FGF21 attenuates salt-sensitive hypertension via regulating HNF4α/ACE2 axis in the hypothalamic paraventricular nucleus of mice.

BACKGROUND: Fibroblast growth factor 21 (FGF21) has a protective effect against cardiovascular disease. However, the role of FGF21 in hypertension remains elusive. METHODS: Ten-week-old male C57BL/6 mice were randomly divided into normal-salt (NS) group, NS+FGF21 group, deoxycorticosterone acetate-salt (DOCA) group and DOCA+FGF21 group. The mice in NS group underwent uninephrectomy without receiving DOCA and 1% NaCl and the mice in DOCA group were subjected to uninephrectomy and DOCA-salt (DOCA and 1% NaCl) treatment for 6 weeks. At the same time, the mice were infused with vehicle (artificial cerebrospinal fluid, aCSF) or FGF21 (1 mg/kg) into the bilateral paraventricular nucleus (PVN) of mice. RESULTS: Here, we showed that FGF21 treatment lowered DOCA salt-induced inflammation and oxidative stress in the PVN, which reduced sympathetic nerve activity and hypertension. Mechanistically, FGF21 treatment decreased the expression of HNF4α and inhibited the binding activity of HNF4α to the promoter region of ACE2 in the PVN of DOCA salt-treated mice, which further up-regulated ACE2/Ang (1-7) signals in the PVN. In addition, ACE2 deficiency abolished the protective effect of FGF21 in DOCA salt-treated mice, suggesting that FGF21-mediated antihypertensive effect was dependent on ACE2. CONCLUSIONS: The results demonstrate that FGF21 protects against salt-sensitive hypertension via regulating HNF4α/ACE2/Ang (1-7) axis in the PVN of DOCA salt-treated mice via multi-organ crosstalk between liver, brain and blood vessels.

The impact of pubertal stress and adult hormone exposure on the transcriptome of the developing hypothalamus.

Why individuals suffer negative consequences following stress is a complex phenomenon that is dictated by individual factors, the timing of stress within the lifespan, and when in the lifespan the consequences are measured. Women who undergo adverse childhood experiences are at risk for lasting biological consequences, including affective and stress dysregulation. We have shown that pubertal adversity is associated with a blunted hypothalamic-pituitary-adrenal axis glucocorticoid response in peripartum humans and mice. In mice, our prior examination of the paraventricular nucleus (PVN) of the hypothalamus showed that pubertal stress led to an upregulation of baseline mRNA expression of six immediate early genes (IEGs) in the PVN of adult, pregnant mice. Separately, we showed that the pregnancy-associated hormone allopregnanolone is necessary and sufficient to produce the blunted stress response phenotype in pubertally stressed mice. In the current study, we further examined a potential mechanistic role for the IEGs in the PVN. We found that in pubertally stressed adult female, but not male, mice, intra-PVN allopregnanolone was sufficient to recapitulate the baseline IEG mRNA expression profile previously observed in pubertally stressed, pregnant mice. We also examined baseline IEG mRNA expression during adolescence, where we found that IEGs have developmental trajectories that showed sex-specific disruption by pubertal stress. Altogether, these data establish that IEGs may act as a key molecular switch involved in increased vulnerability to negative outcomes in adult, pubertally stressed animals. How the factors that produce vulnerability combine throughout the lifespan is key to our understanding of the etiology of stress-related disorders.

Hypothalamic vasopressin sex differentiation is observed by embryonic day 15 in mice and is disrupted by the xenoestrogen bisphenol A.

Arginine vasopressin (AVP) neurons of the hypothalamic paraventricular region (AVPPVN) mediate sex-biased social behaviors across most species, including mammals. In mice, neural sex differences are thought to be established during a critical window around birth ( embryonic (E) day 18 to postnatal (P) day 2) whereby circulating testosterone from the fetal testis is converted to estrogen in sex-dimorphic brain regions. Here, we found that AVPPVN neurons are sexually dimorphic by E15.5, prior to this critical window, and that gestational bisphenol A (BPA) exposure permanently masculinized female AVPPVN neuronal numbers, projections, and electrophysiological properties, causing them to display male-like phenotypes into adulthood. Moreover, we showed that nearly twice as many neurons that became AVP+ by P0 were born at E11 in males and BPA-exposed females compared to control females, suggesting that AVPPVN neuronal masculinization occurs between E11 and P0. We further narrowed this sensitive period to around the timing of neurogenesis by demonstrating that exogenous estrogen exposure from E14.5 to E15.5 masculinized female AVPPVN neuronal numbers, whereas a pan-estrogen receptor antagonist exposed from E13.5 to E15.5 blocked masculinization of males. Finally, we showed that restricting BPA exposure to E7.5-E15.5 caused adult females to display increased social dominance over control females, consistent with an acquisition of male-like behaviors. Our study reveals an E11.5 to E15.5 window of estrogen sensitivity impacting AVPPVN sex differentiation, which is impacted by prenatal BPA exposure.

