Blockade of corticotropin-releasing factor receptor 1 in the central amygdala prevents cocaine-seeking behaviour induced by orexin-A administered to the posterior paraventricular nucleus of the thalamus in male rats.

Background: Orexin-A (OrxA) administration in the posterior paraventricular nucleus of the thalamus (pPVT) reinstates extinguished cocaine-seeking behaviour following extended access to the drug (a model of dependence). The pPVT receives and integrates information associated with emotionally salient events and sends excitatory inputs to brain regions involved in the expression of emotional states, such as those driving cocaine-seeking behaviour (i.e., the nucleus accumbens, the central nucleus of the amygdala [CeA], the basolateral amygdala, the bed nucleus of the stria terminalis [BNST] and the prefrontal cortex). Methods: We monitored the activation pattern of these regions (measured by Fos) during cocaine-seeking induced by OrxA administered to the pPVT. The BNST and CeA emerged as being selectively activated. To test whether the functionality of these regions was pivotal during OrxA-induced cocaine-seeking behaviour, we transiently inactivated these regions concomitantly with OrxA administration to the pPVT. We then tested the participation of corticotropin-releasing factor receptors (CRF1) in the CeA during OrxA-induced cocaine-seeking using the CRF1 antagonist CP154526. Results: We observed selective activation of the CeA and BNST during cocaine-seeking induced by OrxA administered to the pPVT, but only transient inactivation of the CeA prevented cocaine-seeking behaviour. Administration of CP154526 to the CeA prevented OrxAinduced cocaine-seeking behaviour. Limitations: The use of only male rats could have been a limitation. Other limitations could have been the use of an indirect approach to test the hypothesis that administration of OrxA to the pPVT drives cocaine-seeking via CRF1 signalling in the CeA, and a lack of analysis of the participation of CeA subregions. Conclusion: Cocaine-seeking behaviour induced by OrxA administered to the pPVT is driven by activation of the CeA via CRF1 signalling.

Altered Gut Microbiota is Involved in the Anti-Hypertensive Effects of Vitamin C in Spontaneously Hypertensive Rat.

SCOPE: Gut dysbiosis and dysregulation of the gut-brain-axis contributes to the pathogenesis of hypertension. Vitamin C (VC) is a common dietary supplement that shows the ability to lower the elevated blood pressure in hypertensive animals. Thus, the hypothesis that the gut microbiota is involved in the anti-hypertensive effect of VC is proposed. METHODS AND RESULTS: The changes of the gut microbiota and pathology in a spontaneously hypertensive rat (SHR) model after daily oral intake of VC in dosage of 200 or 1000 mg kg-1 are examined. After 4 weeks, the elevated blood pressure of SHRs in both VC-treated groups is attenuated. Sequencing of the gut microbiota shows improvement in its diversity and abundance. Bioinformatic analysis suggests restored metabolism and biosynthesis-related functions of the gut, which are confirmed by the improvement of gut pathology and integrity. Analysis of the hypothalamus paraventricular nucleus (PVN), the central pivot of blood pressure regulation, also shows reduced inflammatory responses and oxidative stress. CONCLUSIONS: The reduced blood pressure, enriched gut microbiota, improved gut pathology and integrity, and reduced inflammatory responses and oxidative stress in the PVN together suggest that the anti-hypertensive effects of VC involve reshaping of gut microbiota composition and function.

An Angiotensin-Responsive Connection from the Lamina Terminalis to the Paraventricular Nucleus of the Hypothalamus Evokes Vasopressin Secretion to Increase Blood Pressure in Mice.

