Blunted nitric oxide-mediated inhibition of renal nerve discharge within PVN of rats with heart failure

We have demonstrated a decreased neuronal nitric oxide (NO) synthase (nNOS) message in the hypothalamus of rats with heart failure (HF). Subsequently, we have demonstrated that NADPH diaphorase (a commonly used marker for nNOS activity) positive neurons are decreased in paraventricular nucleus (PVN) of rats with coronary artery ligation model of HF. The goal of the present study was to examine the influence of endogenous NO within the PVN on renal sympathetic nerve discharge (RSND) during HF. In alpha-chloralose- and urethane-anesthetized rats, an inhibitor of NO synthase, N(G)-monomethyl-L-arginine (L-NMMA) microinjected into the PVN (50, 100, and 200 pmol in 50-200 nl) produced a dose-dependent increase in RSND, blood pressure, and heart rate in control and HF rats. These responses were attenuated in rats with HF compared with control rats. On the other hand, the NO agonist, sodium nitroprusside, microinjected in PVN produced a dose-dependent decrease in RSND and blood pressure in control and HF rats. These responses were less in rats with HF compared with control rats. These data suggest that the endogenous NO-mediated effect within the PVN of HF rats is less potent in suppressing RSND compared with control rats. These data support the conclusion that the NO system within the PVN involved in controlling autonomic outflow is altered during HF and may contribute to the elevated levels of renal sympathoexcitation commonly observed in HF.     

Angiotensin-mediated increase in renal sympathetic nerve discharge within the PVN: role of nitric oxide

The paraventricular nucleus (PVN) of the hypothalamus is known to be an important site of integration in the central nervous system for sympathetic outflow. ANG II and nitric oxide (NO) play an important role in regulation of sympathetic nerve activity. The purpose of the present study was to examine how the interaction between NO and ANG II within the PVN affects sympathetic outflow in rats. Renal sympathetic nerve discharge (RSND), arterial blood pressure (AP), and heart rate (HR) were measured in response to administration of ANG II and N(G)-monomethyl-l-arginine (L-NMMA) into the PVN. Microinjection of ANG II (0.05, 0.5, and 1.0 nmol) into the PVN increased RSND, AP, and HR in a dose-dependent manner, resulting in increases of 53 +/- 9%, 19 +/- 3 mmHg, and 32 +/- 12 beats/min from baseline, respectively, at the highest dose. These responses were significantly enhanced by prior microinjection of L-NMMA and were blocked by losartan, an ANG II type 1 receptor antagonist. Similarly, administration of antisense to neuronal NO synthase within the PVN also potentiated the ANG II responses. Conversely, overexpression of neuronal NOS within the PVN with adenoviral gene transfer significantly attenuated ANG II responses. Push-pull administration of ANG II (1 nmol) into the PVN induced an increase in NO release. Our data indicate that ANG II type 1 receptors within the PVN mediate an excitatory effect on RSND, AP, and HR. NO in the PVN, which can be induced by ANG II stimulation, in turn inhibits the ANG II-mediated increase in sympathetic nerve activity. This negative-feedback mechanism within the PVN may play an important role in maintaining the overall balance and tone of sympathetic outflow.     

Interaction between glutamate and GABA systems in the integration of sympathetic outflow by the paraventricular nucleus of the hypothalamus

The paraventricular nucleus (PVN) of the hypothalamus is a central site known to modulate sympathetic outflow. Excitatory and inhibitory neurotransmitters within the PVN dictate final outflow. The goal of the present study was to examine the role of the interaction between the excitatory neurotransmitter glutamate and the inhibitory neurotransmitter GABA in the regulation of sympathetic activity. In alpha-chloralose- and urethane-anesthetized rats, microinjection of glutamate and N-methyl-D-aspartate (NMDA; 50, 100, and 200 pmol) into the PVN produced dose-dependent increases in renal sympathetic nerve activity, blood pressure, and heart rate. These responses were blocked by the NMDA receptor antagonist DL-2-amino-5-phosphonovaleric acid (AP-5). Microinjection of bicuculline, a GABA(A) receptor antagonist, into the PVN (50, 100, and 200 pmol) also produced significant, dose-dependent increases in renal sympathetic nerve activity, blood pressure, and heart rate; AP-5 also blocked these responses. Using microdialysis and HPLC/electrochemical detection techniques, we observed that bicuculline infusion into the PVN increased glutamate release. Using an in vitro hypothalamic slice preparation, we found that bicuculline increased the frequency of glutamate-mediated excitatory postsynaptic currents in PVN-rostral ventrolateral medullary projecting neurons, supporting a GABA(A)-mediated tonic inhibition of this excitatory input into these neurons. Together, these data indicate that 1) glutamate, via NMDA receptors, excites the presympathetic neurons within the PVN and increases sympathetic outflow and 2) this glutamate excitatory input is tonically inhibited by a GABA(A)-mediated mechanism.     

