A preoptic neuronal population controls fever and appetite during sickness.

During infection, animals exhibit adaptive changes in physiology and behaviour aimed at increasing survival. Although many causes of infection exist, they trigger similar stereotyped symptoms such as fever, warmth-seeking, loss of appetite and fatigue1,2. Yet exactly how the nervous system alters body temperature and triggers sickness behaviours to coordinate responses to infection remains unknown. Here we identify a previously uncharacterized population of neurons in the ventral medial preoptic area (VMPO) of the hypothalamus that are activated after sickness induced by lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid. These neurons are crucial for generating a fever response and other sickness symptoms such as warmth-seeking and loss of appetite. Single-nucleus RNA-sequencing and multiplexed error-robust fluorescence in situ hybridization uncovered the identity and distribution of LPS-activated VMPO (VMPOLPS) neurons and non-neuronal cells. Gene expression and electrophysiological measurements implicate a paracrine mechanism in which the release of immune signals by non-neuronal cells during infection activates nearby VMPOLPS neurons. Finally, we show that VMPOLPS neurons exert a broad influence on the activity of brain areas associated with behavioural and homeostatic functions and are synaptically and functionally connected to circuit nodes controlling body temperature and appetite. Together, these results uncover VMPOLPS neurons as a control hub that integrates immune signals to orchestrate multiple sickness symptoms in response to infection.

Activation of the medial preoptic area (MPOA) ameliorates loss of maternal behavior in a Shank2 mouse model for autism.

Impairments in social relationships and awareness are features observed in autism spectrum disorders (ASDs). However, the underlying mechanisms remain poorly understood. Shank2 is a high-confidence ASD candidate gene and localizes primarily to postsynaptic densities (PSDs) of excitatory synapses in the central nervous system (CNS). We show here that loss of Shank2 in mice leads to a lack of social attachment and bonding behavior towards pubs independent of hormonal, cognitive, or sensitive deficits. Shank2-/- mice display functional changes in nuclei of the social attachment circuit that were most prominent in the medial preoptic area (MPOA) of the hypothalamus. Selective enhancement of MPOA activity by DREADD technology re-established social bonding behavior in Shank2-/- mice, providing evidence that the identified circuit might be crucial for explaining how social deficits in ASD can arise.

Identification of preoptic sleep neurons using retrograde labelling and gene profiling.

In humans and other mammalian species, lesions in the preoptic area of the hypothalamus cause profound sleep impairment, indicating a crucial role of the preoptic area in sleep generation. However, the underlying circuit mechanism remains poorly understood. Electrophysiological recordings and c-Fos immunohistochemistry have shown the existence of sleep-active neurons in the preoptic area, especially in the ventrolateral preoptic area and median preoptic nucleus. Pharmacogenetic activation of c-Fos-labelled sleep-active neurons has been shown to induce sleep. However, the sleep-active neurons are spatially intermingled with wake-active neurons, making it difficult to target the sleep neurons specifically for circuit analysis. Here we identify a population of preoptic area sleep neurons on the basis of their projection target and discover their molecular markers. Using a lentivirus expressing channelrhodopsin-2 or a light-activated chloride channel for retrograde labelling, bidirectional optogenetic manipulation, and optrode recording, we show that the preoptic area GABAergic neurons projecting to the tuberomammillary nucleus are both sleep active and sleep promoting. Furthermore, translating ribosome affinity purification and single-cell RNA sequencing identify candidate markers for these neurons, and optogenetic and pharmacogenetic manipulations demonstrate that several peptide markers (cholecystokinin, corticotropin-releasing hormone, and tachykinin 1) label sleep-promoting neurons. Together, these findings provide easy genetic access to sleep-promoting preoptic area neurons and a valuable entry point for dissecting the sleep control circuit.

Lithium Enhances the GABAergic Synaptic Activities on the Hypothalamic Preoptic Area (hPOA) Neurons.