Intestinal perfusion of unacylated ghrelin alleviated metabolically associated fatty liver disease in rats via a central glucagon-like peptide-1 pathway.

Unacylated ghrelin (UAG), the unacylated form of ghrelin, accounts for 80%-90% of its circulation. Accumulated studies have pointed out that UAG may be used to treat metabolic disorders. This study aimed to investigate the effect of intestinal perfusion of UAG on metabolically associated fatty liver disease (MAFLD) induced by a high-fat diet and its possible mechanisms. Neuronal retrograde tracking combined with immunofluorescence, central administration of a glucagon-like peptide-1 receptor (GLP-1R) antagonist, and hepatic vagotomy was performed to reveal its possible mechanism involving a central glucagon-like peptide-1 (GLP-1) pathway. The results showed that intestinal perfusion of UAG significantly reduced serum lipids, aminotransferases, and food intake in MAFLD rats. Steatosis and lipid accumulation in the liver were significantly alleviated, and lipid metabolism-related enzymes in the liver were regulated. UAG upregulated the expression of GLP-1 receptor (GLP-1R) in the paraventricular nucleus (PVN) and GLP-1 in the nucleus tractus solitarii (NTS), as well as activated GLP-1 neurons in the NTS. Furthermore, GLP-1 fibers projected from NTS to PVN were activated by the intestinal perfusion of UAG. However, hepatic vagotomy and GLP-1R antagonists delivered into PVN before intestinal perfusion of UAG partially attenuated its alleviation of MAFLD. In conclusion, intestinal perfusion of UAG showed a therapeutic effect on MAFLD, which might be related to its activation of the GLP-1 neuronal pathway from NTS to PVN. The present results provide a new strategy for the treatment of MAFLD.NEW & NOTEWORTHY Intestinal perfusion of UAG, the unacylated form of ghrelin, has shown promising potential for treating MAFLD. This study unveils a potential mechanism involving the central GLP-1 pathway, with UAG upregulating GLP-1R expression and activating GLP-1 neurons in specific brain regions. These findings propose a novel therapeutic strategy for MAFLD treatment through UAG and its modulation of the GLP-1 neuronal pathway.

GLP-1 in the Hypothalamic Paraventricular Nucleus Promotes Sympathetic Activation and Hypertension.

Glucagon-like peptide-1 (GLP-1) and its analogs are widely used for diabetes treatment. The paraventricular nucleus (PVN) is crucial for regulating cardiovascular activity. This study aims to determine the roles of GLP-1 and its receptors (GLP-1R) in the PVN in regulating sympathetic outflow and blood pressure. Experiments were carried out in male normotensive rats and spontaneously hypertensive rats (SHR). Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded. GLP-1 and GLP-1R expressions were present in the PVN. PVN microinjection of GLP-1R agonist recombinant human GLP-1 (rhGLP-1) or EX-4 increased RSNA and MAP, which were prevented by GLP-1R antagonist exendin 9-39 (EX9-39) or GLP-1R antagonist 1, superoxide scavenger tempol, antioxidant N-acetylcysteine, NADPH oxidase (NOX) inhibitor apocynin, adenylyl cyclase (AC) inhibitor SQ22536 or protein kinase A (PKA) inhibitor H89. PVN microinjection of rhGLP-1 increased superoxide production, NADPH oxidase activity, cAMP level, AC, and PKA activity, which were prevented by SQ22536 or H89. GLP-1 and GLP-1R were upregulated in the PVN of SHR. PVN microinjection of GLP-1 agonist increased RSNA and MAP in both WKY and SHR, but GLP-1 antagonists caused greater effects in reducing RSNA and MAP in SHR than in WKY. The increased superoxide production and NADPH oxidase activity in the PVN of SHR were augmented by GLP-1R agonists but attenuated by GLP-1R antagonists. These results indicate that activation of GLP-1R in the PVN increased sympathetic outflow and blood pressure via cAMP-PKA-mediated NADPH oxidase activation and subsequent superoxide production. GLP-1 and GLP-1R upregulation in the PVN partially contributes to sympathetic overactivity and hypertension.

