flicking’ and ‘growling’) indicating multiple physiological mechanisms likely modulate aggression in this species. Finally, deslorelin temporarily suppressed hormone concentrations and reproductive function, while clomipramine had no clear effect on either. Ultimately, this information provides important tools for improving male-female pairing success and the overall management of captive clouded leopards. CHARACTERIZATION AND CONTROL OF AGGRESSION AND REPRODUCTION IN THE MALE CLOUDED LEOPARD (NEOFELIS NEBULOSA) By Heather Burton DeCaluwe Dissertation submitted to the Faculty of the Graduate School of the University of Maryland, College Park, in partial fulfillment of the requirements for the degree of Doctor of Philosophy 2012 Advisory Committee: Professor Mary Ann Ottinger, Chair Professor Amy O. Burk Professor Eric S. Haag Dr. Katharine M. Pelican Dr. Budhan S. Pukazhenthi © Copyright by Heather Burton DeCaluwe 2012 Dedication For my beloved husband, Steven And In memory of Dr. JoGayle Howard ii Acknowledgements First, I would like to express my deepest gratitude to my advisors, Dr. Mary Ann Ottinger, Dr. Katey Pelican, and Dr. JoGayle Howard. Together they made the perfect team of advisors! Their support, guidance, expertise, dedication, and encouragement were invaluable; I couldn’t have made it to this point without them. A special thanks to the other members of my committee for their time and attention and, importantly, for being the five (out of nine) to make it to the end – Dr. Eric Haag, Dr. Budhan Pukazhenthi, and Dr. Amy Burk. My deepest gratitude to Dr. Nadja Wielebnowski, an honorary committee member, for her enthusiasm, expertise, and guidance from the very beginning. When I asked for her help she said, “of course, it’s for clouded leopards, right?” Words cannot express how grateful I am to the many many zookeepers, managers, veterinarians, scientists, support staff, and interns at the 12 participating zoological institutions who graciously donated their time and effort to work with us on this research. I will try to name many here and for those I forget, you are not truly forgotten, only momentarily in my exhausted state: Veronica Acosta, Bonnie Breitbeil, Laura Carpenter, Dr. Pierre Comizzoli, Deb Copeland, Christie Eddie, Jilian Fazio, Norah Fletchall, Beth Graham, Kenn Harwood, Dr. Katharine Hope, Kally Koester, Jessica Kordell, Ken Lang, Lisa Laskoski, Elena Less, Bethany Lorentz, Cathi Morrison, Dr. Luis Padilla, Nikki Presley, Sarah Putman, Sara Riger, Jenny Santiestevan, Teresa Shepard, Tracy Sorensen, Dr. Kimberly Terrell, Laurie iii Trechsel, Lisa Ware, Rain Westgard, Drew White, Dr. Megan Wilson, and Michelle Witek. I am eternally grateful to my colleagues and mentors within the Society for Conservation Biology and Smithsonian Conservation Biology Institute, for their support, understanding, inspiration, and most importantly, dedication to conservation – when things got really tough, they helped me remember why it was all worth it! And to my family – I truly could not have accomplished this without you. Dad, you are always in my memory, I know you would be so proud. Mom, your love and support over the past 34 years have made me who I am today. Jenny and Devin, you are like sisters to me, what more can I say? And Steven, you are my rock; you bring constant joy, love, and laughter to my life. Finally, I would like to thank the Conservation Endowment Fund, managed by The Association of Zoos and Aquariums (AZA) - grant #08-801 - for providing financial support for this project. I would also like to acknowledge the partial financial support from University of Maryland, University of Minnesota, and Smithsonian Women’s Committee. iv Table of Contents Dedication..................................................................................................................... ii Acknowledgements......................................................................................................iii Table of Contents.......................................................................................................... v List of Tables .............................................................................................................viii List of Figures.............................................................................................................. ix List of Figures.............................................................................................................. ix Chapter 1: Introduction................................................................................................. 1 1.1 Overview of Study .............................................................................................. 1 1.2 Hypothesis and Objectives.................................................................................. 3 1.3 Background and Significance ............................................................................. 4 1.3.1 Clouded Leopard Natural History................................................................ 4 1.3.2 Neural Pathways and Aggression ................................................................ 6 1.3.3 Testosterone and Aggression ....................................................................... 7 1.3.4 Temperament and Aggression ..................................................................... 9 1.3.5 Stress-Sensitivity and Aggression................................................................ 9 1.3.6 Spermatogenesis and Reproductive Function............................................ 10 1.3.7 Testosterone and Reproductive Function................................................... 11 1.3.8 The Affect of Stress on Reproductive Function ........................................ 12 1.3.9 Psychotropic Drugs, Anxiety and Aggression, and Reproductive Function ............................................................................................................................. 13 1.3.10 GnRH Agonists, Testosterone and Aggression, and Reproductive Function .............................................................................................................. 14 1.4 Summary........................................................................................................... 15 Chapter 2: Behavioral Reaction Tests as a Measure of Stress Reactivity to Determine Temperament in the Male Clouded Leopard (Neofelis nebulosa).............................. 16 2.1 Introduction....................................................................................................... 16 2.2 Methods............................................................................................................. 21 2.2.1 Animals and Study Area ............................................................................ 21 2.2.2 Study Design.............................................................................................. 22 2.2.3 Behavioral Questionnaire........................................................................... 23 2.2.4 Behavioral Reaction Tests ......................................................................... 23 2.2.5 Videotaping and Analysis.........................................................................