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Objective To probe the spatial distribution of respiratory-related nuclei throughout the whole brain, and to investigate the circuit and projection links between the paraventricular hypothalamic nucleus (PVN) and the preBötzinger complex (preBötC) in mice. Methods Using a retrograde tracing strategy from peripheral to central, pseudorabies viruses (PRV) were microinjected into the diaphragm of mice to retrogradely label the upstream inputs brain regions from spinal respiratory phrenic motor neurons. The anterograde non-transsynaptic tracing viruses (AAV2/9-EGFP) were microinjected into the PVN to label the downstream brain region of the PVN; And using the Cre/loxP gene expression strategy, Dio-EGFP and Retro-Cre viruses were injected in the PVN and preBötC, respectively, to validate the PVN-preBötC circuit and the projection relationship between the two. Results The results of PRV-mediated retrograde tracing showed that PRV-EGFP-positive neurons were distributed in several brain regions such as PVN, preBötC, nucleus tractus solitarii (NTS), locus coeruleus (LC) and periaqueductal grey (PAG). Except for the medulla oblongata region, the PVN brain region had the highest percentage of PRV-EGFP-positive neurons (P < 0.000 1). After AAV2/9-EGFP viruses were injected in the PVN, green neurofibrils were observed in the preBötC brain region after viral infection; Dio-EGFP and Retro-Cre viruses were injected in the PVN and preBötC, respectively, and green neuronal cells were seen in the brain region of the PVN after viral infection. Conclusion PVN is an important upstream input nucleus of spinal respiratory phrenic motor neurons, and has the potential to regulate respiration through direct projections from PVN-preBötC circuit.

Understanding of central nervous system mechanisms underlying age-related infertility remains limited. Fibril α-synuclein, distinct from its monomeric form, is implicated in age-related diseases. Notably, fibril α-synuclein spreads among neurons, similar to prions, from damaged old neurons in cortex and hippocampus to healthy neurons. However, less is known whether α-synuclein propagates into oxytocin neurons, which play crucial roles in reproduction. We compared α-synuclein expression in the oxytocin neurons in suprachiasmatic nucleus (SCN), supraoptic nucleus (SON), paraventricular hypothalamic nucleus (PVN), and posterior pituitary (PP) gland of healthy heifers and aged cows to determine its role in age-related infertility. We analyzed mRNA and protein expression, along with Congo red histochemistry and fluorescent immunohistochemistry for oxytocin and α-synuclein, followed by confocal microscopy with Congo red staining. Both mRNA and protein expressions of α-synuclein were confirmed in the bovine cortex, hippocampus, SCN, SON, PVN, and PP tissues. Significant differences in α-synuclein mRNA expressions were observed in the cortex and hippocampus between young heifers and old cows. Western blots showed five bands of α-synuclein, probably reflecting monomers, dimers, and oligomers, in the cortex, hippocampus, SCN, SON, PVN, and PP tissues, and there were significant differences in some bands between the young heifers and old cows. Bright-field and polarized light microscopy did not detect obvious amyloid deposition in the aged hypothalami; however, higher-sensitive confocal microscopy unveiled strong positive signals for Congo red and α-synuclein in oxytocin neurons in the aged hypothalami. α-synuclein was expressed in oxytocin neurons, and some differences were observed between young and old hypothalami.

Objective(s): High levels of resistin are associated with metabolic diseases and their complications, including hypertension. The paraventricular nucleus (PVN) is also involved in metabolic disorders and cardiovascular diseases, such as hypertension. Therefore, this study aimed to study cardiovascular (CV) responses evoked by the injection of resistin into the lateral ventricle (LV) and PVN and determine the mechanism of these responses in the rostral ventrolateral medulla (RVLM).Materials and Methods: Arterial pressure (AP) and heart rate (HR) were evaluated in urethane-anesthetized male rats (1.4 g/kg intraperitoneally) before and after all injections. This study was carried out in two stages. Resistin was injected into LV at the first stage, and AP and HR were evaluated. After that, the paraventricular, supraoptic, and dorsomedial nuclei of the hypothalamus were chosen to evaluate the gene expression of c-Fos. Afterward, resistin was injected into PVN, and cardiovascular responses were monitored. Then to detect possible neural mechanisms of resistin action, agonists or antagonists of glutamatergic, GABAergic, cholinergic, and aminergic transmissions were injected into RVLM. Results: Resistin injection into LV or PVN could increase AP and HR compared to the control group and before injection. Resistin injection into LV also increases the activity of RVLM, paraventricular, supraoptic, and dorsomedial areas. Moreover, the CV reflex created by the administration of resistin in PVN is probably mediated by glutamatergic transmission within RVLM. Conclusion: It can be concluded that hypothalamic nuclei, including paraventricular, are important central areas for resistin actions, and glutamatergic transmission in RVLM may be one of the therapeutic targets for high AP in obese people or with metabolic syndrome.