Trimethylamine-N-oxide aggravated the sympathetic excitation in D-galactose induced aging rats by down-regulating P2Y12 receptor in microglia.

This study aimed to determine whether trimethylamine N-oxide (TMAO) was involved in sympathetic activation in aging and the underlying mechanisms. Our hypothesis is TMAO reduces P2Y12 receptor (P2Y12R) and induces microglia-mediated inflammation in the paraventricular nucleus (PVN), then leading to sympathetic activation in aging. This study involved 18 young adults and 16 old adults. Aging rats were established by injecting D-galactose (D-gal, 200 mg/kg/d) subcutaneously for 12 weeks. TMAO (120 mg/kg/d) or 1% 3, 3-dimethyl-l-butanol (DMB) was administrated via drinking water for 12 weeks to investigate their effects on neuroinflammation and sympathetic activation in aging rats. Plasma TMAO, NE and IL-1ß levels were higher in old adults than in young adults. In addition, standard deviation of all normal to normal intervals (SDNN) and standard deviation of the average of normal to normal intervals (SDANN) were lower in old adults and negatively correlated with TMAO, indicating sympathetic activation in old adults, which is associated with an increase in TMAO levels. Treatment of rats with D-gal showed increased senescence-associated protein levels and microglia-mediated inflammation, as well as decreased P2Y12R protein levels in PVN. Plasma TMAO, NE and IL-1ß levels were increased, accompanied by enhanced renal sympathetic nerve activity (RSNA). While TMAO treatment exacerbated the above phenomenon, DMB mitigated it. These findings suggest that TMAO contributes to sympathetic hyperactivity in aging by downregulating P2Y12R in microglia and increasing inflammation in the PVN. These results may provide promising new target for the prevention and treatment of aging and aging-related diseases.

Calcineurin regulates synaptic Ca2+-permeable AMPA receptors in hypothalamic presympathetic neurons via α2δ-1-mediated GluA1/GluA2 assembly.

Hypertension is a major adverse effect of calcineurin inhibitors, such as tacrolimus (FK506) and cyclosporine, used clinically as immunosuppressants. Calcineurin inhibitor-induced hypertension (CIH) is linked to augmented sympathetic output from the hypothalamic paraventricular nucleus (PVN). GluA2-lacking, Ca2+-permeable AMPA receptors (CP-AMPARs) are a key feature of glutamatergic synaptic plasticity, yet their role in CIH remains elusive. Here, we found that systemic administration of FK506 in rats significantly increased serine phosphorylation of GluA1 and GluA2 in PVN synaptosomes. Strikingly, FK506 treatment reduced GluA1/GluA2 heteromers in both synaptosomes and endoplasmic reticulum-enriched fractions from the PVN. Blocking CP-AMPARs with IEM-1460 induced a larger reduction of AMPAR-mediated excitatory postsynaptic current (AMPAR-EPSC) amplitudes in retrogradely labelled, spinally projecting PVN neurons in FK506-treated rats than in vehicle-treated rats. Furthermore, FK506 treatment shifted the current-voltage relationship of AMPAR-EPSCs from linear to inward rectification in labelled PVN neurons. FK506 treatment profoundly enhanced physical interactions of α2δ-1 with GluA1 and GluA2 in the PVN. Inhibiting α2δ-1 with gabapentin, α2δ-1 genetic knockout, or disrupting α2δ-1-AMPAR interactions with an α2δ-1 C terminus peptide restored GluA1/GluA2 heteromers in the PVN and diminished inward rectification of AMPAR-EPSCs in labelled PVN neurons induced by FK506 treatment. Additionally, microinjection of IEM-1460 or α2δ-1 C terminus peptide into the PVN reduced renal sympathetic nerve discharges and arterial blood pressure elevated in FK506-treated rats but not in vehicle-treated rats. Thus, calcineurin in the hypothalamus constitutively regulates AMPAR subunit composition and phenotypes by controlling GluA1/GluA2 interactions with α2δ-1. Synaptic CP-AMPARs in PVN presympathetic neurons contribute to augmented sympathetic outflow in CIH. KEY POINTS: Systemic treatment with the calcineurin inhibitor increases serine phosphorylation of synaptic GluA1 and GluA2 in the PVN. Calcineurin inhibition enhances the prevalence of postsynaptic Ca2+-permeable AMPARs in PVN presympathetic neurons. Calcineurin inhibition potentiates α2δ-1 interactions with GluA1 and GluA2, disrupting intracellular assembly of GluA1/GluA2 heterotetramers in the PVN. Blocking Ca2+-permeable AMPARs or α2δ-1-AMPAR interactions in the PVN attenuates sympathetic outflow augmented by the calcineurin inhibitor.

