Paraventricular Nucleus Neuronal Activity Research Articles

MGluR5 Upregulation Increases Excitability of Hypothalamic Presympathetic Neurons through NMDA Receptor Trafficking in Spontaneously Hypertensive Rats

The hypothalamic paraventricular nucleus (PVN) is critically involved in elevated sympathetic output and the development of hypertension. However, changes in group I metabotropic glutamate receptors (mGluR1 and mGluR5) and their relevance to the hyperactivity of PVN presympathetic neurons in hypertension remain unclear. Here, we found that selectively blocking mGluR5 significantly reduced the basal firing activity of spinally projecting PVN neurons in spontaneously hypertensive rats (SHRs), but not in normotensive Wistar-Kyoto (WKY) rats. However, blocking mGluR1 had no effect on the firing activity of PVN neurons in either group. The mRNA and protein levels of mGluR5 in the PVN and rostral ventrolateral medulla were significantly higher in SHRs than in WKY rats. The group I mGluR selective agonist (S)-3,5-dihydroxyphenylglycine (DHPG) similarly increased the firing activity of PVN neurons in WKY rats and SHRs. In addition, blocking NMDA receptors (NMDARs) through bath application or intracellular dialysis not only decreased the basal firing in SHRs, but also eliminated DHPG-induced excitation of spinally projecting PVN neurons. DHPG significantly increased the amplitude of NMDAR currents without changing their decay kinetics. Interestingly, DHPG still increased the amplitude of NMDAR currents and caused reappearance of functional NMDAR channels after initially blocking NMDARs. In addition, protein kinase C (PKC) inhibition or intracellular dialysis with synaptosomal-associated protein of 25 kDa (SNAP-25)-blocking peptide abolished DHPG-induced increases in NMDAR currents of PVN neurons in SHRs. Our findings suggest that mGluR5 in the PVN is upregulated in hypertension and contributes to the hyperactivity of PVN presympathetic neurons through PKC- and SNAP-25-mediated surface expression of NMDARs.

Ang II–Salt Hypertension Depends on Neuronal Activity in the Hypothalamic Paraventricular Nucleus but Not on Local Actions of Tumor Necrosis Factor-α

Development of angiotensin II (Ang II)-dependent hypertension involves microglial activation and proinflammatory cytokine actions in the hypothalamic paraventricular nucleus (PVN). Cytokines activate receptor signaling pathways that can both acutely grade neuronal discharge and trigger long-term adaptive changes that modulate neuronal excitability through gene transcription. Here, we investigated contributions of PVN cytokines to maintenance of hypertension induced by subcutaneous infusion of Ang II (150 ng/kg per min) for 14 days in rats consuming a 2% NaCl diet. Results indicate that bilateral PVN inhibition with the GABA-A receptor agonist muscimol (100 pmol/50 nL) caused significantly greater reductions of renal and splanchnic sympathetic nerve activity (SNA) and mean arterial pressure in hypertensive than in normotensive rats (P<0.01). Thus, ongoing PVN neuronal activity seems required for support of hypertension. Next, the role of the prototypical cytokine tumor necrosis factor-α was investigated. Whereas PVN injection of tumor necrosis factor-α (0.3 pmol/50 nL) acutely increased lumbar and splanchnic SNA and mean arterial pressure, interfering with endogenous tumor necrosis factor-α by injection of etanercept (10 μg/50 nL) was without effect in hypertensive and normotensive rats. Next, we determined that although microglial activation in PVN was increased in hypertensive rats, bilateral injections of minocycline (0.5 μg/50 nL), an inhibitor of microglial activation, failed to reduce lumbar or splanchnic SNA or mean arterial pressure in hypertensive or in normotensive rats. Collectively, these findings indicate that established Ang II-salt hypertension is supported by PVN neuronal activity, but short term maintenance of SNA and arterial blood pressure does not depend on ongoing local actions of tumor necrosis factor-α.

Chronic depression of hypothalamic paraventricular neuronal activity produces sustained hypotension in hypertensive rats