Abstract Self-grooming plays an essential role in hygiene maintenance, thermoregulation, and stress response. However, the neural populations involved in self-grooming remain largely unknown. The paraventricular hypothalamic nucleus (PVH) has been implicated in the regulation of self-grooming. Arginine vasopressin-producing neurons are among the major neuronal populations in the PVH (PVHAVP), which play important roles in water homeostasis, blood pressure regulation, feeding, and stress response. Here, we report the critical role of PVHAVP neurons in the induction of self-grooming. Optogenetic activation of PVHAVP neurons immediately induced self-grooming in freely moving mice. Chemogenetic activation of these neurons also increased time spent self-grooming. In contrast, their chemogenetic inhibition significantly reduced naturally occurring self-grooming, suggesting that PVHAVP-induced grooming has physiological relevance. Notably, optogenetic activation of PVHAVP neurons triggered self-grooming over other adaptive behaviors, such as voracious feeding induced by fasting and social interaction with female mice. Thus, our study proposes the novel role of PVHAVP neurons in regulating self-grooming behavior and, consequently, hygiene maintenance and stress response. Furthermore, uncontrolled activation of these neurons may be potentially relevant to diseases characterized by compulsive behaviors and impaired social interaction, such as autism, obsessive–compulsive disorder, and anorexia nervosa.

Background: The Paraventricular Hypothalamic Nucleus (PVN) coordinates autonomic and neuroendocrine systems to maintain homeostasis. Microinjection of angiotensin II (AngII) into the PVN has been previously shown to produce pressor and bradycardia responses. Anatomical evidence has indicated that a substantial proportion of PVN neurons is connected with the neurons in the central amygdala (CeA). The present study aimed to examine the possible contribution of the CeA in cardiovascular responses evoked by microinjection of AngII into the parvocellular portion of PVN (PVNp) before and after microinjection of cobalt chloride (CoCl2) into the CeA.Methods: The experiments were conducted at the Department of Physiology of Shiraz University of Medical Sciences, from April 2019 to November 2019. There were two groups of 21 eight-week-old urethane anesthetized male rats, namely saline (n=9 rats) and AngII (n=12 rats) groups. Drugs (100 nL) were microinjected via a single-glass micropipette into the PVNp and CeA. Their blood pressure (BP) and heart rate (HR) were recorded throughout the experiments. The mean arterial pressure (MAP) and heart rate (HR) were compared to the pre-injection values using paired t test, and to those of the saline group using independent t test.Results: Microinjection of AngII into the PVNp produced pressor response (P

Abstract The nucleus of the solitary tract (NTS) plays a crucial role in integrating peripheral information regarding visceral functions. Glutamate decarboxylase 2 (GAD2) inhibitory neurons are abundant in the NTS, and are known to form local and short-range projections within the NTS and nearby hindbrain areas. Here we performed whole-brain mapping of outputs from GAD2 neurons in the NTS using cell-type specific viral labeling together with ultrahigh-speed 3D imaging at 1-μm resolution. In addition to well-known targets of NTS GAD2 neurons including the principle sensory nucleus of the trigeminal (PSV), spinal nucleus of the trigeminal (SPV), and other short-range targets within the hindbrain, the high sensitivity of our system helps reveal previously unknown long-range projections that target forebrain regions, including the bed nuclei of the stria terminalis (BST) involved in stress and fear responses, and the paraventricular hypothalamic nucleus (PVH) involved in energy balance and stress-related neuroendocrine responses. The long-range projections were further verified by retrograde labeling of NTS GAD2 neurons with cholera toxin B (CTB) injections in the BST and PVH, and by Cre-dependent retrograde tracing with rAAV2-retro injections in the two regions of GAD2-Cre mice. Finally, we performed complete morphological reconstruction of several sparsely labeled neurons projecting to the forebrain and midbrain. These results provide new insights about how NTS might participate in physiological and emotional modulation.