Blood pressure is controlled by endocrine, autonomic, and behavioral responses that maintain blood volume and perfusion pressure at levels optimal for survival. Although it is clear that central angiotensin type 1a receptors (AT1aR; encoded by the Agtr1a gene) influence these processes, the neuronal circuits mediating these effects are incompletely understood. The present studies characterize the structure and function of AT1aR neurons in the lamina terminalis (containing the median preoptic nucleus and organum vasculosum of the lamina terminalis), thereby evaluating their roles in blood pressure control. Using male Agtr1a-Cre mice, neuroanatomical studies reveal that AT1aR neurons in the area are largely glutamatergic and send projections to the paraventricular nucleus of the hypothalamus (PVN) that appear to synapse onto vasopressin-synthesizing neurons. To evaluate the functionality of these lamina terminalis AT1aR neurons, we virally delivered light-sensitive opsins and then optogenetically excited or inhibited the neurons while evaluating cardiovascular parameters or fluid intake. Optogenetic excitation robustly elevated blood pressure, water intake, and sodium intake, while optogenetic inhibition produced the opposite effects. Intriguingly, optogenetic excitation of these AT1aR neurons of the lamina terminalis also resulted in Fos induction in vasopressin neurons within the PVN and supraoptic nucleus. Further, within the PVN, selective optogenetic stimulation of afferents that arise from these lamina terminalis AT1aR neurons induced glutamate release onto magnocellular neurons and was sufficient to increase blood pressure. These cardiovascular effects were attenuated by systemic pretreatment with a vasopressin-1a-receptor antagonist. Collectively, these data indicate that excitation of lamina terminalis AT1aR neurons induces neuroendocrine and behavioral responses that increase blood pressure.SIGNIFICANCE STATEMENT Hypertension is a widespread health problem and risk factor for cardiovascular disease. Although treatments exist, a substantial percentage of patients suffer from "drug-resistant" hypertension, a condition associated with increased activation of brain angiotensin receptors, enhanced sympathetic nervous system activity, and elevated vasopressin levels. The present study highlights a role for angiotensin Type 1a receptor expressing neurons located within the lamina terminalis in regulating endocrine and behavioral responses that are involved in maintaining cardiovascular homeostasis. More specifically, data presented here reveal functional excitatory connections between angiotensin-sensitive neurons in the lamina terminals and vasopressin neurons in the paraventricular nucleus of the hypothalamus, and further indicate that activation of this circuit raises blood pressure. These neurons may be a promising target for antihypertensive therapeutics.

The PGE2 receptor EP3 plays a positive role in the activation of hypothalamic-pituitary-adrenal axis and neuronal activity in the hypothalamus under immobilization stress.

BACKGROUND: Prostaglandin E2 (PGE2) binds to four receptor subtypes (EP1, EP2, EP3 and EP4) and plays an important role in response to stress. However, the identity of the receptor(s) responsible for PGE2 regulation of neuronal activity and signaling through activation of the hypothalamic-pituitary-adrenal (HPA) axis under immobilization stress is unknown. PURPOSE: The present study aimed to investigate the role of the hypothalamic PGE2 receptors in the activation of the HPA axis and neuronal activity in a rat model of stress. METHODS: Stress was induced by immobilization of the animals, after which the stress-induced profile of PGE2 receptor signaling in the rat hypothalamus was determined by real-time polymerase chain reaction and immunohistochemistry. The effect of a selective EP3 receptor antagonist on corticosterone concentrations and c-Fos immunoreactivity was measured. RESULTS: Expression of EP2 and EP3 receptor genes, but not EP1 and EP4, was increased following immobilization stress. The EP3 receptor was localized to the paraventricular nucleus (PVN) of the hypothalamus, and the integrated density of the EP3 receptor was increased after immobilization stress. Rats given L-798,106, a selective antagonist of the EP3 receptor, showed significant attenuation of stress-increased serum corticosterone levels. EP3 antagonist also significantly suppressed the increase in the gene expression of c-Fos and the number of c-Fos-immunoreactive cells in the PVN of the hypothalamus following immobilization stress. CONCLUSIONS: These results suggest that immobilization stress may result in increased activation of the HPA axis and neuronal activity through regulating the function of the EP3 receptor.

Identification of substances which regulate activity of corticotropin-releasing factor-producing neurons in the paraventricular nucleus of the hypothalamus.

