SUMMARY Sexual dimorphisms in the brain underlie behavioral sex differences, but the function of individual sexually dimorphic neuronal populations is poorly understood. Neuronal sexual dimorphisms typically represent quantitative differences in cell number, gene expression, or other features, and it is unknown if these dimorphisms control sex-typical behavior in one sex exclusively or in both sexes. The progesterone receptor (PR) controls female sexual behavior, and we find many sex differences in number, distribution, or projections of PR-expressing neurons in the adult mouse brain. We have ablated one such PR-expressing neuronal population located in the ventromedial hypothalamus (VMH) using a novel genetic strategy. Ablation of these neurons in females greatly diminishes sexual receptivity. Strikingly, the corresponding ablation in males reduces mating and aggression. Our findings reveal the functions of a molecularly-defined, sexually dimorphic neuronal population in the brain. Moreover we show that sexually dimorphic neurons can control distinct sex-typical behaviors in both sexes.

SUMMARY Aromatase-expressing neuroendocrine neurons in the vertebrate male brain synthesize estradiol from circulating testosterone. This locally produced estradiol controls neural circuits underlying courtship vocalization, mating, aggression, and territory marking in male mice. How aromatase-expressing neuronal populations control these diverse estrogen-dependent male behaviors is poorly understood, and the function, if any, of aromatase-expressing neurons in females is unclear. Using targeted genetic approaches, we show that aromatase-expressing neurons within the male posterodorsal medial amygdala (MeApd) regulate components of aggression but not other estrogen-dependent male-typical behaviors. Remarkably, aromatase-expressing MeApd neurons in females are specifically required for components of maternal aggression, which we show is distinct from intermale aggression in pattern and execution. Thus, aromatase-expressing MeApd neurons control distinct forms of aggression in the two sexes. Moreover, our findings indicate that complex social behaviors are separable in a modular manner at the level of genetically identified neuronal populations.

OBJECTIVEHypothalamic leptin resistance is found in most common forms of obesity, such as diet-induced obesity, and is associated with increased expression of suppressor of cytokine signaling 3 (Socs3) in the hypothalamus of diet-induced obese animals. This study aims to determine the functional consequence of Socs3 upregulation on leptin signaling and obesity, and to investigate whether Socs3 upregulation affects energy balance in a cell type–specific way.RESEARCH DESIGN AND METHODSWe generated transgenic mice overexpressing Socs3 in either proopiomelanocortin (POMC) or leptin receptor–expressing neurons, at levels similar to what is observed in diet-induced obesity.RESULTSUpregulation of Socs3 in POMC neurons leads to impairment of STAT3 and mammalian target of rapamycin (mTOR)–S6K-S6 signaling, with subsequent leptin resistance, obesity, and glucose intolerance. Unexpectedly, Socs3 upregulation in leptin receptor neurons results in increased expression of STAT3 protein in mutant hypothalami, but does not lead to obesity.CONCLUSIONSOur study establishes that Socs3 upregulation alone in POMC neurons is sufficient to cause leptin resistance and obesity. Socs3 upregulation impairs both STAT3 and mTOR signaling before the onset of obesity. The lack of obesity in mice with upregulated Socs3 in leptin receptor neurons suggests that Socs3's effect on energy balance could be cell type specific. Our study indicates that POMC neurons are important mediators of Socs3's effect on leptin resistance and obesity, but that other cell types or alteration of other signaling regulators could contribute to the development of obesity.

Chronic consumption of a fat-rich diet leads to attenuation of leptin signaling in hypothalamic neurons, a hallmark feature of cellular leptin resistance. To date, little is known about the temporal and spatial dysregulation of neuronal function under conditions of nutrient excess. We show that agouti-related protein (AgRP)-expressing neurons precede proopiomelanocortin neurons in developing diet-induced cellular leptin resistance. High-fat diet-induced up-regulation of suppressor of cytokine signaling-3 (SOCS3) occurs in AgRP neurons before proopiomelanocortin and other hypothalamic neurons. SOCS3 expression in AgRP neurons increases after 2 d of high-fat feeding, but reduces after switching to a low-fat diet for 1 d. Consistently, transgenic overexpression of SOCS3 in AgRP neurons produces metabolic phenotypes resembling those observed after short-term high-fat feeding. We further show that AgRP neurons are the predominant cell type situated outside the blood-brain barrier in the mediobasal hypothalamus. AgRP neurons are more responsive to low levels of circulating leptin, but they are also more prone to development of leptin resistance in response to a small increase in blood leptin concentrations. Collectively, these results suggest that AgRP neurons are able to sense slight changes in plasma metabolic signals, allowing them to serve as first-line responders to fluctuation of energy intake. Furthermore, modulation of SOCS3 expression in AgRP neurons may play a dynamic and physiological role in metabolic fine tuning in response to short-term changes of nutritional status.M ost common forms of obesity, including diet-induced obesity, are associated with hyperleptinemia and impairment of leptin signaling in hypothalamic neurons, the hallmark feature of cellular leptin resistance. Suppressor of cytokine signaling-3 (SOCS3), a direct transcriptional product of STAT3, is up-regulated in the hypothalamus of diet-induced obese animals (1, 2). Mice with heterozygous mutation of the Socs3 gene, neuronal, or proopiomelanocortin (POMC)-specific deletion of the Socs3 gene are hypersensitive to leptin and resistant to diet-induced obesity (3-5). Conversely, up-regulation of SOCS3 in POMC neurons of chow-fed mice leads to increased body adiposity (6). In addition, wide-spread up-regulation of SOCS3 has been shown to be associated with neuronal inflammation in diet-induced obese animals (7). Thus SOCS3, which is up-regulated in chronic obesity, is commonly thought to play a pathophysiological role in obesity-associated leptin resistance.Multiple neuronal subtypes in several regions of the hypothalamus, including the arcuate nucleus, ventromedial hypothalamus, dorsomedial hypothalamus, and lateral hypothalamic area, have been implicated in the regulation of energy balance and leptin action (8,9). A number of hypothalamic neurons and extrahypothalamic neurons express functional leptin receptor (10, 11). Among these neurons, POMC and agouti-related protein (AgRP) neurons are two key arcuate neuronal subtypes. POMC and AgRP neu...