Objective(s): Resistin, as a 12.5 kDa cysteine-rich polypeptide, is expressed in hypothalamus and regulates sympathetic nerve activity. It is associated with obesity, metabolic syndrome and cardiovascular diseases. In this study, we investigated the neural pathway of cardiovascular responses induced by injection of resistin into paraventricular nucleus (PVN) with rostral ventrolateral medulla (RVLM). Materials and Methods: Adult male rats were anesthetized with urethane (1.4 g/kg intraperitoneally). Resistin (3 µg/1 µl/rat) was first injected into PVN, and the glutamatergic, corticotrophin-releasing factor (CRF)-ergic and angiotensinogenic transmission was inhibited by injecting of their antagonist in RVLM. Arterial pressure (AP) and heart rate (HR) were monitored before and after the injection. Results: The results showed that resistin injection into PVN significantly increased AP and HR compared to control group and prior to its injection (PConclusion: It can be concluded that glutamatergic and CRFergic transmissions have crucial contribution to cardiovascular responses produced by resistin. The results provided new and potentially important insight regarding neural transmission when the plasma level of resistin increases; this reveals the role of resistin in cardiovascular responses such as metabolic syndrome and hypertension.

Insomnia and anxiety are two common clinical diseases that threaten people’s physical and mental health. Insomnia and anxiety may share some similar underlying neural circuit mechanisms in the brain. In this study, we combine techniques including chemo-fMRI, optogenetics, and chemogenetics to reveal that the glutamatergic neurons of the paraventricular hypothalamic nucleus (PVN) regulate both anxiety and arousal through two different downstream neural circuits. Optogenetic activation of the PVN-cingulate cortex (Cg) neural circuit triggers anxiety-like behaviors in mice without affecting the wakefulness, while optogenetic activation of the PVN-paraventricular thalamic nucleus (PVT) neural circuit promotes wakefulness in mice without affecting anxiety-like behaviors. Our research reveals that PVN is a key brain area for controlling anxiety and arousal behaviors. We also provide a neurological explanation for anxiety disorder and insomnia which may offer guidance for treatments including drugs or transcranial magnetic stimulation for the patients.

Hypersomnolence disorder (HD) is characterized by excessive sleep, which is a common sequela following stroke, infection, or tumorigenesis. HD is traditionally thought to be associated with lesions of wake-promoting nuclei. However, lesions of a single wake-promoting nucleus, or even two simultaneously, did not exert serious HD. Therefore, the specific nucleus and neural circuitry for HD remain unknown. Here, we observed that the paraventricular nucleus of the hypothalamus (PVH) exhibited higher c-fos expression during the active period (23:00) than during the inactive period (11:00) in mice. Therefore, we speculated that the PVH, in which most neurons are glutamatergic, may represent one of the key arousal-controlling centers. By using vesicular glutamate transporter 2 (vglut2Cre) mice together with fiber photometry, multichannel electrophysiological recordings, and genetic approaches, we found that PVHvglut2 neurons were most active during wakefulness. Chemogenetic activation of PVHvglut2 neurons induced wakefulness for 9 hr, and photostimulation of PVHvglut2→parabrachial complex/ventral lateral septum circuits immediately drove transitions from sleep to wakefulness. Moreover, lesioning or chemogenetic inhibition of PVHvglut2 neurons dramatically decreased wakefulness. These results indicate that the PVH is critical for arousal promotion and maintenance.

Background: The renal sympathetic nerve activity (RSNA) is enhanced in renal failure. Paraventricular nucleus in hypothalamus is an important central site to regulate sympathetic activity. There are angiotensin II (Ang) II receptors in this nucleus. The aim of this study was to evaluate the effects of angiotensin II in hypothalamic paraventricular nucleus (PVN) on renal ischemia-reperfusion injury and RSNA. Methods: This study was done at 2013 in Physiology department of Tehran University of Medical Sciences. One week before the induction of renal Ischemia-Reperfusion (IR) in Sprague-Dawley rats, a cannula was inserted into the right PVN for microinjection of different doses of Ang II (3, 30, and 300 ng). Then right nephrectomy was done. After one week recovery, renal IR injury was induced by clamping the left renal artery for 45 minute and then reperfusion for 3 or 24 hour. Ten minutes before the induction of renal ischemia-reperfusion, administration of different doses of angiotensin II were done in different groups. In all animals, left renal sympathetic activity was recorded before and during renal ischemia. After 3 or 24 hours reperfusion the blood, kidney and brain were collected to assay renal function and histology and oxidative stress indices Superoxide Dismutase, SOD and Malondialdehyde, MDA) in PVN. Results: Administration of different pharmacological doses of angiotensin II into PVN exaggerated the renal IR injury. Angiotensin II in different doses increased the plasma creatinine and BUN levels and renal histological markers in comparison to renal IR in-jury (P