The stress response is a physiological system for adapting to various internal and external stimuli. Corticotropin-releasing factor-producing neurons in the paraventricular nucleus of the hypothalamus (PVN-CRF neurons) are known to play an important role in the stress response as initiators of the hypothalamic-pituitary-adrenal axis. However, the mechanism by which activity of PVN-CRF neurons is regulated by other neurons and bioactive substances remains unclear. Here, we developed a screening method using calcium imaging to identify how physiological substances directly affect the activity of PVN-CRF neurons. We used acute brain slices expressing a genetically encoded calcium indicator in PVN-CRF neurons using CRF-Cre recombinase mice and an adeno-associated viral vector under Cre control. PVN-CRF neurons were divided into ventral and dorsal portions. Bath application of candidate substances revealed 12 substances that increased and 3 that decreased intracellular calcium concentrations. Among these substances, angiotensin II and histamine mainly increased calcium in the ventral portion of the PVN-CRF neurons via AT1 and H1 receptors, respectively. Conversely, carbachol mainly increased calcium in the dorsal portion of the PVN-CRF neurons via both nicotinic and muscarinic acetylcholine receptors. Our method provides a precise and reliable means of evaluating the effect of a substance on PVN-CRF neuronal activity.

Presynaptic glutamatergic transmission and feedback system of oxytocinergic neurons in the hypothalamus of a rat model of adjuvant arthritis.

The neurohypophysial hormone oxytocin (OXT) is synthesized in the hypothalamic paraventricular and supraoptic nuclei. Recently, some studies have considered OXT to be important in sensory modulation and that the OXT protein is upregulated by acute and chronic nociception. However, the mechanism by which OXT is upregulated in neurons is unknown. In this study, we examined the resting membrane potentials and excitatory postsynaptic currents in OXT-ergic neurons in the paraventricular nucleus in adjuvant arthritis rat model, a model of chronic inflammation, using whole-cell patch-clamping. Transgenic rats expressing OXT and monomeric red fluorescent protein 1 (mRFP1) fusion protein to visualize the OXT-ergic neurons were used, and the OXT-mRFP1 transgenic rat model of adjuvant arthritis was developed by injection of heat-killed Mycobacterium butyricum. Furthermore, the feedback system of synthesized OXT was also examined using the OXT receptor antagonist L-368,899. We found that the resting membrane potentials and frequency of miniature excitatory postsynaptic currents and spontaneous excitatory postsynaptic currents in OXT-monomeric red fluorescent protein 1 neurons in the paraventricular nucleus were significantly increased in adjuvant arthritis rats. Furthermore, L-368,899 dose-dependently increased the frequency of miniature excitatory postsynaptic currents and spontaneous excitatory postsynaptic currents in OXT-ergic neurons. Following bath application of the GABAA receptor antagonist picrotoxin and the cannabinoid receptor 1 antagonist AM 251, L-368,899 still increased the frequency of miniature excitatory postsynaptic currents. However, following bath application of the nitric oxide synthase inhibitor Nω-Nitro-L-arginine methyl ester hydrochloride, L-368,899 did not alter the miniature excitatory postsynaptic current frequency. Thus, it is suggested that OXT-ergic neuron activity is upregulated via an increase in glutamate release, and that the upregulated OXT neurons have a feedback system with released endogenous OXT. It is possible that nitric oxide, but not GABA, may contribute to the feedback system of OXT neurons in chronic inflammation.

Amphetamine-induced sensitization of hypertension and lamina terminalis neuroinflammation.

Psychomotor stimulants are prescribed for many medical conditions, including obesity, sleep disorders, and attention-deficit/hyperactivity disorder. However, despite their acknowledged therapeutic utility, these stimulants are frequently abused, and their use can have both short- and long-term negative consequences. Although stimulants such as amphetamines acutely elevate blood pressure, it is unclear whether they cause any long-term effects on cardiovascular function after use has been discontinued. Previous work in our laboratory has demonstrated that physiological and psychosocial stressors will produce sensitization of the hypertensive response, a heightened pressor response to a hypertensinogenic stimulus delivered after stressor exposure. Here, we tested whether pretreatment with amphetamine for 1 wk can sensitize the hypertensive response in rats. We found that repeated amphetamine administration induced and maintained sensitization of the pressor response to angiotensin II following a 7-day delay after amphetamine injections were terminated. We also found that amphetamine pretreatment altered mRNA expression for molecular markers associated with neuroinflammation and renin-angiotensin-aldosterone system (RAAS) activation in the lamina terminalis, a brain region implicated in the control of sympathetic nervous system tone and blood pressure. The results indicated amphetamine upregulated mRNA expression underlying neuroinflammation and, to a lesser degree, message for components of the RAAS in the lamina terminalis. However, we found no changes in mRNA expression in the paraventricular nucleus. These results suggest that a history of stimulant use may predispose individuals to developing hypertension by promoting neuroinflammation and upregulating activity of the RAAS in the lamina terminalis.