Lithium (Li+) salt is widely used as a therapeutic agent for treating neurological and psychiatric disorders. Despite its therapeutic effects on neurological and psychiatric disorders, it can also disturb the neuroendocrine axis in patients under lithium therapy. The hypothalamic area contains GABAergic and glutamatergic neurons and their receptors, which regulate various hypothalamic functions such as the release of neurohormones, control circadian activities. At the neuronal level, several neurotransmitter systems are modulated by lithium exposure. However, the effect of Li+ on hypothalamic neuron excitability and the precise action mechanism involved in such an effect have not been fully understood yet. Therefore, Li+ action on hypothalamic neurons was investigated using a whole-cell patch-clamp technique. In hypothalamic neurons, Li+ increased the GABAergic synaptic activities via action potential independent presynaptic mechanisms. Next, concentration-dependent replacement of Na+ by Li+ in artificial cerebrospinal fluid increased frequencies of GABAergic miniature inhibitory postsynaptic currents without altering their amplitudes. Li+ perfusion induced inward currents in the majority of hypothalamic neurons independent of amino-acids receptor activation. These results suggests that Li+ treatment can directly affect the hypothalamic region of the brain and regulate the release of various neurohormones involved in synchronizing the neuroendocrine axis.

Key role of estrogen receptor ß in the organization of brain and behavior of the Japanese quail.

Estrogens play a key role in the sexual differentiation of the brain and behavior. While early estrogen actions exert masculinizing effects on the brain of male rodents, a diametrically opposite effect is observed in birds where estrogens demasculinize the brain of females. Yet, the two vertebrate classes express similar sex differences in the brain and behavior. Although ERα is thought to play a major role in these processes in rodents, the role of ERß is still controversial. In birds, the identity of the estrogen receptor(s) underlying the demasculinization of the female brain remains unclear. The aim of the present study was thus to determine in Japanese quail the effects of specific agonists of ERα (propylpyrazole triol, PPT) and ERß (diarylpropionitrile, DPN) administered at the beginning of the sensitive period (embryonic day 7, E7) on the sexual differentiation of male sexual behavior and on the density of vasotocin-immunoreactive (VT-ir) fibers, a known marker of the organizational action of estrogens on the quail brain. We demonstrate that estradiol benzoate and the ERß agonist (DPN) demasculinize male sexual behavior and decrease the density of VT-ir fibers in the medial preoptic nucleus and the bed nucleus of the stria terminalis, while PPT has no effect on these measures. These results clearly indicate that ERß, but not ERα, is involved in the estrogen-induced sexual differentiation of brain and sexual behavior in quail.

Nature and nurture of sexual partner preference: Teachings from prenatal administration of acetaminophen in male rats.

The organizational-activational hypothesis indicates that activation of adult sexual behavior in males depends on organization of the masculine brain during the perinatal sensitive period. In the medial preoptic area such masculinization depends on a neuroendocrine cascade that includes exposure to testosterone, aromatization to estradiol, activation of estrogen receptors, synthesis of cyclooxygenase (COX), increase of prostaglandins, release of glutamate, and activation of AMPA receptors that result in the formation of more dendritic spines. Thus, in the present study we assessed the sexual partner preference (SPP) of adult male rats prenatally treated with acetaminophen (APAP), an analgesic/antipyretic drug that inhibits COX-2 and is commonly used and prescribed during pregnancy. Female rats received either saline (2 ml/kg s.c.) or APAP (50 mg/kg s.c.) every 12 h, during days 16-20 of pregnancy. At postnatal day PD60 half of the male offspring were exposed to sexual experience with receptive females during 5 trials, and the other half remained sexually naïve. At PD90 all them were tested for SPP with one sexually receptive female and one stud male. The results indicated that only APAP-naïve males failed to display SPP. However, APAP-experienced males displayed SPP for females. We discuss the effects of prenatal APAP in the disruption of unconditioned responses towards females (nature mechanisms), and the effects of sexual experience (nurture mechanisms) in the development of conditioned heterosexual preference.

Dietary phytoestrogens modulate aggression and activity in social behavior circuits of male mice.