Changes in the sympathetic nervous system are responsible for the initiation, development and maintenance of hypertension. An important central sympathoexcitatory region is the paraventricular nucleus (PVN) of the hypothalamus, which may become more active in hypertensive conditions, as shown in acute studies previously. Our objective was to depress PVN neuronal activity chronically by the overexpression of an inwardly rectifying potassium channel (hKir2.1), while evaluating the consequences on blood pressure (BP) and its reflex regulation. In spontaneously hypertensive rats (SHRs) and Wistar rats (WKY) lentiviral vectors (LVV-hKir2.1; LV-TREtight-Kir-cIRES-GFP5 4 × 10(9) IU and LV-Syn-Eff-G4BS-Syn-Tetoff 6.2 × 10(9) IU in a ratio 1:4) were stereotaxically microinjected bilaterally into the PVN. Sham-treated SHRs and WKY received bilateral PVN microinjections of LVV-eGFP (LV-Syn-Eff-G4BS-Syn-Tetoff 6.2 × 10(9) IU and LV-TREtight-GFP 5.7 × 10(9) IU in a ratio 1:4). Blood pressure was monitored continuously by radio-telemetry and evaluated over 75 days. Baroreflex gain was evaluated using phenylephrine (25 μg ml(-1), i.v.), whereas lobeline (25 μg ml(-1), i.v.) was used to stimulate peripheral chemoreceptors. In SHRs but not normotensive WKY rats, LVV-hKir2.1 expression in the PVN produced time-dependent and significant decreases in systolic (from 158 ± 3 to 132 ± 6 mmHg; P < 0.05) and diastolic BP (from 135 ± 4 to 113 ± 5 mmHg; P < 0.05). The systolic BP low-frequency band was reduced (from 0.79 ± 0.13 to 0.42 ± 0.09 mmHg(2); P < 0.05), suggesting reduced sympathetic vasomotor tone. Baroreflex gain was increased and peripheral chemoreflex depressed after PVN microinjection of LVV-hKir2.1. We conclude that the PVN plays a major role in long-term control of BP and sympathetic nervous system activity in SHRs. This is associated with reductions in both peripheral chemosensitivity and respiratory-induced sympathetic modulation and an improvement in baroreflex sensitivity. Our results support the PVN as a powerful site to control BP in neurogenic hypertension.

Chronic intermittent hypoxia increases sympathetic control of blood pressure: role of neuronal activity in the hypothalamic paraventricular nucleus

Like humans with sleep apnea, rats exposed to chronic intermittent hypoxia (CIH) experience arterial hypoxemias and develop hypertension characterized by exaggerated sympathetic nerve activity (SNA). To gain insights into the poorly understood mechanisms that initiate sleep apnea/CIH-associated hypertension, experiments were performed in rats exposed to CIH for only 7 days. Compared with sham-treated normoxic control rats, CIH-exposed rats (n = 8 rats/group) had significantly increased hematocrit (P < 0.001) and mean arterial pressure (MAP; P < 0.05). Blockade of ganglionic transmission caused a significantly (P < 0.05) greater reduction of MAP in rats exposed to CIH than control rats (n = 8 rats/group), indicating a greater contribution of SNA in the support of MAP even at this early stage of CIH hypertension. Chemical inhibition of neuronal discharge in the hypothalamic paraventricular nucleus (PVN) (100 pmol muscimol) had no effect on renal SNA but reduced lumbar SNA (P < 0.005) and MAP (P < 0.05) more in CIH-exposed rats (n = 8) than control rats (n = 7), indicating that CIH increased the contribution of PVN neuronal activity in the support of lumbar SNA and MAP. Because CIH activates brain regions controlling body fluid homeostasis, the effects of internal carotid artery injection of hypertonic saline were tested and determined to increase lumbar SNA more (P < 0.05) in CIH-exposed rats than in control rats (n = 9 rats/group). We conclude that neurogenic mechanisms are activated early in the development of CIH hypertension such that elevated MAP relies on increased sympathetic tonus and ongoing PVN neuronal activity. The increased sensitivity of Na(+)/osmosensitive circuitry in CIH-exposed rats suggests that early neuroadaptive responses among body fluid regulatory neurons could contribute to the initiation of CIH hypertension.

Rats selectively bred for differences in aerobic capacity have similar hypertensive responses to chronic intermittent hypoxia

Exposure to chronic intermittent hypoxia (CIH) is an animal model that mimics the repetitive bouts of hypoxemia experienced by humans with sleep apnea. Rats exposed to CIH develop hypertension that depends on the activation of sympathetic nerve activity (SNA). Since obesity and metabolic syndrome have been linked to neurogenic hypertension and sleep apnea, and because sleep apnea can adversely affect aerobic exercise capacity, we tested the hypothesis that rats bred for selection of low aerobic capacity running (LCR) would have a greater hypertensive response to CIH than rats bred for high aerobic capacity running (HCR). Blockade of ganglionic transmission was performed to compare the contribution of SNA to the maintenance of resting mean arterial pressure (MAP). Next, hypertensive responses to 7 days of CIH were compared across LCR and HCR rats (14-16 mo old). Finally, the contribution of the hypothalamic paraventricular nucleus (PVN) to the maintenance of SNA and hypertension after CIH was determined and compared across groups. Although LCR rats were less active and had greater body weights than HCR rats, resting MAP, the contribution of ongoing SNA to the maintenance of MAP, and hypertensive responses to CIH were similar between groups. Contrary to our hypothesis, chemical inhibition of the PVN with muscimol (1 mmol/100 nl) caused a larger fall of MAP in HCR rats than in LCR rats. We conclude that LCR rats do not have resting hypertension or an exaggerated hypertensive response to CIH. Interestingly, the maintenance of CIH hypertension in LCR rats compared with HCR rats appears less reliant on ongoing PVN neuronal activity.