Oxytocin and CD38 in the paraventricular nucleus play a critical role in paternal aggression in mice.

In mammals, the development of healthy offspring requires maternal care. Behavior by lactating mothers toward other individuals is an important component of maternal aggression. However, it is unclear whether fathers display aggression primed by pups (an external factor), and the protection mechanism is poorly understood. To address this question, we examined paternal aggression in the ICR mouse strain. We found that sires exposed to cues from pups and lactating dams showed stronger aggression toward intruders than did sires that were deprived of family cues or exposed to nonlactating mates. c-Fos immunohistochemistry showed that cells in both the paraventricular and supraoptic nuclei (PVN and SON, respectively) in the hypothalamus of sires exposed to any cues were highly activated. However, c-Fos activation in oxytocinergic neurons was increased only in sires exposed to pup cues and solely in the PVN. In Cd38-knockout sires, the presence of pups induced no or reduced parental aggression; however, this phenotype was recovered, that is, aggression increased to the wild-type level, after intraperitoneal administration of oxytocin (OT). Specific c-Fos activation patterns induced by pup cues were not found in the PVN of knockout sires. These results demonstrate that the PVN is one of the primary hypothalamic areas involved in paternal aggression and suggest that a CD38-dependent OT mechanism in oxytocinergic neurons is critical for part of the behavior associated with the protection of offspring by nurturing male mice.

The anorexigenic effect of neuropeptide K in chicks involves the paraventricular nucleus and arcuate nucleus of the hypothalamus.

Neuropeptide K (NPK) induces satiety in birds and mammals. We demonstrated that in birds this effect was associated with the hypothalamus, but beyond this little is known in any species regarding the central mechanism of action. Thus, this study was designed to identify hypothalamic molecular mechanisms associated with the food intake-inhibiting effects of NPK in chicks. In Experiment 1, intracerebroventricular (ICV) injection of 1.0 and 3.0 nmol of NPK reduced food intake and we identified an effective dose for microinjection. In Experiment 2, food intake was reduced when NPK was microinjected into the PVN. In Experiment 3, whole hypothalamus was collected from chicks at 1 h post-ICV NPK injection. The abundance of corticotropin-releasing factor (CRF) and agouti-related peptide (AgRP) mRNA was reduced in NPK-injected chicks. In Experiment 4, within the isolated paraventricular nucleus (PVN) there was less CRF mRNA, and within the arcuate nucleus (ARC) there was less AgRP mRNA, in NPK- than vehicle-treated chicks at 1 h post-injection. We conclude that there are first order neurons for NPK that reside within the PVN, and the anorexigenic effect of NPK is associated with a decrease in AgRP in the ARC.

Acupuncture reduces nicotine-induced norepinephrine release in the hypothalamus via the solitary NMDA receptor/NOS pathway.

Noradrenergic projections from the nucleus tractus solitarius (NTS) to the hypothalamic paraventricular nucleus (PVN) are involved in nicotine (Nic) dependence. Nic induces hypothalamic norepinephrine (NE) release through N-methyl-d-aspartate receptors (NMDARs) and nitric oxide in the NTS. However, acupuncture attenuates Nic withdrawal-induced anxiety. Therefore, this study investigated the effects of acupuncture on Nic-induced hypothalamic NE release. Rats received an intravenous infusion of Nic (90 µg/kg, over 60 s) and extracellular NE levels in the PVN were determined by in vivo microdialysis. Immediately after Nic administration, the rats were bilaterally treated with acupuncture at acupoint HT7 (Shen-Men) or PC6 (Nei-Guan), or a non-acupoint (tail) for 60 s. Acupuncture at HT7, but not at PC6 or the tail, significantly reduced Nic-induced NE release. However, this was abolished by a post-acupuncture infusion of either NMDA or sodium nitroprusside into the NTS. Additionally, acupuncture at HT7, but not the control points, prevented Nic-induced plasma corticosterone secretion and inhibited Nic-induced increases in the phosphorylation of neuronal nitric oxide synthase (nNOS) and endothelial NOS in the NTS. These findings suggest that acupuncture at HT7 reduces Nic-induced NE release in the PVN via inhibition of the solitary NMDAR/NOS pathway.