Taurocholic acid ameliorates hypertension through the activation of TGR5 in the hypothalamic paraventricular nucleus.

Diets rich in taurine can increase the production of taurine-conjugated bile acids, which are known to exert antihypertensive effects. Despite their benefits to the heart, kidney and arteries, their role in the central nervous system during the antihypertensive process remains unclear. Since hypothalamic paraventricular nucleus (PVN) plays a key role in blood pressure regulation, we aimed to investigate the function of bile acids in the PVN. The concentration of bile acids in the PVN of spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto rats (WKY) fed with normal chow was measured using LC-MS/MS, which identified taurocholic acid (TCA) as the most down-regulated bile acid. To fully understand the mechanism of TCA's functions in the PVN, bi-lateral PVN micro-infusion of TCA was carried out. TCA treatment in the PVN led to a significant reduction in the blood pressure of SHRs, with decreased plasma levels of norepinephrine and improved morphology of cardiomyocytes. It also decreased the number of c-fos+ neurons, reduced the inflammatory response, and suppressed oxidative stress in the PVN of the SHRs. Most importantly, the TGR5 receptors in neurons and microglia were activated. PVN infusion of SBI-115, a TGR5 specific antagonist, was able to counteract with TCA in the blood pressure regulation of SHRs. In conclusion, TCA supplementation in the PVN of SHRs can activate TGR5 in neurons and microglia, reduce the inflammatory response and oxidative stress, suppress activated neurons, and attenuate hypertension.

Decreased levels of hydrogen sulfide in the hypothalamic paraventricular nucleus contribute to sympathetic hyperactivity induced by cerebral infarction.

Paroxysmal sympathetic hyperactivity (PSH) is a common clinical feature secondary to ischemic stroke (IS), but its mechanism is poorly understood. We aimed to investigate the role of H2S in the pathogenesis of PSH. IS patients were divided into malignant (MCI) and non-malignant cerebral infarction (NMCI) group. IS in rats was induced by the right middle cerebral artery occlusion (MCAO). H2S donor (NaHS) or inhibitor (aminooxy-acetic acid, AOAA) were microinjected into the hypothalamic paraventricular nucleus (PVN). Compared with the NMCI group, patients in the MCI group showed PSH, including tachycardia, hypertension, and more plasma norepinephrine (NE) that was positively correlated with levels of creatine kinase, glutamate transaminase, and creatinine respectively. The 1-year survival rate of patients with high plasma NE levels was lower. The hypothalamus of rats with MCAO showed increased activity, especially in the PVN region. The levels of H2S in PVN of the rats with MCAO were reduced, while the blood pressure and renal sympathetic discharge were increased, which could be ameliorated by NaHS and exacerbated by AOAA. NaHS completely reduced the disulfide bond of NMDAR1 in PC12 cells. The inhibition of NMDAR by MK-801 microinjected in PVN of rats with MCAO also could lower blood pressure and renal sympathetic discharge. In conclusion, PSH may be associated with disease progression and survival in patients with IS. Decreased levels of H2S in PVN were involved in regulating sympathetic efferent activity after cerebral infarction. Our results might provide a new strategy and target for the prevention and treatment of PSH.

Dexmedetomidine Inhibits Paraventricular Corticotropin-releasing Hormone Neurons that Attenuate Acute Stress-induced Anxiety-like Behavior in Mice.