Inhibition of Cyclooxygenase-2 Reduces Hypothalamic Excitation in Rats with Adriamycin-Induced Heart Failure

BackgroundThe paraventricular nucleus (PVN) of the hypothalamus plays an important role in the progression of heart failure (HF). We investigated whether cyclooxygenase-2 (COX-2) inhibition in the PVN attenuates the activities of sympathetic nervous system (SNS) and renin-angiotensin system (RAS) in rats with adriamycin-induced heart failure.Methodology/Principal FindingHeart failure was induced by intraperitoneal injection of adriamycin over a period of 2 weeks (cumulative dose of 15 mg/kg). On day 19, rats received intragastric administration daily with either COX-2 inhibitor celecoxib (CLB) or normal saline. Treatment with CLB reduced mortality and attenuated both myocardial atrophy and pulmonary congestion in HF rats. Compared with the HF rats, ventricle to body weight (VW/BW) and lung to body weight (LW/BW) ratios, heart rate (HR), left ventricular end-diastolic pressure (LVEDP), left ventricular peak systolic pressure (LVPSP) and maximum rate of change in left ventricular pressure (LV±dp/dtmax) were improved in HF+CLB rats. Angiotensin II (ANG II), norepinephrine (NE), COX-2 and glutamate (Glu) in the PVN were increased in HF rats. HF rats had higher levels of ANG II and NE in plasma, higher level of ANG II in myocardium, and lower levels of ANP in plasma and myocardium. Treatment with CLB attenuated these HF-induced changes. HF rats had more COX-2-positive neurons and more corticotropin releasing hormone (CRH) positive neurons in the PVN than did control rats. Treatment with CLB decreased COX-2-positive neurons and CRH positive neurons in the PVN of HF rats.ConclusionsThese results suggest that PVN COX-2 may be an intermediary step for PVN neuronal activation and excitatory neurotransmitter release, which further contributes to sympathoexcitation and RAS activation in adriamycin-induced heart failure. Treatment with COX-2 inhibitor attenuates sympathoexcitation and RAS activation in adriamycin-induced heart failure.

Transient Receptor Potential Vanilloid Type 1–Dependent Regulation of Liver-Related Neurons in the Paraventricular Nucleus of the Hypothalamus Diminished in the Type 1 Diabetic Mouse

The paraventricular nucleus (PVN) of the hypothalamus controls the autonomic neural output to the liver, thereby participating in the regulation of hepatic glucose production (HGP); nevertheless, mechanisms controlling the activity of liver-related PVN neurons are not known. Transient receptor potential vanilloid type 1 (TRPV1) is involved in glucose homeostasis and colocalizes with liver-related PVN neurons; however, the functional role of TRPV1 regarding liver-related PVN neurons has to be elucidated. A retrograde viral tracer was used to identify liver-related neurons within the brain-liver circuit in control, type 1 diabetic, and insulin-treated mice. Our data indicate that TRPV1 regulates liver-related PVN neurons. This TRPV1-dependent excitation diminished in type 1 diabetic mice. In vivo and in vitro insulin restored TRPV1 activity in a phosphatidylinositol 3-kinase/protein kinase C–dependent manner and stimulated TRPV1 receptor trafficking to the plasma membrane. There was no difference in total TRPV1 protein expression; however, increased phosphorylation of TRPV1 receptors was observed in type 1 diabetic mice. Our data demonstrate that TRPV1 plays a pivotal role in the regulation of liver-related PVN neurons. Moreover, TRPV1-dependent excitation of liver-related PVN neurons diminishes in type 1 diabetes, thus indicating that the brain-liver autonomic circuitry is altered in type 1 diabetes and may contribute to the autonomic dysfunction of HGP.