Phytoestrogens comprise biologically active constituents of human and animal diet that can impact on systemic and local estrogen functions in the brain. Here we report on the importance of dietary phytoestrogens for maintaining activity in a brain circuit controlling aggressive and social behavior of male mice. After six weeks of low-phytoestrogen chronic diet (diadzein plus genistein <20 µg/g) a reduction of intermale aggression and altered territorial marking behavior could be observed, compared to littermates on a standard soy-bean based diet (300 µg/g). Further, mice on low-phyto diet displayed a decrease in sociability and a reduced preference for social odors, indicating a general disturbance of social behavior. Underlying circuits were investigated by analysing the induction of the activity marker c-Fos upon social encounter. Low-phyto diet led to a markedly reduced c-Fos induction in the medial as well as the cortical amygdala, the lateral septum, medial preoptic area and bed nucleus of the stria terminalis. No difference between groups was observed in the olfactory bulb. Together our data suggest that dietary phytoestrogens critically modulate social behavior circuits in the male mouse brain.

Testosterone-related behavioral and neural mechanisms associated with location preferences: A model for territorial establishment.

Territoriality is an adaptive behavioral trait that is important for animal's fitness and there still remains much to learn about the proximate mechanisms underlying the development of territoriality. We speculate that the formation of a conditioned place preference (CPP), an increased time allocation to the environment where a rewarding experience occurred, contributes to territoriality. Testosterone (T) plays an important role in modulating territorial behaviors and T pulses can induce a CPP. We confirmed previous findings in California mice (Peromyscus californicus) that T pulses can induce a CPP in singly-housed, but not group-housed males. Housing singly may be similar enough to dispersal in nature to initiate similar hormonal and neuroanatomical changes needed for the development of territoriality. We further revealed that T pulses interact with the single housing experience and appear to enhance the motivation to be aggressive towards a stimulus male. On a neural level, being singly housed upregulated levels of androgen receptors in the preoptic area, which positively correlated with the strength of the CPP. We speculate that this change in androgen sensitivity in the preoptic area is characteristic of males that have dispersed, making them more sensitive to T pulses. Also, single housing increased markers of synaptic plasticity in the nucleus accumbens, ventral and dorsal hippocampus, neural changes that may be associated with dispersal, reproduction and territory establishment. These behavioral and neural changes may reflect the life history transition from residing in the natal territory to dispersing and establishing a new territory.

Involvement of glutamatergic mechanisms in the median preoptic nucleus in the dipsogenic response induced by angiotensinergic activation of the subfornical organ in rats.

Experiments were done to investigate the role of glutamatergic systems in the median preoptic nucleus (MnPO) in the water ingestion induced by administration of angiotensin II (ANG II) in the subfornical organ (SFO) in the awake rat. Microdialysis methods were utilized to quantify the extracellular content of glutamate (Glu) in the region of MnPO. Microinjection of ANG II (10-10 M) into the SFO significantly increased the release of Glu in the MnPO in the rats under the condition that water is available for drinking and the rats under the condition that water is not available for drinking. The amount of initial maximal increases in the Glu levels elicited by the ANG II injection was quite similar in drinking and non-drinking rats, whereas the duration of the response was much longer in non-drinking than in drinking rats. The amount of water ingestion in 20 min immediately after the ANG II injection was significantly enhanced by previous injections of N-methyl-D-aspartate (NMDA, 10 µM) into the MnPO, while the ANG II-induced water ingestion was attenuated by pretreatment with the NMDA antagonist dizocilpine (MK-801, 10 µM). The amount of water intake elicited by the ANG II injection into the SFO was enhanced by previous injections of either the non-NMDA agonist kainic acid (KA, 50 µM) or quisqualic acid (QA, 50 µM) into the MnPO. On the contrary, the ANG II-induced drinking response was diminished by pretreatment with the non-NMDA antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 µM) in the MnPO. Each injection of NMDA, KA, and QA into the MnPO produced drinking behavior. These results imply that the glutamatergic neural pathways to the MnPO may transmit the information for eliciting drinking in response to ANG II acting at the SFO. Our data further provide evidence that the ANG II-induced dipsogenic response may be mediated through both NMDA and non-NMDA glutamatergic receptor mechanisms in the MnPO.

Prepubertal overexposure to manganese induce precocious puberty through GABAA receptor/nitric oxide pathway in immature female rats.