In vivo calcium imaging from axons provides direct interrogation of afferent neural activity, informing the neural representations that a local circuit receives. Unlike in somata and dendrites, axonal recording of neural activity-both electrically and optically-has been difficult to achieve, thus preventing comprehensive understanding of neuronal circuit function. Here we developed an active transportation strategy to enrich GCaMP6, a genetically encoded calcium indicator, uniformly in axons with sufficient brightness, signal-to-noise ratio, and photostability to allow robust, structure-specific imaging of presynaptic activity in awake mice. Axon-targeted GCaMP6 enables frame-to-frame correlation for motion correction in axons and permits subcellular-resolution recording of axonal activity in previously inaccessible deep-brain areas. We used axon-targeted GCaMP6 to record layer-specific local afferents without contamination from somata or from intermingled dendrites in the cortex. We expect that axon-targeted GCaMP6 will facilitate new applications in investigating afferent signals relayed by genetically defined neuronal populations within and across specific brain regions.

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Leptin is a fat-derived hormone that exerts pleiotropic effects on energy balance and neuroendocrine functions. Mice defective in leptin or its receptor [leptin receptor, isoform b (LepRb)] exhibit profound obesity, infertility, and reduced linear growth. Leptin binding to its receptor triggers multiple signaling pathways, including signal transducer and activator of transcription 3 (Stat 3), phosphatidylinositol-3-kinase, and ERK. A considerable amount of effort has been focused on how these signaling pathways mediate diverse leptin functions. Mice containing a mutant LepRb incapable of Stat3 signaling are obese but remain fertile with enhanced linear growth. In contrast, deletion of Stat3 in the whole brain with Nestin-Cre results in infertility and decreased linear growth, in addition to obesity. The additional phenotypes of the Nestin-mediated deletion could reflect Stat3 action in non-LepRb neurons or leptin-independent Stat3 actions in LepRb neurons. To resolve this discrepancy and to gain more insight into the metabolic actions of Stat3, we have generated mice in which Stat3 is disrupted specifically in LepRb neurons after the onset of leptin receptor expression. We show that mutant mice exhibit profound obesity with increased linear growth and normal fertility. In addition, impaired glycemic control in these animals correlates with their degree of obesity. These results demonstrate that Stat3 in LepRb neurons does not regulate linear growth or fertility. These results further suggest that leptin's effects on growth and reproduction are mediated by other signaling pathways, and that Stat3-mediated control of these functions is mediated independently of leptin and LepRb neurons.

c-Jun-N-terminal kinase (JNK) is a signaling molecule that is activated by proinflammatory signals, endoplasmic reticulum (ER) stress, and other environmental stressors. Although JNK has diverse effects on immunological responses and insulin resistance in peripheral tissues, a functional role for JNK in feeding regulation has not been established. In this study, we show that central inhibition of JNK activity potentiates the stimulatory effects of glucocorticoids on food intake and that this effect is abolished in mice whose agouti-related peptide (AgRP) neurons are degenerated. JNK1-deficient mice feed more upon central administration of glucocorticoids, and glucocorticoid receptor nuclear immunoreactivity is enhanced in the AgRP neurons. JNK inhibition in hypothalamic explants stimulates Agrp expression, and JNK1-deficient mice exhibit increased Agrp expression, heightened hyperphagia, and weight gain during refeeding. Our study shows that JNK1 is a novel regulator of feeding by antagonizing glucocorticoid function in AgRP neurons. Paradoxically, JNK1 mutant mice feed less and lose more weight upon central administration of insulin, suggesting that JNK1 antagonizes insulin function in the brain. Thus, JNK may integrate diverse metabolic signals and differentially regulate feeding under distinct physiological conditions.