Abstract
Neurons in the hypothalamic arcuate nucleus (ARC) that co-express kisspeptin, neurokinin B and dynorphin (KNDy cells) are essential for mammalian reproduction as key regulators of gonadotropin-releasing hormone (GnRH) secretion. Although multiple endogenous and exogenous signals act indirectly via KNDy neurons to regulate GnRH, the identity of upstream neurons that provide synaptic input to this subpopulation is unclear. We used rabies-mediated tract-tracing in transgenic Kiss1-Cre mice combined with whole-brain optical clearing and multiple-label immunofluorescence to create a comprehensive and quantitative brain-wide map of neurons providing monosynaptic input to KNDy cells, as well as identify the estrogen receptor content and peptidergic phenotype of afferents. Over 90% of monosynaptic input to KNDy neurons originated from hypothalamic nuclei in both male and female mice. The greatest input arose from non-KNDy ARC neurons, including proopiomelanocortin-expressing cells. Significant female-dominant sex differences in afferent input were detected from estrogen-sensitive hypothalamic nuclei critical for reproductive endocrine function and sexual behavior in mice, indicating KNDy cells may provide a unique site for the coordination of sex-specific behavior and gonadotropin release. These data provide key insight into the structural framework underlying the ability of KNDy neurons to integrate endogenous and environmental signals important for the regulation of reproductive function.
Highlights
Reproductive capacity is dependent on the pulsatile secretion of the gonadotrophin luteinizing hormone (LH), which is driven by the pulsatile release of gonadotrophin-releasing hormone (GnRH) from gonadotropinreleasing hormone (GnRH) neurons in the hypothalamus[1]
As the KNDy peptides are difficult to visualize in the arcuate nucleus (ARC) of mice using immunolabelling, the percentage of Cre-positive kisspeptin cells transfected by AAV-TVA/GFP was first determined through injection of AAV vectors into the ARC of Kiss1-Cre+/−/tdTomato+/− reporter mice (n = 3 male, n = 3 female)
This study suggests that KNDy neurons receive direct synaptic input from a remarkably diverse number of afferent populations throughout the brain, which may form the anatomical basis for the control of gonadotropin secretion over different reproductive and non-reproductive states
Summary
Reproductive capacity is dependent on the pulsatile secretion of the gonadotrophin luteinizing hormone (LH), which is driven by the pulsatile release of gonadotrophin-releasing hormone (GnRH) from GnRH neurons in the hypothalamus[1]. One upstream population of cells in the arcuate nucleus (ARC) that co-express the neuropeptides kisspeptin, neurokinin B and dynorphin, known as KNDy cells, are proposed to form the long elusive GnRH/LH pulse generator[9] As such, they provide an ideal nodal point through which peripheral signals can alter gonadotropin secretion and reproductive capacity. Kisspeptin-specific deletion of the insulin[36] or leptin[37] receptor does not affect puberty onset and reproductive capacity in mice, indicating transmission through upstream populations Consistent with this hypothesis, KNDy neurons express receptors for neurochemicals that regulate gonadotropin release using indirect circuits. Using a shorter transfection period to avoid toxicity induced by the rabies virus, we determined peptidergic phenotypes of afferent populations to KNDy cells and identified subpopulations that respond to steroid hormone signals
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