Feeding neurons integrate metabolic and reproductive states in mice.

Balance between metabolic and reproductive processes is important for survival, particularly in mammals that gestate their young. How the nervous system coordinates this balance is an active area of study. Herein, we demonstrate that somatostatin (SST) neurons of the tuberal hypothalamus alter feeding in a manner sensitive to metabolic and reproductive states in mice. Whereas chemogenetic activation of SST neurons increased food intake across sexes, ablation decreased food intake only in female mice during proestrus. This ablation effect was only apparent in animals with low body mass. Fat transplantation and bioinformatics analysis of SST neuronal transcriptomes revealed white adipose as a key modulator of these effects. These studies indicate that SST hypothalamic neurons integrate metabolic and reproductive cues by responding to varying levels of circulating estrogens to modulate feeding differentially based on energy stores. Thus, gonadal steroid modulation of neuronal circuits can be context dependent and gated by metabolic status.

Feeding Neurons Integrate Metabolic and Reproductive States in Mice.

Trade-offs between metabolic and reproductive processes are important for survival, particularly in mammals that gestate their young. Puberty and reproduction, as energetically taxing life stages, are often gated by metabolic availability in animals with ovaries. How the nervous system coordinates these trade-offs is an active area of study. We identify somatostatin neurons of the tuberal nucleus (TNSST) as a node of the feeding circuit that alters feeding in a manner sensitive to metabolic and reproductive states in mice. Whereas chemogenetic activation of TNSST neurons increased food intake across sexes, selective ablation decreased food intake only in female mice during proestrus. Interestingly, this ablation effect was only apparent in animals with a low body mass. Fat transplantation and bioinformatics analysis of TNSST neuronal transcriptomes revealed white adipose as a key modulator of the effects of TNSST neurons on food intake. Together, these studies point to a mechanism whereby TNSST hypothalamic neurons modulate feeding by responding to varying levels of circulating estrogens differentially based on energy stores. This research provides insight into how neural circuits integrate reproductive and metabolic signals, and illustrates how gonadal steroid modulation of neuronal circuits can be context-dependent and gated by metabolic status.

Deletion of Androgen Receptor in LepRb Cells Improves Estrous Cycles in Prenatally Androgenized Mice.

Androgens are steroid hormones crucial for sexual differentiation of the brain and reproductive function. In excess, however, androgens may decrease fertility as observed in polycystic ovary syndrome, a common endocrine disorder characterized by oligo/anovulation and/or polycystic ovaries. Hyperandrogenism may also disrupt energy homeostasis, inducing higher central adiposity, insulin resistance, and glucose intolerance, which may exacerbate reproductive dysfunction. Androgens bind to androgen receptors (ARs), which are expressed in many reproductive and metabolic tissues, including brain sites that regulate the hypothalamo-pituitary-gonadal axis and energy homeostasis. The neuronal populations affected by androgen excess, however, have not been defined. We and others have shown that, in mice, AR is highly expressed in leptin receptor (LepRb) neurons, particularly in the arcuate (ARH) and the ventral premammillary nuclei (PMv). Here, we assessed if LepRb neurons, which are critical in the central regulation of energy homeostasis and exert permissive actions on puberty and fertility, have a role in the pathogenesis of female hyperandrogenism. Prenatally androgenized (PNA) mice lacking AR in LepRb cells (LepRbΔAR) show no changes in body mass, body composition, glucose homeostasis, or sexual maturation. They do show, however, a remarkable improvement of estrous cycles combined with normalization of ovary morphology compared to PNA controls. Our findings indicate that the prenatal androgenization effects on adult reproductive physiology (ie, anestrus and anovulation) are mediated by a subpopulation of LepRb neurons directly sensitive to androgens. They also suggest that the effects of hyperandrogenism on sexual maturation and reproductive function in adult females are controlled by distinct neural circuits.

Distribution of androgen receptor mRNA in the prepubertal male and female mouse brain.

