SHORT COURSE 1: Sex Differences in the Brain: Balancing Sex in Preclinical Research Organized by Jill Becker, PhD, and Jessica Tollkuhn, PhDNEW COVER TK Short Course 1 Sex Differences in the Brain: Balancing Sex in Preclinical Research Organized by Jill Becker, PhD, and Jessica Tollkuhn, PhD Please cite articles using the model: [AUTHOR’S LAST NAME, AUTHOR’S FIRST & MIDDLE INITIALS] (2018) [CHAPTER TITLE] Sex Differences in the Brain: Balancing Sex in Preclinical Research. (Becker J, Tollkuhn J, eds) pp. [xx-xx]. Washington, DC : Society for Neuroscience. All articles and their graphics are under the copyright of their respective authors. Cover graphics and design © 2018 Society for Neuroscience. SHORT COURSE 1 Sex Differences in the Brain: Balancing Sex in Preclinical Research Organized by Jill Becker, PhD and Jessica Tollkuhn, PhD Friday, November 2, 2018 8 a.m.–6 p.m. Location: San Diego Convention Center • Room: 6B TIME TOPIC SPEAKER 7:30–8 a.m. CHECK-IN Jill Becker, PhD • University of Michigan 8–8:10 a.m. Opening Remarks Jessica Tollkuhn, PhD • Cold Spring Harbor Laboratory 8:10–9 a.m. Developmental Origins of Sex Differences in the Brain Margaret M. McCarthy, PhD • University of Maryland 9–9:30 a.m. The Stress Response: Sex-Specific Neural Mechanisms Debra Bangasser, PhD • Temple University 9:30–10 a.m. Behavior Paradigms I: Sex-Specific Responses Rebecca Shansky, PhD • Northeastern University 10–10:30 a.m. BREAK 10:30–11:20 a.m. Sex-Dependent Mechanisms of Synaptic Modulation Catherine Woolley, PhD • Northwestern University 11:20–11:50 a.m. Behavior Paradigms II: Reward and Affective Behaviors Laura O’Dell, PhD • University of Texas, El Paso 11:50 a.m.–12:20 p.m. Dissecting Sex-Typical Neural Circuitry Jessica Tollkuhn, PhD • Cold Spring Harbor Laboratory 12:20–1:20 p.m. LUNCH — ROOM 33 1:20–2:10 p.m. Adolescence and Puberty Deena Walker, PhD • Icahn School of Medicine at Mount Sinai 2:10–3 p.m. The Importance of Estradiol as a Neuroprotectant in Both Sexes Colin Saldanha, PhD • American University 3–3:50 p.m. How to Study Female and Male Rodents — Takeaway Lessons Jill Becker, PhD • University of Michigan 3:50–4 p.m. Summary and Introduction of Breakout Sessions Jill Becker, PhD and Jessica Tollkuhn, PhD 4–4:15 p.m. BREAK AFTERNOON BREAKOUT SESSIONS • PARTICIPANTS SELECT FIRST DISCUSSION GROUPS AT 4:15 P.M. TIME BREAKOUT SESSIONS SPEAKERS ROOM Debra Bangasser, PhD and Rebecca 4:15–5 p.m. Group 1: 29A Stress, Learning, and Vulnerability Shansky, PhD Group 2: Reward Circuitry and Addiction: Differential Effects of Drugs of Jill Becker, PhD and Laura O’Dell, PhD 29B Abuse in Females and Males Margaret M. McCarthy, PhD and Colin Group 3: 29C Neuroimmune Interactions: The Role of Microglia Saldanha, PhD Group 4: Deena Walker, PhD and Jessica Sex Differences in Gene Expression: Contributions of Steroid 29D Tollkuhn, PhD Hormones and Sex Chromosomes 5–5:15 p.m. BREAK AFTERNOON BREAKOUT SESSIONS • PARTICIPANTS SELECT SECOND DISCUSSION GROUPS AT 5:15 P.M. 5:15–6 p.m. Same as sessions above over Table of Contents Agenda .........................................................................3 Introduction .....................................................................5 How to Study Female and Male Rodents Jill B. Becker, PhD ..............................................................6 Genetic and Neural Circuit Approaches to Studying Sex Differences Jessica Tollkuhn, PhD ...........................................................17 How to Study the Origins of Sex Differences in Brain and Behavior Margaret M. McCarthy, PhD .....................................................29 Adolescence and Reward: Making Sense of Neural and Behavioral Changes Amid the Chaos Deena M. Walker, PhD, Margaret R. Bell, PhD, Cecilia Flores, PhD, Joshua Gulley, PhD, Jari Willing, PhD, and Matthew J. Paul, PhD .....................40 Sex Differences in Behavioral Strategies: Avoiding Interpretational Pitfalls Rebecca M. Shansky, PhD. 53 Sex Differences and the Role of Ovarian Hormones in Modulating the Behavioral Effects of Nicotine in Rodent Models Rodolfo J. Flores, MA, Bryan Cruz, BA, Kevin P. Uribe, MS, Luis M. Carcoba, MD, PhD, and Laura E. O’Dell, PhD ................................59 Neuroinflammation and Neurosteroidogenesis: Reciprocal Modulation During Injury to the Adult Zebra Finch Brain Alyssa L. Pedersen, PhD, Jenna L. Brownrout, BA, and Colin J. Saldanha, PhD. .68 Sex-Dependent Mechanisms of Synaptic Modulation Catherine S. Woolley, PhD .......................................................78 The Stress Response: Sex-Specific Neural Mechanisms Debra A. Bangasser, PhD, and Kimberly R. Wiersielis, PhD ..............................86 Introduction Many, if not most, neurological and psychiatric disorders show a