Casein Kinase 2-mediated Synaptic GluN2A Up-regulation Increases N-Methyl-d-aspartate Receptor Activity and Excitability of Hypothalamic Neurons in Hypertension

Increased glutamatergic input, particularly N-methyl-D-aspartate receptor (NMDAR) activity, in the paraventricular nucleus (PVN) of the hypothalamus is closely associated with high sympathetic outflow in essential hypertension. The molecular mechanisms underlying augmented NMDAR activity in hypertension are unclear. GluN2 subunit composition at the synaptic site critically determines NMDAR functional properties. Here, we found that evoked NMDAR-excitatory postsynaptic currents (EPSCs) of retrogradely labeled spinally projecting PVN neurons displayed a larger amplitude and shorter decay time in spontaneously hypertensive rats (SHRs) than in Wistar-Kyoto (WKY) rats. Blocking GluN2B caused a smaller decrease in NMDAR-EPSCs of PVN neurons in SHRs than in WKY rats. In contrast, GluN2A blockade resulted in a larger reduction in evoked NMDAR-EPSCs and puff NMDA-elicited currents of PVN neurons in SHRs than in WKY rats. Blocking presynaptic GluN2A, but not GluN2B, significantly reduced the frequency of miniature EPSCs and the firing activity of PVN neurons in SHRs. The mRNA and total protein levels of GluN2A and GluN2B in the PVN were greater in SHRs than in WKY rats. Furthermore, the GluN2B Ser(1480) phosphorylation level and the synaptosomal GluN2A protein level in the PVN were significantly higher in SHRs than in WKY rats. Inhibition of protein kinase CK2 normalized the GluN2B Ser(1480) phosphorylation level and the contribution of GluN2A to NMDAR-EPSCs and miniature EPSCs of PVN neurons in SHRs. Collectively, our findings suggest that CK2-mediated GluN2B phosphorylation contributes to increased synaptic GluN2A, which potentiates pre- and postsynaptic NMDAR activity and the excitability of PVN presympathetic neurons in hypertension.

Enhanced activation of RVLM-projecting PVN neurons in rats with chronic heart failure

Previous studies have indicated that there is increased activation of the paraventricular nucleus (PVN) in rats with chronic heart failure (CHF); however, it is not clear if the preautonomic neurons within the PVN are specifically overactive. Also, it is not known if these neurons have altered responses to baroreceptor or osmotic challenges. Experiments were conducted in rats with CHF (6-8 wk after coronary artery ligation). Spontaneously active neurons were recorded in the PVN, of which 36% were antidromically activated from the rostral ventrolateral medulla (RVLM). The baseline discharge rate in RVLM-projecting PVN (PVN-RVLM) neurons from CHF rats was significantly greater than in sham-operated (sham) rats (6.0 ± 0.6 vs. 2.6 ± 0.3 spikes/s, P < 0.05). Picoinjection of the N-methyl-D-aspartate (NMDA) receptor antagonist D,L-2-amino-5-phosphonovaleric acid significantly decreased the basal discharge of PVN-RVLM neurons by 80% in CHF rats compared with 37% in sham rats. Fifty-two percent of spontaneously active PVN-RVLM neurons responded to changes in the mean arterial pressure (MAP). The changes in discharge rate in PVN-RVLM neurons after a reduction in MAP (+52 ± 7% vs. +184 ± 61%) or an increase in MAP (-42 ± 8% vs. -71 ± 6%) were significantly attenuated in rats with CHF compared with sham rats. Most PVN-RVLM neurons (63%), including all barosensitive PVN-RVLM neurons, were excited by an internal carotid artery injection of hypertonic NaCl (2.1 osmol/l), whereas a smaller number (7%) were inhibited. The increase in discharge rate in PVN-RVLM neurons to hypertonic stimulation was significantly enhanced in rats with CHF compared with sham rats (134 ± 15% vs. 92 ± 13%). Taken together, these data suggest that PVN-RVLM neurons are more active under basal conditions and this overactivation is mediated by an enhanced glutamatergic tone in rats with CHF. Furthermore, this enhanced activation of PVN-RVLM neurons may contribute to the altered responses to baroreceptor and osmotic challenges observed during CHF.

Exposure to Acute Intermittent Hypoxia (IH) Prolongs Sympathoexcitatory and Pressor Responses Evoked by NMDA in the Hypothalamic Paraventricular Nucleus (PVN)