Gamma-aminobutyric acid (GABA) plays a critical role in regulation of gonadotropin-releasing hormone (GnRH) through GABAA receptor (GABAAR). Nitric oxide (NO) production has correlation with GABA and regulates GnRH secretion. This study was performed to examine the mechanisms by which manganese (Mn) accelerate puberty onset involves GABAAR/NO pathway in the preoptic area-anterior hypothalamus (POA-AH) in immature female rats. First, female rats received daily dose of MnCl2 0 (saline), 2.5, 5 and 10 mg/kg b.w by oral gavage during postnatal day (PND) 21-32. Animals administered with 10 mg/kg MnCl2 exhibited earlier puberty onset age and advanced ovary and uterus development than these in saline-treatment group. Furthermore, we found that decrease of GABAAR result in elevated production of nitric oxide synthase1 (NOS1), NO and GnRH in the POA-AH. Second, we recorded the neuronal spikes alternation after perfusion with GABAAR inhibitor bicuculline (BIC), GABAAR agonist isoguvacine (isog), and MnCl2 from the POA-AH in acute brain slices of PND21 rats. Spontaneous firing revealed a powerful GABAAR-mediated action on immature POA-AH and confirm that MnCl2 has a significant effect on GABAAR. Third, we revealed that decrease in NOS1 and NO production by treatment with isog-alone or isog+MnCl2 contribute to the decrease of GnRH in the POA-AH and a delayed puberty onset age compared to treatment with MnCl2-alone. Together, these results suggested that excessive exposure to MnCl2 stimulates NO production through decreased GABAAR in the POA-AH to advance puberty onset in immature female rats.

Mast Cells in the Developing Brain Determine Adult Sexual Behavior.

Many sex differences in brain and behavior are programmed during development by gonadal hormones, but the cellular mechanisms are incompletely understood. We found that immune-system-derived mast cells are a primary target for the masculinizing hormone estradiol and that mast cells are in turn primary mediators of brain sexual differentiation. Newborn male rats had greater numbers and more activated mast cells in the preoptic area (POA), a brain region essential for male copulatory behavior, than female littermates during the critical period for sexual differentiation. Inhibiting mast cells with a stabilizing agent blunted the masculinization of both POA neuronal and microglial morphology and adult sex behavior, whereas activating mast cells in females, even though fewer in number, induced masculinization. Treatment of newborn females with a masculinizing dose of estradiol increased mast cell number and induced mast cells to release histamine, which then stimulated microglia to release prostaglandins and thereby induced male-typical synaptic patterning. These findings identify a novel non-neuronal origin of brain sex differences and resulting motivated behaviors.SIGNIFICANCE STATEMENT We found that immune-system-derived mast cells are a primary target for the masculinizing hormone estradiol and that mast cells are in turn primary mediators of brain sexual differentiation. These findings identify a novel non-neuronal origin of brain sex differences and resulting motivated behaviors.

Alterations of estradiol-induced histone H3 acetylation in the preoptic area and anteroventral periventricular nucleus of middle-aged female rats.

In this study we investigated the characteristics of histone H3 acetylation in the anterior hypothalamus under E2 positive feedback to gain a better understanding of the mechanism underlying reduced GnRH neuron activation and altered gene expression in female reproductive aging. Young and middle-aged female rats were ovariectomized (OVX) and treated with estradiol (E2) or oil. C-Fos expression, the number of GnRH neurons co-localized with c-Fos in the preoptic area (POA), and the number of acetylated histone H3 cells in the POA and anteroventral periventricular nucleus (AVPV) were quantified at the time of the expected GnRH neuron activation. We used real-time PCR to evaluate the expression of Esr1 target genes including Kiss1 and VGluT2 and genes known as Esr1 coregulators in the anterior hypothalamus. Our results show that in the young females, E2 markedly increased histone H3 acetylation in the POA and AVPV, coincident with increased c-Fos and GnRH neuron activation in the POA. In middle-aged females, E2-induced histone H3 acetylation was reduced in the POA but was not significantly altered in the AVPV. This occurred in association with a reduction of c-Fos expression and the number of GnRH cells expressing c-Fos in the POA as well as a down-regulation of Kiss1 and VGluT2 mRNA expression in the anterior hypothalamus of the animals. E2 caused significant decreases in Ncoa2 and Crebbp mRNA expression in the anterior hypothalamus of young, but not middle-aged females. Taken together, these data suggest that alterations of histone H3 acetylation in the POA and AVPV and the inability of Ncoa2 and Crebbp to respond to E2 in the middle-aged anterior hypothalamus partially contribute to the decline of GnRH neuron activation and E2 target gene expression changes that occur in female along with reproductive aging.