Role of the paraventricular nucleus in renal excretory responses to acute volume expansion: role of nitric oxide

Acute volume expansion (VE) produces a suppression of renal sympathetic nerve discharge (RSND) resulting in diuresis and natriuresis. Recently, we have demonstrated that the endogenous nitric oxide (NO) system within the paraventricular nucleus (PVN) produces a decrease in RSND. We hypothesized that endogenous NO in the PVN is involved in the suppression of RSND leading to diuretic and natriuretic responses to acute VE. To test this hypothesis, we first measured the VE-induced increase in renal sodium excretion and urine flow with and without blockade of NO, with microinjection of NG-monomethyl-L-arginine (L-NMMA; 200 pmol in 200 nl), within the PVN of Inactin-anesthetized male Sprague-Dawley rats. Acute VE produced significant increases in urine flow and sodium excretion, which were diminished in rats treated with L-NMMA within the PVN. This effect of NO blockade within the PVN on VE-induced diuresis and natriuresis was abolished by renal denervation. Consistent with these data, acute VE induced a decrease in RSND (52% of the baseline level), which was significantly blunted by prior administration of L-NMMA into the PVN (28% of the baseline level) induced by a comparable level of acute VE. Using the push-pull perfusion technique, we found that acute VE induced a significant increase in NOx concentration in the perfusate from the PVN region. Taken together, these results suggest that acute VE induces an increase in NO production within the PVN that leads to renal sympathoinhibition, resulting in diuresis and natriuresis. We conclude that NO within the PVN plays an important role in regulation of sodium and water excretions in the volume reflex via modulating renal sympathetic outflow.     

Exercise training improves endogenous nitric oxide mechanisms within the paraventricular nucleus in rats with heart failure

Previously, we have demonstrated that an altered endogenous nitric oxide (NO) mechanism within the paraventricular nucleus (PVN) contributes to increased renal sympathetic nerve activity (RSNA) in heart failure (HF) rats. The goal of this study was to examine the effect of exercise training (ExT) in improving the endogenous NO mechanism within the PVN involved in the regulation of RSNA in rats with HF. ExT significantly restored the decreased number of neuronal NO synthase (nNOS)-positive neurons in the PVN (129 +/- 17 vs. 99 +/- 6). nNOS mRNA expression and protein levels in the PVN were also significantly increased in HF-ExT rats compared with HF-sedentary rats. To examine the functional role of NO within the PVN, an inhibitor of NOS, N(G)-monomethyl-L-arginine, was microinjected into the PVN. Dose-dependent increases in RSNA, arterial blood pressure (BP), and heart rate (HR) were produced in all rats. There was a blunted increase in these parameters in HF rats compared with the sham-operated rats. ExT significantly augmented RSNA responses in rats with HF (33% vs. 20% at the highest dose), thus normalizing the responses. The NO donor sodium nitroprusside, microinjected into the PVN, produced dose-dependent decreases in RSNA, BP, and HR in both sham and HF rats. ExT significantly improved the blunted decrease in RSNA in HF rats (36% vs. 17% at the highest dose). In conclusion, our data indicate that ExT improves the altered NO mechanism within the PVN and restores NO-mediated changes in RSNA in rats with HF.     