Exposure of rats to acute IH induces long term facilitation (LTF) of sympathetic nerve activity (SNA). Neurons of PVN subserve cardio‐respiratory function, project to brainstem and spinal cord regions controlling SNA, and are activated by short‐term (7 days) exposure to chronic IH. Here we tested the hypothesis that acute IH not only induces LTF of SNA, but facilitates sympathoexcitatory responses to acute PVN neuronal activation. Sprague Dawley rats (300–350g) were anesthetized (urethane/chloralose), ventilated (100% O2) and prepared for recording of splanchnic (n=3) or renal (n=3) SNA and mean arterial pressure (MAP, n=6). Responses to unilateral PVN injection of N‐Methyl‐D‐aspartic acid (NMDA, 200 pmol in 50 nl) were determined before and after exposure to acute IH (n=6), which was achieved by alternately changing FiO2 from 100% to 15% every 3 minutes. Compared to pre‐acute IH values, ongoing SNA, MAP and ETCO2 were unchanged after an average of 10 hypoxic cycles. Thus, LTF of SNA was not observed. Likewise, the amplitude of SNA and MAP responses to PVN NMDA was unaltered by acute IH. By contrast, whereas NMDA evoked responses of SNA and MAP prior to AIH were transient as expected, responses following acute IH were persistent (lasting &gt;1 h). These data suggest that exposure to acute IH prolongs sympathetic and pressor responses to PVN NMDA even in the absence of sympathetic LTF. Support: HL102310 &amp; HL 088052 (GMT)

Median preoptic nucleus and subfornical organ drive renal sympathetic nerve activity via a glutamatergic mechanism within the paraventricular nucleus.

The paraventricular nucleus (PVN) of the hypothalamus is involved in the neural control of sympathetic drive, but the precise mechanism(s) that influences the PVN is not known. The activation of the PVN may be influenced by input from higher forebrain areas, such as the median preoptic nucleus (MnPO) and the subfornical organ (SFO). We hypothesized that activation of the MnPO or SFO would drive the PVN through a glutamatergic pathway. Neuroanatomical connections were confirmed by the recovery of a retrograde tracer in the MnPO and SFO that was injected bilaterally into the PVN in rats. Microinjection of 200 pmol of N-methyl-d-aspartate (NMDA) or bicuculline-induced activation of the MnPO and increased renal sympathetic activity (RSNA), mean arterial pressure, and heart rate in anesthetized rats. These responses were attenuated by prior microinjection of a glutamate receptor blocker AP5 (4 nmol) into the PVN (NMDA - ΔRSNA 72 ± 8% vs. 5 ± 1%; P < 0.05). Using single-unit extracellular recording, we examined the effect of NMDA microinjection (200 pmol) into the MnPO on the firing activity of PVN neurons. Of the 11 active neurons in the PVN, 6 neurons were excited by 95 ± 17% (P < 0.05), 1 was inhibited by 57%, and 4 did not respond. The increased RSNA after activation of the SFO by ANG II (1 nmol) or bicuculline (200 pmol) was also reduced by AP5 in the PVN (for ANG II - ΔRSNA 46 ± 7% vs. 17 ± 4%; P < 0.05). Prior microinjection of ANG II type 1 receptor blocker losartan (4 nmol) into the PVN did not change the response to ANG II or bicuculline microinjection into the SFO. The results from this study demonstrate that the sympathoexcitation mediated by a glutamatergic mechanism in the PVN is partially driven by the activation of the MnPO or SFO.

Switch to glutamate receptor 2-lacking AMPA receptors increases neuronal excitability in hypothalamus and sympathetic drive in hypertension.

Glutamatergic synaptic input in the hypothalamic paraventricular nucleus (PVN) plays a critical role in regulating sympathetic outflow in hypertension. GluR2-lacking AMPA receptors (AMPARs) are permeable to Ca(2+), and their currents show unique inward rectification. However, little is known about changes in the AMPAR composition and its functional significance in hypertension. In this study, we found that AMPAR-mediated EPSCs (AMPAR-EPSCs) of retrogradely labeled spinally projecting PVN neurons exhibited a linear current-voltage relationship in Wistar-Kyoto (WKY) rats. However, AMPAR-EPSCs of labeled PVN neurons in spontaneously hypertensive rats (SHR) displayed inward rectification at positive holding potentials, which were not altered by lowering blood pressure with celiac ganglionectomy. Blocking GluR2-lacking AMPARs with 1-naphthyl acetyl spermine (NAS) caused a greater reduction in the AMPAR-EPSC amplitude and firing activity of PVN neurons in SHR than in WKY rats. Furthermore, blocking NMDA receptors and inhibition of calpain or calcineurin abolished inward rectification of AMPAR-EPSCs of PVN neurons in SHR. The GluR2 protein level was significantly less in the plasma membrane but greater in the cytosolic vesicle fraction in SHR than in WKY rats. In addition, microinjection of NAS into the PVN decreased blood pressure and lumbar sympathetic nerve activity in SHR but not in WKY rats. Our study reveals that increased GluR2-lacking AMPAR activity of PVN neurons results from GluR2 internalization through NMDA receptor-calpain-calcineurin signaling in hypertension. This phenotype switch in synaptic AMPARs contributes to increased excitability of PVN presympathetic neurons and sympathetic vasomotor tone in hypertension.