BACKGROUND: Dexmedetomidine has repeatedly shown to improve anxiety, but the precise neural mechanisms underlying this effect remain incompletely understood. This study aims to explore the role of corticotropin-releasing hormone-producing hypothalamic paraventricular nucleus (CRHPVN) neurons in mediating the anxiolytic effects of dexmedetomidine. METHODS: A social defeat stress mouse model was used to evaluate the anxiolytic effects induced by dexmedetomidine through the elevated plus maze, open-field test, and measurement of serum stress hormone levels. In vivo Ca2+ signal fiber photometry and ex vivo patch-clamp recordings were used to determine the excitability of CRHPVN neurons and investigate the specific mechanism involved. CRHPVN neuron modulation was achieved through chemogenetic activation or inhibition. RESULTS: Compared with saline, dexmedetomidine (40 µg/kg) alleviated anxiety-like behaviors. Additionally, dexmedetomidine reduced CRHPVN neuronal excitability. Chemogenetic activation of CRHPVN neurons decreased the time spent in the open arms of the elevated plus maze and in the central area of the open-field test. Conversely, chemogenetic inhibition of CRHPVN neurons had the opposite effect. Moreover, the suppressive impact of dexmedetomidine on CRHPVN neurons was attenuated by the α2-receptor antagonist yohimbine. CONCLUSIONS: The results indicate that the anxiety-like effects of dexmedetomidine are mediated via α2-adrenergic receptor-triggered inhibition of CRHPVN neuronal excitability in the hypothalamus.

Central administration of AICAR attenuates hypertension via AMPK/Nrf2 pathway in the hypothalamic paraventricular nucleus of hypertensive rats.

BACKGROUND: Oxidative stress and inflammatory cytokines in the hypothalamus paraventricular nucleus (PVN) have been implicated in sympathetic nerve activity and the development of hypertension, but the specific mechanisms underlying their production in the PVN remains to be elucidated. Previous studies have demonstrated that activation of nuclear transcription related factor-2 (Nrf2) in the PVN reduced the production of reactive oxygen species (ROS) and inflammatory mediators. Moreover, AMP-activated protein kinase (AMPK), has been observed to decrease ROS and inflammatory cytokine production when activated in the periphery. 5-amino-1-ß-D-ribofuranosyl-imidazole-4-carboxamide (AICAR) is an AMPK agonist. However, little research has been conducted on the role of AMPK in the PVN during hypertension. Therefore, we hypothesized that AICAR in the PVN is involved in regulating AMPK/Nrf2 pathway, affecting ROS and inflammatory cytokine expression, influencing sympathetic nerve activity. METHODS: Adult male Sprague-Dawley rats were utilized to induce two-kidney, one-clip (2K1C) hypertension via constriction of the right renal artery. Bilateral PVN was microinjected with either artificial cerebrospinal fluid or AICAR once a day for 4 weeks. RESULTS: Compared to the SHAM group, the PVN of 2K1C hypertensive rats decreased p-AMPK and p-Nrf2 expression, increased Fra-Like, NAD(P)H oxidase (NOX)2, NOX4, tumor necrosis factor-α and interleukin (IL)-1ß expression, elevated ROS levels, decreased superoxide dismutase 1 and IL-10 expression, and elevated plasma norepinephrine levels. Bilateral PVN microinjection of AICAR significantly ameliorated these changes. CONCLUSION: These findings suggest that repeated injection of AICAR in the PVN suppresses ROS and inflammatory cytokine production through the AMPK/Nrf2 pathway, reducing sympathetic nerve activity and improving hypertension.

Beneficial effects of metformin supplementation in hypothalamic paraventricular nucleus and arcuate nucleus of type 2 diabetic rats.

Background Oxidative stress and inflammation play important roles in the development of diabetes. Metformin (MET) is considered as the first-line therapy for patients with type 2 diabetes (T2D). Hypothalamic paraventricular nucleus (PVN) and hypothalamic arcuate nucleus (ARC) are vital in obesity and diabetes. However, there have been few studies on the effects of MET on inflammatory reaction and oxidative stress in the PVN and ARC of T2D diabetic rats. Methods Male Sprague-Dawley (SD) rats were fed with high-fat diet (HFD), and intraperitoneally injected with low-dose streptozotocin (STZ, 30 mg/kg) at 6th week to induce T2D diabetes. After injection of STZ, they were fed with HFD continually. Starting from the 8th week of HFD feeding, T2D rats received intragastrical administration of MET (150 mg/kg/day) in addition to the HFD for another 8 weeks. At the end of the 15th week, the rats were anaesthetized to record the sympathetic nerve activity and collect blood and tissue samples. Results In comparison with control rats, T2D diabetic rats had higher levels of pro-inflammatory cytokines (PICs) and excessive oxidative stress in the PVN and ARC, accompanied with more activated astrocytes. The renal sympathetic nerve activity (RSNA) and the plasma norepinephrine (NE) increased in T2D diabetic rats. The expression of tyrosine hydroxylase (TH) increased and the expression of 67-kDa isoform of glutamate decarboxylase (GAD67) decreased in T2D diabetic rats. Supplementation of MET decreased blood glucose, suppressed RSNA, decreased PICs (TNF-α, IL-1ß and IL-6) in PVN and ARC, attenuated oxidative stress and activation of astrocytes in ARC and PVN of T2D diabetic rats, as well as restored the balance of neurotransmitter synthetase. The number of Fra-LI (chronic neuronal excitation marker) positive neurons in the ARC and PVN of T2D diabetic rats increased. Chronic supplementation of MET also decreased the number of Fra-LI positive neurons in the ARC and PVN of T2D diabetic rats. Conclusion These findings suggest that the PVN and ARC participate in the beneficial effects of MET in T2D diabetic rats, which is possibly mediated via down-regulating of inflammatory molecules, attenuating oxidative stress and restoring the balance of neurotransmitter synthetase by MET in the PVN and ARC.