Angiotensin type 1a receptors in the median preoptic nucleus support intermittent hypoxia-induced hypertension.

Chronic intermittent hypoxia (CIH) is a model of the hypoxemia from sleep apnea that causes a sustained increase in blood pressure. Inhibition of the central renin-angiotensin system or FosB in the median preoptic nucleus (MnPO) prevents the sustained hypertensive response to CIH. We tested the hypothesis that angiotensin type 1a (AT1a) receptors in the MnPO, which are upregulated by CIH, contribute to this hypertension. In preliminary experiments, retrograde tract tracing studies showed AT1a receptor expression in MnPO neurons projecting to the paraventricular nucleus. Adult male rats were exposed to 7 days of intermittent hypoxia (cycling between 21% and 10% O2 every 6 min, 8 h/day during light phase). Seven days of CIH was associated with a FosB-dependent increase in AT1a receptor mRNA without changes in the permeability of the blood-brain barrier in the MnPO. Separate groups of rats were injected in the MnPO with an adeno-associated virus containing short hairpin (sh)RNA against AT1a receptors to test their role in intermittent hypoxia hypertension. Injections of shRNA against AT1a in MnPO blocked the increase in mRNA associated with CIH, prevented the sustained component of the hypertension during normoxia, and reduced circulating advanced oxidation protein products, an indicator of oxidative stress. Rats injected with shRNA against AT1a and exposed to CIH had less FosB staining in MnPO and the rostral ventrolateral medulla after intermittent hypoxia than rats injected with the control vector that were exposed to CIH. Our results indicate AT1a receptors in the MnPO contribute to the sustained blood pressure increase to intermittent hypoxia.

Estrogen Receptors Alpha and Beta in POA-AHA Region Regulate Asymmetrically Ovulation.

We examined the role of the estrogen receptors alpha (ERα) and beta (ERß) in of the preoptic-anterior hypothalamic area (POA-AHA) in the regulation of ovulation in rats. The number of ERα- and ERß-immunoreactive (-ir) cells was determined at 09:00, 13:00, and 17:00 h of each stage of the estrous cycle in intact rats. Additionally, the effects of blocking ERα and ERß on ovulation rate at 09:00 h on diestrus-2 or proestrus day through the microinjection of methyl-piperidino-pyrazole (MPP) or cyclofenil in either side of POA-AHA were evaluated. The number of ERα-ir and ERß-ir cells in POA-AHA varied in each phase of estrous cycle. Either MPP or cyclofenil in the right side of POA-AHA on diestrus-2 day reduced the ovulation rate, while at proestrus day it was decreased in rats treated in either side with MPP, and in those treated with cyclofenil in the left side. MPP or cyclofenil produced a decrease in the surge of luteinizing hormone levels (LH) and an increase in progesterone and follicle stimulating hormone (FSH). Replacement with synthetic luteinizing hormone-releasing hormone in non-ovulating rats treated with MPP or cyclofenil restored ovulation. These results suggest that activation of estrogen receptors on the morning of diestrus-2 and proestrus day asymmetrically regulates ovulation and appropriately regulates the secretion of FSH and progesterone in the morning and afternoon of proestrus day. This ensures that both, the preovulatory secretion of LH and ovulation, occur at the right time.

Role of oxytocin in the medial preoptic area (MPOA) in the modulation of paternal behavior in mandarin voles.