Water deprivation increases respiratory rhythmic sympathetic discharge through a mechanism that depends on neuronal activity in the hypothalamic paraventricular nucleus (PVN)

As in specific neurogenic cardiovascular diseases, activation of the hypothalamic PVN during water deprivation likely plays a key role in supporting sympathetic nerve activity (SNA) and arterial blood pressure (ABP). To determine if PVN support of SNA in water deprived (WD) rats involves increasing specific rhythmic bursting patterns, vagal intact euhydrated (control) and 48 h WD male rats were instrumented to record ABP, ECG, expired CO2, and splanchnic SNA (SSNA) or renal SNA (RSNA). Autospectral analysis (0.2 Hz resolution) revealed that respiratory power (1.3 - 1.7 Hz) of SSNA and RSNA was greater in WD rats (n=3/4) than controls (n=3). Bilateral inhibition of PVN with the GABA-AR agonist muscimol (100pmol in 100nl/side) reduced mean ABP (P<0.001), SSNA and RSNA (P<0.05/0.005) in WD rats but not controls. Muscimol also tended to reduce respiratory power of SSNA only in WD rats (P<0.09) such that respiratory power : total power (0.2 – 15 Hz) ratios were no longer different between groups. PVN muscimol did not affect respiratory power of RSNA in either group. These data indicate that PVN neuronal activity in WD rats supports respiratory rhythmic bursting of SSNA, but not RSNA. Ongoing studies seek to determine if reduced respiratory rhythmic SSNA by PVN muscimol in WD rats is a direct effect of suppressed PVN output to respiratory/sympathetic networks or an indirect effect of reduced mean ABP. Support: HL102310 (GMT)

Stimulation of the prelimbic cortex differentially modulates neuroendocrine responses to psychogenic and systemic stressors

The medial prefrontal cortex is important for normal regulation of stress responses, and is implicated in stress-related affective disease states (e.g. depression). In the current study, we investigated the role of the prelimbic division of the prefrontal cortex in control of responses to psychogenic and systemic stressors (restraint and hypoxia, respectively). Acute stimulation of the prelimbic cortical region with bicuculline methiodide (BMI) caused significant reduction of ACTH and corticosterone responses to restraint and reduced Fos activation of paraventricular nucleus neurons, consistent with a role in central inhibition of acute psychogenic stress responses. In contrast, BMI enhanced corticosterone (but not ACTH) responses to hypoxia via a mechanism suggestive of central PVN drive and enhanced adrenal sensitivity. Acute BMI increased restraint stress-induced Fos activation in known downstream targets of the prelimbic cortex (e.g., the basolateral amygdala and central amygdaloid nuclei), suggesting a connection between modulation of amygdalar signaling and stress inhibition. In contrast, hypoxia caused robust Fos activation in the basolateral and central amygdala, which was not affected by prelimbic BMI injection. The data suggest that the prelimbic cortex stimulation is sufficient to trigger inhibition of the HPA axis to psychogenic stress, but may play a very different role in enhancing HPA responsiveness to physical threats.

Sympathetic Response to Insulin Is Mediated by Melanocortin 3/4 Receptors in the Hypothalamic Paraventricular Nucleus

Hyperinsulinemia increases sympathetic nerve activity and contributes to cardiovascular dysfunction in obesity and diabetes. Neurons of the hypothalamic paraventricular nucleus (PVN) regulate sympathetic nerve activity through mono- and poly-synaptic connections to preganglionic neurons in the spinal cord. The purpose of the present study was to determine whether PVN neurons mediate the sympathetic response to insulin. Hyperinsulinemic-euglycemic clamps were performed in α-chloralose-anesthetized, male Sprague-Dawley rats (280-420 g) by an infusion of insulin (3.75 mU/kg per min) and 50% dextrose (0.75-2.0 mL/h) for 120 minutes. At 90 minutes, insulin significantly increased lumbar sympathetic nerve activity without any change in renal sympathetic nerve activity, heart rate, or blood glucose levels. Inhibition of the PVN with bilateral injection of the GABA(A) receptor agonist muscimol completely reversed the sympathoexcitatory response. However, direct injection of insulin into the PVN did not alter lumbar sympathetic nerve activity, and thereby suggests that insulin activates neurons upstream of the PVN. Interestingly, the sympathetic response to insulin was eliminated by PVN injection of the melanocortin 3/4 receptor antagonist SHU9119, but was unaffected by the angiotensin II type 1 receptor antagonist losartan. A final set of experiments suggests activation of PVN neurons during hyperinsulinemia increases glutamatergic drive to the rostral ventrolateral medulla. Collectively, these findings indicate that insulin activates a melanocortin-dependent pathway to the PVN that increases glutamatergic drive to the rostral ventrolateral medulla and alters cardiovascular function.