Haloperidol and aripiprazole affects CRH system and behaviour of animals exposed to chronic mild stress.

CRH system integrates responses to stress challenges, whereas antipsychotics may impinge on this process. Effect of haloperidol (HAL) and aripiprazole (ARI) on chronic mild stress (CMS) induced neurobehavioral and CRH/CRHR1 system changes was studied in functionally interconnected rat brain areas including prefrontal cortex (PFC), bed nucleus of the stria terminalis (BNST), hypothalamic paraventricular nucleus (PVN), hippocampus (HIP), and amygdala (AMY). Animals were exposed to CMS for 3-weeks and since the 7th day of CMS injected with vehicle (VEH), HAL (1 mg/kg) or ARI (10 mg/kg) for 4-weeks. Expression levels of CRH, CRHR1, and c-fos genes and anxiety-like and anhedonia behavioural patterns were evaluated. CMS in VEH animals suppressed CRH gene expression in the PFC and BNST, c-fos expression in all areas, except HIP, and increased CRHR1 gene expression in the HIP. Antipsychotics decreased CRH gene expression in all areas, except HIP and by CMS elevated CRHR1 expression in the HIP (ARI also in AMY). CMS and antipsychotics decreased the sucrose preference. Aripiprazole prevented CRH expression decrease in the BNST and sucrose preference induced by CMS. Haloperidol increased time spent in the EPM open arms. These data indicate that HAL and ARI selectively influenced behavioural parameters and CRH/CRHR1 gene expression levels in CMS animals.

Repeated Administration of Orexin into the Thalamic Paraventricular Nucleus Inhibits the Development of Morphine-Induced Analgesia.

BACKGROUND: Hypothalamic neuropeptides, orexins, play pivotal roles in nociception and pain modulation. OBJECTIVE: In this study, we investigated the effect of the administration of orexin into the paraventricular nucleus (PVT) on the development of morphine-induced analgesia in rats. METHOD: Male Wistar rats weighing 250-300g received subcutaneous (s.c.) chronic morphine (6, 16, 26, 36, 46, 56 and 66 mg/kg, 2 ml/kg) at an interval of 24 hours for 7 days. Animals were divided into two experimental groups in which the orexin (100 µM, 200 nl) and its vehicle were microinjected into the PVT nucleus for 7 days before each morphine injection. Then, the formalin test was performed for the assessment of pain-related behaviors. RESULTS: The results demonstrated that the rats pretreated by intra-PVT orexin exhibited higher pain-related behaviors than the morphine-treated group. The analgesic effects of morphine were significantly lower in orexin plus morphine-treated rats than the vehicle plus morphine-treated ones. CONCLUSION: Our findings suggested that the animals receiving the prolonged intra-PVT application of orexin before morphine injection demonstrated a significant increase in the development of nociceptive behaviors in all phases. There fore, the present study highlighted a new area of the brain involved in the effect of orexin on analgesia induced by morphine.

Effects of chronic exposure to bisphenol A in adult female mice on social behavior, vasopressin system, and estrogen membrane receptor (GPER1).

