Sex differences in the cellular differentiation of the developing brain when paired with altered development can lead to abnormal circuitries that may be associated with sex differences in susceptibility to major mental disorders. We are determining mechanisms in development by which male and female brains differ and where and how genetic and hormonal factors interact to result in brain sexual differentiation. Cells of the mammalian hypothalamus are strongly affected by gonadal steroid hormone signals during development. Over the last 15 years a better understanding of direct genetic influences on brain sexual differentiation have emerged. This proposal concentrates on the formation of the hypothalamus (HYP) as a key target for studying this process. The HYP is important for regulating homeostatic, neuroendocrine, and behavioral functions. It is a region where hormones dramatically influence development and where hormone-concentrating cells regulate physiology and behavior. We have established methods that render developmental processes accessible to live observation, and direct manipulation in vitro. We will raise SF-1 knockout (KO) mice to adulthood to assess consequences of brain sexual differentiation without endogenous gonadal steroids. Factors that regulate the arrangement of cells in subcortical brain regions are poorly understood. This proposal centers on two fundamental questions: 1. do hormones influence the position of neurons during development, and 2. What are the consequences of having hormone responsive cells moved to inappropriate positions? Two aims focus on mechanisms during development and differentiation, and a third focuses on the adult consequences: First we ask how are sex differences in the positions of cells in the embryonic HYP regulated by gonadal steroids? We will take advantage of genetically disrupted mice (SF-1) in which fetal steroid synthesis is disturbed to test the hypothesis that sex differences in the distribution of cells in the HYP depend upon specific receptor subtypes. We will take advantage of the same mice to determine the long term consequences of moving hormone responsive cells to different places in the region of the ventromedial nucleus of the hypothalamus (VMH) as part of our third aim. We will utilize live video microscopy in slices taken from mice with cells that are fluorescent under the control of specific promoters driving green or yellow fluorescent protein expression. Second, we will ask how does nitric oxide (NO) generation contributes to the development of the HYP by influencing the characteristics of cell movement or survival and what is the pattern of live NO exposure? The embryonic HYP is rich in neuronal nitric oxide synthase (nNOS). We will use newly available NO donors to help directly test the hypothesis that NO influences cell behavior. We will trace the real time production, release and diffusion of NO using a novel electrochemical biosensor array that will provide for 'chemical vision' in live slices. Finally, we will ask what is the role(s) of steroid hormones during specific periods of development for setting behavioral potentials? Our data shows that SF-1 knockout mice still develop some sex differences in brain and behavior even though they are not exposed to endogenous gonadal steroids during development. We will use SF-1 KO male and female mice and their littermate controls as model animals for examining social and anxiety-like behaviors after developmental treatment with exogenous steroid hormones at different periods during development. This will provide a test of the function of particular critical periods for the differentiation of brain and behavior on a null hormonal background. Brains from adult animals will be examined by histological, immunohistochemical and in situ hybridization procedures to determine sex differences in HYP structure, outputs, and gene expression.

Public Health Relevance

This proposal addresses a major issue of sex differences in brain development. A number of diseases and disorders arise with significant sex-dependent bias ranging from purely physiological to largely psychological. We have established methods that render developmental processes accessible to live observation, and direct manipulation in vitro and developed methods to raise SF-1 knockout (KO) mice to adulthood to assess consequences of brain sexual differentiation without endogenous gonadal steroids. The plan marshals an array of new and evolving technologies to attack the problem of brain sexual differentiation in vivo and in vitro. By the use of converging approaches - and a research team that is practiced in communicating around the world, we have the opportunity to transform our understanding of sex biases among major health issues.

National Institute of Health (NIH)
National Institute of Mental Health (NIMH)
Research Project (R01)
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Special Emphasis Panel (ZRG1-IFCN-Z (02))
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Panchision, David M
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Colorado State University-Fort Collins
Veterinary Sciences
Schools of Veterinary Medicine
Fort Collins
United States
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Büdefeld, Tomaz; Tobet, Stuart A; Majdic, Gregor (2016) The Influence of Gonadal Steroid Hormones on Immunoreactive Kisspeptin in the Preoptic Area and Arcuate Nucleus of Developing Agonadal Mice with a Genetic Disruption of Steroidogenic Factor 1. Neuroendocrinology 103:248-58
Spanic, Tanja; Fabjan, Teja; Majdic, Gregor (2015) Expression levels of mRNA for neurosteroidogenic enzymes 17?-HSD, 5?-reductase, 3?-HSD and cytochrome P450 aromatase in the fetal wild type and SF-1 knockout mouse brain. Endocr Res 40:44-8
Babwah, Andy V; Navarro, Víctor M; Ahow, Maryse et al. (2015) GnRH Neuron-Specific Ablation of G?q/11 Results in Only Partial Inactivation of the Neuroendocrine-Reproductive Axis in Both Male and Female Mice: In Vivo Evidence for Kiss1r-Coupled G?q/11-Independent GnRH Secretion. J Neurosci 35:12903-16
Kercmar, Jasmina; Tobet, Stuart A; Majdic, Gregor (2014) Social isolation during puberty affects female sexual behavior in mice. Front Behav Neurosci 8:337
Ahow, Maryse; Min, Le; Pampillo, Macarena et al. (2014) KISS1R signals independently of G?q/11 and triggers LH secretion via the ?-arrestin pathway in the male mouse. Endocrinology 155:4433-46
Kercmar, Jasmina; Snoj, Tomaz; Tobet, Stuart A et al. (2014) Gonadectomy prior to puberty decreases normal parental behavior in adult mice. Horm Behav 66:667-73
Knoll, John Gabriel; Clay, Colin M; Bouma, Gerrit J et al. (2013) Developmental profile and sexually dimorphic expression of kiss1 and kiss1r in the fetal mouse brain. Front Endocrinol (Lausanne) 4:140
Grgurevic, Neza; Budefeld, Tomaz; Spanic, Tanja et al. (2012) Evidence that sex chromosome genes affect sexual differentiation of female sexual behavior. Horm Behav 61:719-24
Nugent, B M; Tobet, S A; Lara, H E et al. (2012) Hormonal programming across the lifespan. Horm Metab Res 44:577-86
Majdic, Gregor; Tobet, Stuart (2011) Cooperation of sex chromosomal genes and endocrine influences for hypothalamic sexual differentiation. Front Neuroendocrinol 32:137-45

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