Chronic infusion of angiotensin receptor antagonists in the hypothalamic paraventricular nucleus prevents hypertension in a rat model of sleep apnea

Sleep apnea is characterized by increased sympathetic activity and is associated with systemic hypertension. Angiotensin (Ang) peptides have previously been shown to participate in the regulation of sympathetic tone and arterial pressure in the hypothalamic paraventricular nucleus (PVN) neurons. We investigated the role of endogenous Ang peptides within the PVN to control blood pressure in a rat model of sleep apnea-induced hypertension. Male Sprague–Dawley rats (250 g), instrumented with bilateral guide cannulae targeting the PVN, received chronic infusion of Ang antagonists (A-779, Ang-(1–7) antagonist; losartan and ZD7155, AT 1 antagonists; PD123319, AT 2 receptor antagonist, or saline vehicle). A separate group received an infusion of the GABA A receptor agonist (muscimol) to inhibit PVN neuronal activity independent of angiotensin receptors. After cannula placement, rats were exposed during their sleep period to eucapnic intermittent hypoxia (IH; nadir 5% O 2; 5% CO 2 to peak 21% O 2; 0% CO 2) 20 cycles/h, 7 h/day, for 14 days while mean arterial pressure (MAP) was measured by telemetry. In rats receiving saline, IH exposure significantly increased MAP (+ 12 ± 2 mm Hg vs. Sham −2 ± 1 mm Hg P < 0.01). Inhibition of PVN neurons with muscimol reversed the increase in MAP in IH rats (MUS: −9 ± 4 mm Hg vs. vehicle + 12 ± 2 mm Hg; P < 0.01). Infusion of any of the Ang antagonists also prevented the rise in MAP induced by IH (A-779: −5 ± 1 mm Hg, losartan: -9 ± 4 mm Hg, ZD7155: -11 ± 4 mm Hg and PD123319: -4 ± 3 mm Hg; P < 0.01). Our results suggest that endogenous Ang peptides acting in the PVN contribute to IH-induced increases in MAP observed in this rat model of sleep apnea-induced hypertension.

Adenosine inhibits paraventricular pre‐sympathetic neurons through ATP‐dependent potassium channels

Adenosine produces cardiovascular depressor effects in various brain regions. However, the cellular mechanisms underlying these effects remain unclear. The pre-sympathetic neurons in the hypothalamic paraventricular nucleus (PVN) play an important role in regulating arterial blood pressure and sympathetic outflow through projections to the spinal cord and brainstem. In this study, we performed whole-cell patch-clamp recordings on retrogradely labeled PVN neurons projecting to the intermediolateral cell column of the spinal cord in rats. Adenosine (10-100 microM) decreased the firing activity in a concentration-dependent manner, with a marked hyperpolarization in 12 of 26 neurons tested. Blockade of A(1) receptors with the adenosine A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine or intracellular dialysis of guanosine 5'-O-(2-thodiphosphate) eliminated the inhibitory effect of adenosine on labeled PVN neurons. Immunocytochemical labeling revealed that A(1) receptors were expressed on spinally projecting PVN neurons. Also, blocking ATP-dependent K(+) (K(ATP)) channels with 100 microM glibenclamide or 200 microM tolbutamide, but not the G protein-coupled inwardly rectifying K(+) channels blocker tertiapin-Q, abolished the inhibitory effect of adenosine on the firing activity of PVN neurons. Furthermore, glibenclamide or tolbutamide significantly decreased the adenosine-induced outward currents in labeled neurons. The reversal potential of adenosine-induced currents was close to the K(+) equilibrium potential. In addition, adenosine decreased the frequency of both spontaneous and miniature glutamatergic excitatory post-synaptic currents and GABAergic inhibitory post-synaptic currents in labeled neurons, and these effects were also blocked by 8-cyclopentyl-1,3-dipropylxanthine. Collectively, our findings suggest that adenosine inhibits the excitability of PVN pre-sympathetic neurons through A(1) receptor-mediated opening of K(ATP) channels.