Parental care plays an important role in individual survival and development in mammals. Many studies have focused on the mechanisms underlying maternal behavior. However, the underlying neural mechanisms of paternal behavior are less understood. Using monogamous mandarin voles (Microtus mandarinus), the present study found that fathers initiated more paternal behavior and the virgin male showed more infanticide. Moreover fathers had shorter latency to approach a pup at the postnatal day (PND) 10 than PND1, PND20 than nonfathers. Fathers had a shorter latency to take care of unfamiliar pups than nonfathers. They had higher levels of paternal behavior at PND 10 than PND1 and PND20 toward the mandarin vole pups. Fathers had a significantly higher serum concentration of oxytocin (OT) than virgin males. Both RT-PCR and Western blot results indicated that the levels of the oxytocin receptor (OTR) in the medial preoptic area (MPOA) of fathers were significantly higher than in virgin males, but the levels of vasopressin 1a receptor (V1AR) mRNA and protein expression in the MPOA did not show significant differences. Microinjection of an oxytocin receptor antagonist into the MPOA significantly reduced the total duration of paternal behavior and increased the latency to approach the pup and initiate paternal behavior. Our results indicated that OT plays a key role in the modulation of paternal behavior via the MPOA.

Anorexigenic effects of estradiol in the medial preoptic area occur through membrane-associated estrogen receptors and metabotropic glutamate receptors.

Activation of membrane-associated estrogen receptors (mER) decreases food and water intake in female rats. Additional studies suggest these effects are mediated, at least in part, by membrane-associated estrogen receptor alpha (ERα). Nevertheless, the critical site of action and the intracellular signaling required for the ingestive effects of ERα remain unclear. Estradiol given to the medial preoptic area (mPOA) decreases ingestive behaviors, and membrane-associated ERα has been shown to affect intracellular signaling through interactions with metabotropic glutamate receptor (mGluR) subtypes, but an involvement of this signaling pathway, in the mPOA, in ingestive behavior remains untested. To address these open questions, we first showed that activation of mER in the mPOA decreased both overnight food and water intake, and did so in a time course consistent with a genomic mechanism of action. Next, we tested the requirement of mGluR1a signaling in the mPOA for the anorexigenic and anti-dipsogenic effects of estradiol. As expected, estradiol in the mPOA decreased food intake, but only in the absence of an mGluR1a antagonist. The same was not true for estradiol effects on water intake, which were unaffected by an mGluR1a antagonist. These results suggest that estrogens require mGluR activation for at least some of their effects on ingestive behaviors, and indicate that the mPOA is a critical site of action. The results also reveal an interesting divergence in the estrogenic control of ingestive behavior by which mGluR signaling in the mPOA plays a role in the control of food intake, but not water intake.

Site-specific effects of aromatase inhibition on the activation of male sexual behavior in male Japanese quail (Coturnix japonica).

Aromatization within the medial preoptic nucleus (POM) is essential for the expression of male copulatory behavior in Japanese quail. However, several nuclei within the social behavior network (SBN) also express aromatase. Whether aromatase in these loci participates in the behavioral activation is not known. Castrated male Japanese quail were implanted with 2 subcutaneous Silastic capsules filled with crystalline testosterone and with bilateral stereotaxic implants filled with the aromatase inhibitor Vorozole targeting the POM, the bed nucleus of the stria terminalis (BST) or the ventromedial nucleus of the hypothalamus (VMN). Control animals were implanted with testosterone and empty bilateral stereotaxic implants. Starting 2 days after the surgery, subjects were tested for the expression of consummatory sexual behavior (CSB) every other day for a total of 10 tests. They were also tested once for appetitive sexual behavior (ASB) as measured by the rhythmic cloacal sphincter movements displayed in response to the visual presentation of a female. CSB was drastically reduced when the Vorozole implants were localized in the POM, but not in the BST nor in the VMN. Birds with implants in the BST took longer to show CSB in the first 6 tests than controls, suggesting a role of the BST in the acquisition of the full copulatory ability. ASB was not significantly affected by aromatase blockade in any region. These data confirm the key role played by the POM in the control of male sexual behavior and suggest a minor role for aromatization in the BST or VMN.

AT1a influences GABAA-mediated inhibition through regulation of KCC2 expression.