Androgens are steroid hormones that play a critical role in brain development and sexual maturation by acting upon both androgen receptors (AR) and estrogen receptors (ERα/ß) after aromatization. The contribution of estrogens from aromatized androgens in brain development and the central regulation of metabolism, reproduction, and behavior is well defined, but the role of androgens acting on AR has been unappreciated. Here, we map the sex specific expression of Ar in the adult and developing mouse brain. Postnatal days (PND) 12 and 21 were used to target a critical window of prepubertal development. Consistent with previous literature in adults, sex-specific differences in Ar expression were most profound in the bed nucleus of the stria terminalis (BST), medial amygdala (MEA) and medial preoptic area (MPO). Ar expression was also high in these areas at PND 12 and 21 in both sexes. In addition, we describe extra-hypothalamic and extra-limbic areas that show moderate, consistent and similar Ar expression in both sexes at both prepubertal time points. Briefly, Ar expression was observed in olfactory areas of the cerebral cortex, the hippocampus, several thalamic nuclei, and cranial nerve nuclei involved in autonomic sensory and motor function. To further characterize forebrain populations of Ar expressing neurons and determine whether they also coexpress estrogen receptors, we examined expression of Ar, Esr1 and Esr2 in prepubertal mice in selected nuclei. We found populations of neurons in the BST, MEA and MPO that coexpress Ar, but not Esr1 or Esr2, whereas others express a combination of the three receptors. Our findings indicate that various brain areas express Ar during prepubertal development and may play an important role in female neuronal development and physiology.

ERα Signaling in GHRH/Kiss1 Dual-Phenotype Neurons Plays Sex-Specific Roles in Growth and Puberty.

Gonadal steroids modulate growth hormone (GH) secretion and the pubertal growth spurt via undefined central pathways. GH-releasing hormone (GHRH) neurons express estrogen receptor α (ERα) and androgen receptor (AR), suggesting changing levels of gonadal steroids during puberty directly modulate the somatotropic axis. We generated mice with deletion of ERα in GHRH cells (GHRHΔERα), which displayed reduced body length in both sexes. Timing of puberty onset was similar in both groups, but puberty completion was delayed in GHRHΔERα females. Lack of AR in GHRH cells (GHRHΔAR mice) induced no changes in body length, but puberty completion was also delayed in females. Using a mouse model with two reporter genes, we observed that, while GHRHtdTom neurons minimally colocalize with Kiss1hrGFP in prepubertal mice, ∼30% of GHRH neurons coexpressed both reporter genes in adult females, but not in males. Developmental analysis of Ghrh and Kiss1 expression suggested that a subpopulation of ERα neurons in the arcuate nucleus of female mice undergoes a shift in phenotype, from GHRH to Kiss1, during pubertal transition. Our findings demonstrate that direct actions of gonadal steroids in GHRH neurons modulate growth and puberty and indicate that GHRH/Kiss1 dual-phenotype neurons play a sex-specific role in the crosstalk between the somatotropic and gonadotropic axes during pubertal transition.SIGNIFICANCE STATEMENT Late maturing adolescents usually show delayed growth and bone age. At puberty, gonadal steroids have stimulatory effects on the activation of growth and reproductive axes, but the existence of gonadal steroid-sensitive neuronal crosstalk remains undefined. Moreover, the neural basis for the sex differences observed in the clinical arena is unknown. Lack of ERα in GHRH neurons disrupts growth in both sexes and causes pubertal delay in females. Deletion of androgen receptor in GHRH neurons only delayed female puberty. In adult females, not males, a subset of GHRH neurons shift phenotype to start producing Kiss1. Thus, direct estrogen action in GHRH/Kiss1 dual-phenotype neurons modulates growth and puberty and may orchestrate the sex differences in endocrine function observed during pubertal transition.

Lack of AR in LepRb Cells Disrupts Ambulatory Activity and Neuroendocrine Axes in a Sex-Specific Manner in Mice.

Disorders of androgen imbalance, such as hyperandrogenism in females or hypoandrogenism in males, increase risk of visceral adiposity, type 2 diabetes, and infertility. Androgens act upon androgen receptors (AR) which are expressed in many tissues. In the brain, AR are abundant in hypothalamic nuclei involved in regulation of reproduction and energy homeostasis, yet the role of androgens acting via AR in specific neuronal populations has not been fully elucidated. Leptin receptor (LepRb)-expressing neurons coexpress AR predominantly in hypothalamic arcuate and ventral premammillary nuclei (ARH and PMv, respectively), with low colocalization in other LepRb neuronal populations, and very low colocalization in the pituitary gland and gonads. Deletion of AR from LepRb-expressing cells (LepRbΔAR) has no effect on body weight, energy expenditure, and glucose homeostasis in male and female mice. However, LepRbΔAR female mice show increased body length later in life, whereas male LepRbΔAR mice show an increase in spontaneous ambulatory activity. LepRbΔAR mice display typical pubertal timing, estrous cycles, and fertility, but increased testosterone levels in males. Removal of sex steroid negative feedback action induced an exaggerated rise in luteinizing hormone in LepRbΔAR males and follicle-stimulating hormone in LepRbΔAR females. Our findings show that AR can directly affect a subset of ARH and PMv neurons in a sex-specific manner and demonstrate specific androgenic actions in the neuroendocrine hypothalamus.