Bisphenol A (BPA), an organic synthetic compound found in some plastics and epoxy resins, is classified as an endocrine disrupting chemical. Exposure to BPA is especially dangerous if it occurs during specific "critical periods" of life, when organisms are more sensitive to hormonal changes (i.e., intrauterine, perinatal, juvenile or puberty periods). In this study, we focused on the effects of chronic exposure to BPA in adult female mice starting during pregnancy. Three months old C57BL/6J females were orally exposed to BPA or to vehicle (corn oil). The treatment (4 µg/kg body weight/day) started the day 0 of pregnancy and continued throughout pregnancy, lactation, and lasted for a total of 20 weeks. BPA-treated dams did not show differences in body weight or food intake, but they showed an altered estrous cycle compared to the controls. In order to evidence alterations in social and sociosexual behaviors, we performed the Three-Chamber test for sociability, and analyzed two hypothalamic circuits (well-known targets of endocrine disruption) particularly involved in the control of social behavior: the vasopressin and the oxytocin systems. The test revealed some alterations in the displaying of social behavior: BPA-treated dams have higher locomotor activity compared to the control dams, probably a signal of high level of anxiety. In addition, BPA-treated dams spent more time interacting with no-tester females than with no-tester males. In brain sections, we observed a decrease of vasopressin immunoreactivity (only in the paraventricular and suprachiasmatic nuclei) of BPA-treated females, while we did not find any alteration of the oxytocin system. In parallel, we have also observed, in the same hypothalamic nuclei, a significant reduction of the membrane estrogen receptor GPER1 expression.

Conditional ablation of vasopressin-synthesizing neurons in transgenic rats.

Vasopressin-synthesizing neurons are located in several brain regions, including the hypothalamic paraventricular nucleus (PVN), supraoptic nucleus (SON) and suprachiasmatic nucleus (SCN). Vasopressin has been shown to have various functions in the brain, including social recognition memory, stress responses, emotional behaviors and circadian rhythms. The precise physiological functions of vasopressin-synthesizing neurons in specific brain regions remain to be clarified. Conditional ablation of local vasopressin-synthesizing neurons may be a useful tool for investigation of the functions of vasopressin neurons in the regions. In the present study, we characterized a transgenic rat line that expresses a mutated human diphtheria toxin receptor under control of the vasopressin gene promoter. Under a condition of salt loading, which activates the vasopressin gene in the hypothalamic PVN and SON, transgenic rats were i.c.v. injected with diphtheria toxin. Intracerebroventricular administration of diphtheria toxin after salt loading depleted vasopressin-immunoreactive cells in the hypothalamic PVN and SON, but not in the SCN. The number of oxytocin-immunoreactive cells in the hypothalamus was not significantly changed. The rats that received i.c.v. diphtheria toxin after salt loading showed polydipsia and polyuria, which were rescued by peripheral administration of 1-deamino-8-d-arginine vasopressin via an osmotic mini-pump. Intrahypothalamic administration of diphtheria toxin in transgenic rats under a normal hydration condition reduced the number of vasopressin-immunoreactive neurons, but not the number of oxytocin-immunoreactive neurons. The transgenic rat model can be used for selective ablation of vasopressin-synthesizing neurons and may be useful for clarifying roles of vasopressin neurons at least in the hypothalamic PVN and SON in the rat.

Acetylcholinesterase inhibition with Pyridostigmine attenuates hypertension and neuroinflammation in the paraventricular nucleus in rat model for Preeclampsia.

Preeclampsia (PE) is characterized by hypertension, autonomic imbalance and inflammation. The subfornical organ (SFO) reportedly relays peripheral inflammatory mediator's signals to the paraventricular nucleus (PVN), a brain autonomic center shown to mediate hypertension in hypertensive rat but not yet in PE rat models. Additionally, we previously showed that Pyridostigmine (PYR), an acetylcholinesterase inhibitor, attenuated placental inflammation and hypertension in PE models. In this study, we investigated the effect of PYR on the activities of these brain regions in PE model. PYR (20 mg/kg/day) was administered to reduced uterine perfusion pressure (RUPP) Sprague-Dawley rat from gestational day (GD) 14 to GD19. On GD19, the mean arterial pressure (MAP) was recorded and samples were collected for analysis. RUPP rats exhibited increased MAP (P = 0.0025), elevated circulating tumor necrosis factor-α (TNF-α, P = 0.0075), reduced baroreflex sensitivity (BRS), increased neuroinflammatory markers including TNF-α, interleukin-1ß (IL-1ß), microglial activation (P = 0.0039), oxidative stress and neuronal excitation within the PVN and the SFO. Changes in MAP, in molecular and cellular expression induced by RUPP intervention were improved by PYR. The ability of PYR to attenuate TNF-α mediated central effect was evaluated in TNF-α-infused pregnant rats. TNF-α infusion-promoted neuroinflammation in the PVN and SFO in dams was abolished by PYR. Collectively, our data suggest that PYR improves PE-like symptoms in rat by dampening placental ischemia and TNF-α-promoted inflammation and pro-hypertensive activity in the PVN. This broadens the therapeutical potential of PYR in PE.