sex bias in incidence, etiology, or age of onset. Yet most neuroscientists still perform their experiments in a single sex, usually males. To address this discrepancy, the National Institutes of Health now mandates that researchers consider “sex as a biological variable” in their study design. It is therefore crucial for scientists to possess a sound understanding of how sex differences in the brain manifest and how these differences can influence development, brain function, and disease susceptibility. In addition to contributions from sex chromosomes, sex differences in the vertebrate brain are regulated largely by steroid hormones that originate both in the gonads and locally in the brain. These hormones act in early life to specify sex-typical neural circuitry and, following puberty, to modulate neural activity and signaling pathways within the sexually differentiated brain. Recent work has made significant progress toward illuminating sex differences in sophisticated behavior paradigms, mapping the circuitry that mediates sex-typical behavioral responses, and defining molecular programs that both instruct and reflect these differences. The study of sex differences is now rapidly expanding beyond the realm of a specialized field to emerge as a principle consideration in the design of reproducible, clinically relevant research. This course is designed to enable neuroscientists to incorporate both sexes into their preclinical research. Participants will become familiar with molecular, neural circuit, and behavioral differences between the sexes, with a focus on rodents. Leading experts will review fundamental concepts and the latest discoveries, including the developmental origins of sex differences, gonadal steroid hormones, stress and vulnerability, reward and affective behaviors, and adolescence and puberty. Experimental design considerations and statistical analyses will also be discussed. How to Study Female and Male Rodents Jill B. Becker, PhD Molecular and Behavioral Neuroscience Institute Department of Psychology University of Michigan Ann Arbor, Michigan © 2018 Becker How to Study Female and Male Rodents 7 Introduction Becker, 2016). When a sex difference is found, some NOTES This chapter discusses how to think about and investigators will want to determine more about the determine the appropriate manipulations and neurobiological processes that are responsible for the procedures for investigating sex differences in, and differences. the effects of gonadal hormones on, experimental outcomes in adult rats and mice. I will also discuss Effect of Gonadal Hormones on estrous cycles, surgical procedures, and hormone a Trait treatments. I will conclude with a discussion of One of the next questions that will arise is whether variability and statistical methods that can be used gonadal hormones have an effect on the trait. Two to minimize animal numbers when adding sex as a approaches can help determine whether this is biological variable to your research. the case. One can examine whether the female’s behavior varies with the estrous cycle. Alternatively, What Is a Sex Difference? one can remove the gonads by ovariectomy (OVX) The first question researchers usually ask is whether or castration (CAST) and then selectively replace there is a sex difference in a trait. The answer to this hormones. We will address the estrous cycle first. question is not a simple “yes” or “no”; it turns out to be more complicated. As illustrated in Figure 1A, Determining Estrous Cycle Stages males and females can exhibit different traits, as is The estrous cycle is the product of the hypothalamic- true for reproduction. For many traits, however, both pituitary-gonadal (HPG) axis that results in a cyclic females and males exhibit the trait, but there are release of ovarian hormones, ovulation, and sexual differences in how it is expressed (Figs. 1B,C) or the receptivity. It is analagous to the human menstrual mechanisms that mediate it (Fig. 1D) (Becker et al., cycle, except that in rats and mice, the cycle is much 2016; Becker and Koob, 2016; Sanchis-Segura and Figure 1. Four types of sex differences that can be observed: qualitative, quantitative, population, and mechanistic, also referred to as compensatory, divergent, or latent sex differences. Reprinted with permission from Becker and Koob (2016), Fig. 1. Copy- right 2016, American Society for Pharmacology and Experimental Therapeutics. © 2018 Becker How to Study Female and Male Rodents 8 NOTES Figure 2. Patterns of estradiol, progesterone, and luteinizing hormone (LH) in the female human (A) and rat (B) during the reproductive cycle. Time unit of the x-axis in A is days; in B, x-axis is hours. B, Dark bars, Dark period of the day/night cycle. Note that during the follicular