Estrogen receptor beta and G protein-coupled estrogen receptor 1 are involved in the acute estrogenic regulation of arginine-vasopressin immunoreactive levels in the supraoptic and paraventricular hypothalamic nuclei of female rats.

The ovarian hormone 17ß-estradiol is known to regulate the release, expression and immunoreactivity of arginine-vasopressin (AVP) in the supraoptic and paraventricular hypothalamic nuclei of rodents. Previous studies have shown that estrogen receptor α is involved in the effects of chronic estradiol administration on arginine-vasopressin immunoreactivity in the female rat hypothalamus. In this study we have examined the effect of an acute administration of estradiol or specific agonists for estrogen receptors α, ß and G protein-coupled estrogen receptor 1 on the immunoreactivity of arginine-vasopressin in the hypothalamus of adult ovariectomized female rats. Acute estradiol administration resulted in a significant decrease in the number of arginine-vasopressin immunoreactive neurons in the supraoptic and paraventricular nuclei after 24 h. The effects of the specific estrogen receptors agonists suggest that the action of estradiol on arginine-vasopressin immunoreactivity is mediated in the supraoptic nucleus by G protein-coupled estrogen receptor 1 and in the paraventricular nucleus by both estrogen receptor ß and G protein-coupled estrogen receptor 1. Thus, in contrast to previous studies on the effect of chronic estrogenic treatments, the present findings suggest that estrogen receptor ß and G protein-coupled estrogen receptor 1 mediate the acute effects of estradiol on arginine-vasopressin immunoreactivity in the hypothalamus of ovariectomized rats.

Morphological analysis for neuronal pathway from the hindbrain ependymocytes to the hypothalamic kisspeptin neurons.

Hindbrain ependymocytes are postulated to have a glucose-sensing role in regulating gonadal functions. Previous studies have suggested that malnutrition-induced suppression of gonadotropin secretion is mediated by noradrenergic inputs from the A2 region in the solitary tract nucleus to the paraventricular nucleus (PVN), and by corticotropin-releasing hormone (CRH) release in the hypothalamus. However, no morphological evidence to indicate the neural pathway from the hindbrain ependymocytes to hypothalamic kisspeptin neurons, a center for reproductive function in mammals, currently exists. The present study aimed to examine the existence of a neuronal pathway from the hindbrain ependymocytes to kisspeptin neurons in the arcuate nucleus (ARC) and anteroventral periventricular nucleus (AVPV). To determine this, wheat-germ agglutinin (WGA), a trans-synaptic tracer, was injected into the fourth ventricle (4V) in heterozygous Kiss1-tandem dimer Tomato (tdTomato) rats, where kisspeptin neurons were visualized by tdTomato fluorescence. 48 h after the WGA injection, brain sections were taken from the forebrain, midbrain and hindbrain and subjected to double immunohistochemistry for WGA and dopamine ß-hydroxylase (DBH) or CRH. WGA immunoreactivities were found in vimentin-immunopositive ependymocytes of the 4V and the central canal (CC), but not in the third ventricle. The WGA immunoreactivities were detected in some tdTomato-expressing cells in the ARC and AVPV, DBH-immunopositive cells in the A1-A7 noradrenergic nuclei, and CRH-immunopositive cells in the PVN. These results suggest that the hindbrain ependymocytes have neuronal connections with the kisspeptin neurons, most probably via hindbrain noradrenergic and CRH neurons to relay low energetic signals for regulation of reproduction.