Circadian neurons in the paraventricular nucleus entrain and sustain daily rhythms in glucocorticoids.

Signals from the central circadian pacemaker, the suprachiasmatic nucleus (SCN), must be decoded to generate daily rhythms in hormone release. Here, we hypothesized that the SCN entrains rhythms in the paraventricular nucleus (PVN) to time the daily release of corticosterone. In vivo recording revealed a critical circuit from SCN vasoactive intestinal peptide (SCNVIP)-producing neurons to PVN corticotropin-releasing hormone (PVNCRH)-producing neurons. PVNCRH neurons peak in clock gene expression around midday and in calcium activity about three hours later. Loss of the clock gene Bmal1 in CRH neurons results in arrhythmic PVNCRH calcium activity and dramatically reduces the amplitude and precision of daily corticosterone release. SCNVIP activation reduces (and inactivation increases) corticosterone release and PVNCRH calcium activity, and daily SCNVIP activation entrains PVN clock gene rhythms by inhibiting PVNCRH neurons. We conclude that daily corticosterone release depends on coordinated clock gene and neuronal activity rhythms in both SCNVIP and PVNCRH neurons.

Astaxanthin Ameliorates Blood Pressure in Salt-Induced Prehypertensive Rats Through ROS/MAPK/NF-κB Pathways in the Hypothalamic Paraventricular Nucleus.

Astaxanthin (AST) has a variety of biochemical effects, including anti-inflammatory, antioxidative, and antihypertensive functions. The aim of the present study was to determine whether AST ameliorates blood pressure in salt-induced prehypertensive rats by ROS/MAPK/NF-κB pathways in hypothalamic paraventricular nucleus.To explore the central effects of AST on the development of blood pressure, prehypertensive rats were induced by a high-salt diet (HS, 8% NaCl) and its control groups were treated with normal-salt diet (NS, 0.3% NaCl). The Dahl salt-sensitive (S) rats with HS diet for 6 weeks received AST or vehicle by gastric perfusion for 6 weeks. Compared to those with NS diet, rats with HS diet exhibited increased mean arterial pressure (MAP) and heart rate (HR). These increases were associated with higher plasma level of norepinephrine (NE), interleukin 1ß (IL-1ß), and interleukin 6 (IL-6); elevated PVN level of reactive oxygen species (ROS), NOX2, and NOX4, that of IL-1ß, IL-6, monocyte chemotactic protein 1 (MCP-1), tyrosine hydroxylase (TH), phosphorylation extracellular-signal-regulated kinase (p-ERK1/2), phosphorylation Jun N-terminal kinases (p-JNK), nuclear factor-kappa B (NF-κB) activity; and lower levels of IL-10, superoxide dismutase (SOD), and catalase (CAT) in the PVN. In addition, our data demonstrated that chronic AST treatment ameliorated these changes in the HS but not NS diet rats. These data suggested that AST could alleviate prehypertensive response in HS-induced prehypertension through ROS/MAPK/NF-κB pathways in the PVN.

Curcumin ameliorates hypertension via gut-brain communication in spontaneously hypertensive rat.

Gut dysbiosis and dysregulation of gut-brain communication have been identified in hypertensive patients and animal models. Previous studies have shown that probiotic or prebiotic treatments exert positive effects on the pathophysiology of hypertension. This study aimed to examine the hypothesis that the microbiota-gut-brain axis is involved in the antihypertensive effects of curcumin, a potential prebiotic obtained from Curcuma longa. Male 8- to 10-week-old spontaneously hypertensive rats (SHRs) and Wistar Kyoto (WKY) rats were divided into four groups: WKY rats and SHRs treated with vehicle and SHRs treated with curcumin in dosage of 100 or 300 mg/kg/day for 12 weeks. Our results show that the elevated blood pressure of SHRs was markedly decreased in both curcumin-treated groups. Curcumin treatment also altered the gut microbial composition and improved intestinal pathology and integrity. These factors were associated with reduced neuroinflammation and oxidative stress in the hypothalamus paraventricular nucleus (PVN). Moreover, curcumin treatment increased butyrate levels in the plasma, which may be the result of increased butyrate-producing gut microorganisms. In addition, curcumin treatment also activated G protein-coupled receptor 43 (GPR 43) in the PVN. These results indicate that curcumin reshapes the composition of the gut microbiota and ameliorates the dysregulation of the gut-brain communication to induce antihypertensive effects.