The median preoptic nucleus (MnPO) is an integrative site involved in body fluid homeostasis, cardiovascular control, thermoregulation, and sleep homeostasis. Angiotensin II (ANG II), a neuropeptide shown to have excitatory effects on MnPO neurons, is of particular interest with regard to its role in body fluid homeostasis and cardiovascular control. The present study investigated the role of angiotensin type 1a (AT1a) receptor activation on neuronal excitability in the MnPO. Male Sprague-Dawley rats were infused with an adeno-associated virus with an shRNA against the AT1a receptor or a scrambled control. In vitro loose-patch voltage-clamp recordings of spontaneous action potential activity were made from labeled MnPO neurons in response to brief focal application of ANG II or the GABAA receptor agonist muscimol. Additionally, tissue punches from MnPO were taken to asses mRNA and protein expression. AT1a receptor knockdown neurons were insensitive to ANG II and showed a marked reduction in GABAA-mediated inhibition. The reduction in GABAA-mediated inhibition was not associated with reductions in mRNA or protein expression of GABAA ß-subunits. Knockdown of the AT1a receptor was associated with a reduction in the potassium-chloride cotransporter KCC2 mRNA as well as a reduction in pS940 KCC2 protein. The impaired GABAA-mediated inhibition in AT1a knockdown neurons was recovered by bath application of phospholipase C and protein kinase C activators. The following study indicates that AT1a receptor activation mediates the excitability of MnPO neurons, in part, through the regulation of KCC2. The regulation of KCC2 influences the intracellular [Cl-] and the subsequent efficacy of GABAA-mediated currents.

BDNF infusion into the MPN mag is sufficient to restore copulatory behavior in the castrated Syrian hamster.

Testosterone plays a key role in the expression of male sex behavior by influencing cellular activity and synapses within the magnocellular medial preoptic nucleus (MPN mag), a sub-nucleus of the medial preoptic area (MPOA) in the Syrian hamster. Although the mechanisms underlying hormonally-induced synaptic plasticity in this region remain elusive, the data suggests that an increase in synaptic density may mediate testosterone's effects on copulation. As brain derived neurotrophic factor (BDNF) plays an integral role in regulating synaptic plasticity and gonadal steroids regulate the levels of BDNF, we hypothesize that BDNF may mediate the effects of gonadal hormones on copulatory behavior. To test this hypothesis, we infused BDNF or controls into the MPN mag of long-term castrates. Our results indicate that BDNF, but not the controls, restored copulatory behavior in castrated male Syrian hamsters. Furthermore, the rise of BDNF expression in the MPOA preceded the rise of synaptophysin following testosterone replacement in castrated males. These data are consistent with our hypothesis, implicating a role for BDNF in mediating testosterone's action on copulation and suggest that the delay in testosterone's restoration of copulation is, in part, due to the delay in the increase of BDNF and synaptophysin.

TrkB is necessary for male copulatory behavior in the Syrian Hamster (Mesocricetus auratus).

The magnocellular medial preoptic nucleus (MPN mag), a subdivision of the medial preoptic area (MPOA), plays a critical role in the regulation of copulation in the male Syrian hamster; in part by mediating the effects of gonadal steroids. For example, ablation of the MPN mag eliminates mating and testosterone placed in the MPN mag restores mating in castrated males. Furthermore, testosterone treatment enhances synaptic density and dendritic spines in the MPN mag. Thus, copulatory behaviors are correlated with increases in synaptic morphology in the MPN mag. As brain derived neurotrophic factor (BDNF) and its receptor, tyrosine receptor kinase-B (TrkB), effect neuronal growth and synaptic plasticity, this study explored the role of TrkB and BDNF in mediating testosterone's effects on the MPN mag and behavior. Testosterone treatment increased BDNF expression and conversely lowered TrkB expression in the MPOA. siRNA-mediated TrkB knockdown in the MPN mag eliminated copulation two-days post injection and the behavior was restored one week later. These data indicate that testosterone influences the expression of BDNF and TrkB in the MPOA and that expression of copulation is dependent on the presence of TrkB. Taken together our findings support a role for TrkB and BDNF in mediating the effects of testosterone on copulatory behavior in the Syrian hamster.