Effect of Dp71 deficiency on the oxytocin hypothalamic axis in osmoregulation function in mice.

Dp71 is the major form of dystrophins (Dp) in the supraoptic nucleus (SON) and in the neural lobe of hypophysis (NL/HP). Dp71-null mice exhibit a hypo-osmolar status attributed to an altered osmosensitivity of the SON and to a perturbed vasopressinergic axis. Because oxytocin (OT) is implicated in osmoregulation via natriuresis, this study explored the oxytocinergic axis in Dp71-null mice after salt-loading (SL). Under normosmolar conditions, OT-mRNA expression was higher in the Dp71-null SON compared to wild-type (wt) and the OT peptide level has not changed. Dp-immunostaining was localized in astrocytes end-feet surrounding vessels in wt SON. This distribution changed in Dp71-null SON, Dp being detected in OT-soma of MCNs. nNOS and NADPH-diaphorase levels increased in the OT area of the Dp71-null SON compared to wt. In the NL/HP, OT level reduced in Dp71-null mice and Dp localization changed from pituicytes end-feet in wt SON to OT terminals in Dp71-null SON. Salt-Loading resulted in an increase of OT-mRNA and peptide levels in wt SON but had no effect in Dp71-null SON. In the NL/HP, OT content was reduced after SL. For Dp71-null mice, OT level, already low in control, was not modified by SL. Dp level was not affected by SL in the SON nor in the NL/HP. Our data confirmed the importance of Dp71 for the SON functionality in osmoregulation. The localization of Dp71 at the glial-vascular interface could be associated with SON osmosensitivity, leading to an adequate OT synthesis in the SON and release from the NL/HP upon plasmatic hyperosmolality.

Chronic infusion of berberine into the hypothalamic paraventricular nucleus attenuates hypertension and sympathoexcitation via the ROS/Erk1/2/iNOS pathway.

BACKGROUND: Berberine (BBR), a Chinese traditional herbal medicine, has many pharmacologic benefits such as anti-inflammation and anti-oxidation. It is widely used in clinical treatment of cardiovascular diseases such as hypertension. However, the mechanism of how BBR attenuates hypertension through affecting central neural system is not clear. PURPOSE: This study was designed to determine whether chronic infusion of BBR into the hypothalamic paraventricular nucleus (PVN) attenuates hypertension and sympathoexcitation via the ROS/Erk1/2/iNOS pathway. METHODS: Two-kidney, one-clip (2K1C) renovascular hypertensive rats were randomly assigned and treated with bilateral PVN infusion of BBR (2µg/h) or vehicle (artificial cerebrospinal fluid) via osmotic minipumps for 28 days. RESULTS: 2K1C rats showed higher mean arterial pressure (MAP) and PVN Fra-like activity, plasma levels of norepinephrine (NE), PVN levels of NOX2, NOX4, Erk1/2 and iNOS, and lower PVN levels of copper/zinc superoxide dismutase (Cu/Zn-SOD). Chronic infusion of BBR reduced MAP, PVN Fra-like activity and plasma levels of NE, reduced NOX2, NOX4, Erk1/2, iNOS and induced Cu/Zn-SOD in the PVN. CONCLUSIONS: These results suggest that BBR attenuates hypertension and sympathoexcitation via the ROS/Erk1/2/iNOS pathway in 2K1C renovascular hypertensive rats.

Resetting the Stress System with a Mifepristone Challenge.