Impact of environmental stress upon pathophysiology of the metabolic syndrome (MetS) has been substantiated by epidemiological, psychophysiological, and endocrinological studies. This review discusses recent advances in the understanding of causative roles of nutritional factors, sympathomedullo-adrenal (SMA) and hypothalamic-pituitary adrenocortical (HPA) axes, and adipose tissue chronic low-grade inflammation processes in MetS. Disturbances in the neuroendocrine systems for leptin, melanocortin, and neuropeptide Y (NPY)/agouti-related protein systems have been found resulting directly in MetS-like conditions. The review identifies candidate risk genes from factors shown critical for the functioning of each of these neuroendocrine signalling cascades. In its meta-analytic part, recent studies in epigenetic modification (histone methylation, acetylation, phosphorylation, ubiquitination) and posttranscriptional gene regulation by microRNAs are evaluated. Several studies suggest modification mechanisms of early life stress (ELS) and diet-induced obesity (DIO) programming in the hypothalamic regions with populations of POMC-expressing neurons. Epigenetic modifications were found in cortisol (here HSD11B1 expression), melanocortin, leptin, NPY, and adiponectin genes. With respect to adiposity genes, epigenetic modifications were documented for fat mass gene cluster APOA1/C3/A4/A5, and the lipolysis gene LIPE. With regard to inflammatory, immune and subcellular metabolism, PPARG, NKBF1, TNFA, TCF7C2, and those genes expressing cytochrome P450 family enzymes involved in steroidogenesis and in hepatic lipoproteins were documented for epigenetic modifications.

This study used immunohistochemical methods to investigate the possibility that hypothalamic neurons that contain 11-β-hydroxysteroid dehydrogenase type 2 (HSD2) are involved in the control of feeding by rats via neuroanatomical associations with the α subtype of estrogen receptor (ERα), catecholamines, and/or oxytocin. An aggregate of HSD2-containing neurons is located laterally in the hypothalamus, and the numbers of these neurons were greatly increased by estradiol treatment in ovariectomized rats compared to numbers in male rats and in ovariectomized rats that were not given estradiol. However, HSD2-containing neurons were anatomically segregated from ERα-containing neurons in the Ventromedial Hypothalamus and the Arcuate Nucleus. There was an absence of oxytocin-immunolabeled fibers in the area of HSD2-labeled neurons. Taken together, these findings provide no support for direct associations between hypothalamic HSD2 and ERα or oxytocin neurons in the control of feeding. In contrast, there was catecholamine-fiber labeling in the area of HSD2-labeled neurons, and these fibers occasionally were in close apposition to HSD2-labeled neurons. Therefore, we cannot rule out interactions between HSD2 and catecholamines in the control of feeding; however, given the relative sparseness of the appositions, any such interaction would appear to be modest. Thus, these studies do not conclusively identify a neuroanatomical substrate by which HSD2-containing neurons in the hypothalamus may alter feeding, and leave the functional role of hypothalamic HSD2-containing neurons subject to further investigation.

Chronic social stress has been associated with increased caloric intake and adiposity. These effects have been linked to stress induced changes in the secretion of ghrelin, a hormone that targets a number of brain regions to increase food intake and energy expenditure and promote increased body fat content. One of the brain sites targeted by ghrelin is the hypothalamic paraventricular nucleus (PVN), a region critical for both the regulation of the stress response and the regulation of energy balance. Given these data, we examined the contribution of ghrelin receptors in the PVN to the metabolic and behavioral changes that are seen during chronic social stress in mice. To do this, mice were implanted with cannulae attached to osmotic minipumps and delivering either vehicle or the ghrelin receptor (growth hormone secretagogue receptor) antagonist [D-Lys-3]-GHRP-6 (20nmol/day/mouse). Following a week of recovery, half of the animals in each group were exposed to chronic social defeat stress for a period of three weeks whereas the other half were left undisturbed. During this time, all animals were given ad libitum access to standard laboratory chow and presented a high-fat diet for 4 hours during the day. Results showed that the ghrelin receptor antagonism did not decrease stressed induced caloric intake, but paradoxically increased the intake of the high fat diet. This would suggest that ghrelin acts on the PVN to promote the intake of carbohydrate rich diets while decreasing fat intake and blockade of ghrelin receptors in the PVN leads to more consumption of foods that are high in fat.