Neuronal Activity in the Hypothalamic Paraventricular Nucleus Varies Across the Estrous Cycle in Anesthetized Female Rats: Effects of Dopamine Receptor Agonism

The central nervous system plays a pivotal role in sexual behavior. The role of the paraventricular nucleus (PVN) of the hypothalamus in female sexual behavior is poorly characterized. In males, there is a correlation between PVN neuron activity and erectile activity, and in mediating dopamine receptor agonist-induced sexual arousal. To understand the role of the PVN in female sexual function, baseline PVN neuronal activity and responses to dopamine receptor agonism were assessed in anesthetized rats. Single unit recordings were used to assess the firing properties of individual PVN neurons; and local field potentials quantified PVN network activity (combined activity of large numbers of PVN neurons). Baseline and apomorphine-stimulated activity was measured across the estrous cycle. Baseline firing rates of single units were found to differ across the stages of the estrous cycle with metestrus showing the highest firing rate (3.7 vs. 0.9 Hz in diestrus). Apomorphine administration caused significant increases in firing rate in 29% of neurons, and significant decreases in 71%. Basal local field potentials also varied between estrous stages and in response to apomorphine; significant differences in the total power of alpha1 and beta1 bands were observed in both cases. This study shows that PVN neuronal activity varies with hormonal state, and these neurons are differentially affected by apomorphine, suggesting two different populations. These data are indicative of a critical role for the PVN in female sexual function. It is important that the hormonal state should be considered when investigating sexual physiology and the effect of pharmacological agents.

Reduction in synaptic GABA release contributes to target-selective elevation of PVN neuronal activity in rats with myocardial infarction.

Neuronal activity in the paraventricular nucleus (PVN) is known to be elevated in rats with heart failure. However, the type of neurons involved and the underlying synaptic mechanisms remain unknown. Here we examined spontaneous firing activity and synaptic currents in presympathetic PVN neurons in rats with myocardial infarction (MI), using slice patch clamp combined with the retrograde labeling technique. In PVN neurons projecting to the rostral ventrolateral medulla (PVN-RVLM), MI induced a significant increase in basal firing rate (1.79 to 3.02 Hz, P < 0.05) and a reduction in the frequency of spontaneous (P < 0.05) and miniature (P < 0.01) inhibitory postsynaptic currents (IPSCs). In addition, MI induced an increase in the paired-pulse ratio of evoked IPSCs (P < 0.05). Bicuculline, a GABA(A) receptor antagonist, increased the firing rate of PVN-RVLM neurons in sham-operated (1.21 to 2.74 Hz, P < 0.05) but not MI (P > 0.05) rats. In contrast, in PVN neurons projecting to the intermediolateral horn of the spinal cord (PVN-IML), MI did not induce any significant changes in the basal firing rate and the properties of spontaneous and miniature IPSCs. The properties of spontaneous excitatory postsynaptic currents (EPSCs) were not altered in either neuron group. In conclusion, our results indicate that MI induces an elevation of firing activity in PVN-RVLM but not in PVN-IML neurons and that the elevated firing rate is largely due to a decrease in GABA release. These results provide evidence for a novel target-selective synaptic plasticity in the PVN that is associated with the sympathetic hyperactivity commonly seen in heart failure.

Regulation of tonic GABA inhibitory function, presympathetic neuronal activity and sympathetic outflow from the paraventricular nucleus by astroglial GABA transporters

Neuronal activity in the hypothalamic paraventricular nucleus (PVN), as well as sympathetic outflow from the PVN, is basally restrained by a GABAergic inhibitory tone. We recently showed that two complementary GABA(A) receptor-mediated modalities underlie inhibition of PVN neuronal activity: a synaptic, quantal inhibitory modality (IPSCs, I(phasic)) and a sustained, non-inactivating modality (I(tonic)). Here, we investigated the role of neuronal and/or glial GABA transporters (GATs) in modulating these inhibitory modalities, and assessed their impact on the activity of RVLM-projecting PVN neurons (PVN-RVLM neurons), and on PVN influence of renal sympathetic nerve activity (RSNA). Patch-clamp recordings were obtained from retrogradely labelled PVN-RVLM neurons in a slice preparation. The non-selective GAT blocker nipecotic acid (100-300 microM) caused a large increase in GABA(A)I(tonic), and reduced IPSC frequency. These effects were replicated by beta-alanine (100 microM), but not by SKF 89976A (30 microM), relatively selective blockers of GAT3 and GAT1 isoforms, respectively. Similar effects were evoked by the gliotoxin L-alpha-aminodipic acid (2 mM). GAT blockade attenuated the firing activity of PVN-RVLM neurons. Moreover, PVN microinjections of nipecotic acid in the whole animal diminished ongoing RSNA. A robust GAT3 immunoreactivity was observed in the PVN, which partially colocalized with the glial marker GFAP. Altogether, our results indicate that by modulating ambient GABA levels and the efficacy of GABA(A)I(tonic), PVN GATs, of a likely glial location, contribute to setting a basal tone of PVN-RVLM firing activity, and PVN-driven RSNA.