Psychotic depression is characterized by elevated circulating cortisol, and high daily doses of the glucocorticoid/progesterone antagonist mifepristone for 1 week are required for significant improvement. Using a rodent model, we find that such high doses of mifepristone are needed because the antagonist is rapidly degraded and poorly penetrates the blood-brain barrier, but seems to facilitate the entry of cortisol. We also report that in male C57BL/6J mice, after a 7-day treatment with a high dose of mifepristone, basal blood corticosterone levels were similar to that of vehicle controls. This is surprising because after the first mifepristone challenge, corticosterone remained elevated for about 16 h, and then decreased towards vehicle control levels at 24 h. At that time, stress-induced corticosterone levels of the 1xMIF were sevenfold higher than the 7xMIF group, the latter response being twofold lower than controls. The 1xMIF mice showed behavioral hyperactivity during exploration of the circular hole board, while the 7xMIF mice rather engaged in serial search patterns. To explain this rapid reset of corticosterone secretion upon recurrent mifepristone administration, we suggest the following: (i) A rebound glucocorticoid feedback after cessation of mifepristone treatment. (ii) Glucocorticoid agonism in transrepression and recruitment of cell-specific coregulator cocktails. (iii) A more prominent role of brain MR function in control of stress circuit activity. An overview table of neuroendocrine MIF effects is provided. The data are of interest for understanding the mechanistic underpinning of stress system reset as treatment strategy for stress-related diseases.

Neuronal nitric oxide synthase activity mediates Lycium barbarum polysaccharides-enhanced sexual performance without stimulating noncontact erection in rats.

RATIONALE: Lycium barbarum polysaccharide (LBP) is known to promote reproductive functions. However, its role in noncontact erection (NCE) of penis initiated by brain regions including medial preoptic area (MPOA) and paraventricular nucleus (PVN) regions responsible for sexual behavior has not been investigated. OBJECTIVES: Therefore, this study initially investigated the effects of LBP on male sexual function, and subsequently, the mechanistic insight was investigated through assessing the expression of neuronal nitric oxide synthase (nNOS) in the MPOA and PVN. METHODS: The adult male rats were treated with 100 mg/kg of LBP or vehicle by oral gavage. Before and after 14 days of treatment, copulatory behavior and noncontact erection (NCE) were recorded. After the last behavioral test, the brain was isolated to measure nNOS expression in the MPOA and PVN. RESULTS: Data showed that LBP treatment significantly increased both the frequencies of intromission as well as ejaculation, compared to the control group. Whereas, a reduced post-ejaculatory interval was observed compared to same group on day 0. Furthermore, the treatment led to an increased intromission ratio, inter-intromission interval, and the number of MPOA nNOS-immunoreactive cells (nNOS-ir). Additionally, a significantly positive correlation between ejaculation frequency and MPOA nNOS-ir cells was recorded. Of note, LBP treatment had no effects on NCE and PVN nNOS-ir expression. CONCLUSION: These findings suggest that LBP enhances sexual behavior through increased nNOS expression in the MPOA in male rats.

Central action of CART induces neuronal activation in the paraventricular and dorsomedial hypothalamus of diet-induced obese and lean mice.

The cocaine- and amphetamine-regulated transcript (CART) is a peptide commonly studied in the feeding behavior, but it exerts an important role in the autonomic and cardiovascular control as well. It is known that exogenous administration of CART in the central nervous system can elicit increase in blood pressure of both conscious and anesthetized rodents, but little is known whether these central effects might differ between lean and obese animals. We have recently shown that diet-induced obese mice that developed hypertension presented an upregulation of CART levels in the dorsomedial nucleus of hypothalamus, while obese normotensive ones had not. Herein we investigate whether the central action of CART could activate differently the hypothalamic nuclei of diet-induced obese mice compared to the lean counterparts by using Fos protein expression, C57BL/6 mice were randomly assigned to two cohorts, one fed with a high-fat diet for 8 weeks (obese), and the other fed with regular rodent chow (lean). Both groups received an ICV injection of CART at the dose of 400µM, 1 mM or vehicle. Subsequently, the brains were processed for Fos protein immunohistochemical in order to identify hypothalamic neuronal activation. Significantly greater numbers of Fos-positive neurons were observed in the PVN and DMH of obese mice that received CART 1 mM, when compared to the lean control. These results indicate that the central action of CART induces neuronal activation in the hypothalamic nuclei of obese and lean mice, and this could be relevant to the different autonomic and cardiovascular adjustments that an organism exposed to different diet and metabolic condition.

Estrogen effects on oxytocinergic pathways that regulate food intake.

Multiple stimulatory and inhibitory neural circuits control eating, and these circuits are influenced by an array of hormonal, neuropeptide, and neurotransmitter signals. For example, estrogen and oxytocin (OT) both are known to decrease food intake, but the mechanisms by which these signal molecules influence eating are not fully understood. These studies investigated the interaction between estrogen and OT in the control of food intake. RT-qPCR studies revealed that 17ß-estradiol benzoate (EB)-treated rats showed a two-fold increase in OT mRNA in the paraventricular nucleus of the hypothalamus (PVN) compared to Oil-treated controls. Increased OT mRNA expression may increase OT protein levels, and immunohistochemistry studies showed that EB-treated rats had more intense OT labeling in the nucleus of the solitary tract (NTS), a region known to integrate signals for food intake. Food intake measurements showed that EB treatment reduced food intake, as expected. EB-treated rats lost weight over the course of the experiment, as expected, and EB-treated rats that received the highest dose of OT lost more weight than EB-treated rats that did not receive OT. Finally, OT antagonist administered to EB-treated rats reversed the effect of EB on food intake, suggesting that estrogen effects to decrease food intake may involve the oxytocinergic pathway.

Acylated ghrelin suppresses the cytokine response to lipopolysaccharide and does so independently of the hypothalamic-pituitary-adrenal axis.

Ghrelin, one of the major metabolic hormones involved in controlling energy balance, has recently been shown to have other properties including regulating the hypothalamic-pituitary-adrenal (HPA) axis response to psychological stress and being a potent anti-inflammatory agent. Ghrelin's HPA axis and anti-inflammatory actions have previously been identified as principally due to the acylated form (AG). However, our recent work has also suggested a role for des-acylated ghrelin (DAG) in these functions. Here we hypothesized ghrelin's anti-inflammatory activity is mediated by the HPA axis and this effect is differentially executed by AG and DAG. We gave adult male Wistar rats a concomitant injection of AG or DAG and lipopolysaccharide (LPS) and measured their effects on circulating cytokines, stress hormones and neuronal activation of the paraventricular nucleus of the hypothalamus (PVN). AG, but not DAG significantly suppressed the pro- and anti-inflammatory cytokine response induced by LPS in vivo. DAG also had no effects on any components of the HPA axis. AG, despite stimulating neuronal activation in the PVN in vivo and stimulating ACTH release from the pituitary in vitro, did not affect the HPA axis response to LPS. These findings suggest AG's anti-inflammatory effects are independent of its actions on the HPA axis and have implications for the potential use of this peptide for treatment of inflammatory conditions without compromising HPA axis activity.

Serotonin receptor regulation as a potential mechanism for sexually dimorphic oxytocin dysregulation in a model of Autism.

Perinatal administration of serotonin (5HT) agonist 5-methoxytryptamine (5MT) induces developmental hyperserotonemia (DHS; elevated blood serotonin) and produces behavioral and neurochemical changes in rats relevant to Autism Spectrum Disorder (ASD), such as oxytocin dysregulation. Disruption of the oxytocin system may underlie many of the social deficits present in ASD individuals, thus we investigated the mechanism(s) underlying DHS-induced oxytocin dysregulation. The most parsimonious mechanism of 5HT action would be alteration of 5HT receptors on oxytocin cells; 5HT is known to influence cell survival as well as influence oxytocin release via 5HT1A and 5HT2A receptors, which co-localize in oxytocin-expressing (OXT+) cells in the paraventricular nucleus (PVN) of the hypothalamus. We report that both male and female DHS rats have a lower percentage of OXT+ cells co-localized with excitatory 5HT2A receptors than control animals, while only DHS females have a higher percentage of OXT+ cells co-localized with inhibitory 5HT1A receptors compared to controls. Importantly, DHS also reduces the number of OXT+ cells in the PVN of adult male, but not female, rats. This pattern suggests that females, but not males, can regulate 5HT receptors in response to DHS in a manner that promotes oxytocin cell survival and functional efficiency. In addition, it has been previously reported that DHS alters normal juvenile play, especially in males, thus we also tested play partner preference among juvenile control and DHS males. Sex differences observed using the DHS model of ASD add to its validity, given the pronounced male sex bias in the prevalence of ASD, and emphasize the need for inclusion of both sexes in ASD research.