Exercise training normalizes enhanced sympathetic activation from the paraventricular nucleus in chronic heart failure: role of angiotensin II

Exercise training (ExT) normalizes the increased sympathetic outflow in heart failure (HF), but the underlying mechanisms are not known. We hypothesized ExT would normalize the augmented activation of the paraventricular nucleus (PVN) via an angiotensinergic mechanism during HF. Four groups of rats used were the following: 1) sham-sedentary (Sed); 2) sham-ExT; 3) HF-Sed, and 4) HF-ExT. HF was induced by left coronary artery ligation. Four weeks after surgery, 3 wk of treadmill running was performed in ExT groups. The number of FosB-positive cells in the PVN was significantly increased in HF-Sed group compared with the sham-Sed group. ExT normalized (negated) this increase in the rats with HF. In anesthetized condition, the increases in renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP), and heart rate (HR) in response to microinjection of angiotensin (ANG) II (50～200 pmol) in the PVN of HF-Sed group were significantly greater than of the sham-Sed group. In the HF-ExT group the responses to microinjection of ANG II were not different from sham-Sed or sham-ExT groups. Blockade of ANG II type 1 (AT(1)) receptors with losartan in the PVN produced a significantly greater decrease in RSNA, MAP, and HR in HF-Sed group compared with sham-Sed group. ExT prevented the difference between HF and sham groups. AT(1) receptor protein expression was increased 50% in HF-Sed group compared with sham-Sed group. In the HF-ExT group, AT(1) receptor protein expression was not significantly different from sham-Sed or sham-ExT groups. In conclusion, one mechanism by which ExT alleviates elevated sympathetic outflow in HF may be through normalization of angiotensinergic mechanisms within the PVN.     

侧脑室注射一叶萩碱的心血管效应及作用机制

在麻醉大鼠侧脑室注射左旋一叶Ｑｉｕ碱（Ｌ－Ｓｅｃ）,记录动脉血压（ＡＰ）、心率（ＨＲ）及肾交感神经放电（ＲＳＮＤ）,观察前脑室周系统ＧＡＢＡ能紧张性活动改变引起的心血管效应。结果如下：（１）Ｌ－Ｓｅｃ可引起ＲＳＮＤ增加、ＡＰ升高和ＨＲ加快,并呈一定剂量－效应关系;但Ｌ－Ｓ盈余 于ｂｉｃｕｃｕｌｌｉｎｅ（Ｂｉｃ）。（２）Ｌ－Ｓｅｃ既能拮抗ｍｕｓｃｉｍｏｌ（Ｍｕｓ）,又能拮抗ｂａｃｌｏｆｅｎ（Ｂａｃ）     

Enhanced angiotensin II-mediated central sympathoexcitation in streptozotocin-induced diabetes: role of superoxide anion

Studies have shown that the superoxide mechanism is involved in angiotensin II (ANG II) signaling in the central nervous system. We hypothesized that ANG II activates sympathetic outflow by stimulation of superoxide anion in the paraventricular nucleus (PVN) of streptozotocin (STZ)-induced diabetic rats. In α-chloralose- and urethane-anesthetized rats, microinjection of ANG II into the PVN (50, 100, and 200 pmol) produced dose-dependent increases in renal sympathetic nerve activity (RSNA), arterial pressure (AP), and heart rate (HR) in control and STZ-induced diabetic rats. There was a potentiation of the increase in RSNA (35.0 ± 5.0 vs. 23.0 ± 4.3%, P < 0.05), AP, and HR due to ANG II type I (AT(1)) receptor activation in diabetic rats compared with control rats. Blocking endogenous AT(1) receptors within the PVN with AT(1) receptor antagonist losartan produced significantly greater decreases in RSNA, AP, and HR in diabetic rats compared with control rats. Concomitantly, there were significant increases in mRNA and protein expression of AT(1) receptor with increased superoxide levels and expression of NAD(P)H oxidase subunits p22(phox), p47(phox), and p67(phox) in the PVN of rats with diabetes. Pretreatment with losartan (10 mg·kg(-1)·day(-1) in drinking water for 3 wk) significantly reduced protein expression of NAD(P)H oxidase subunits (p22(phox) and p47(phox)) in the PVN of diabetic rats. Pretreatment with adenoviral vector-mediated overexpression of human cytoplasmic superoxide dismutase (AdCuZnSOD) within the PVN attenuated the increased central responses to ANG II in diabetes (RSNA: 20.4 ± 0.7 vs. 27.7 ± 2.1%, n = 6, P < 0.05). These data support the concept that superoxide anion contributes to an enhanced ANG II-mediated signaling in the PVN involved with the exaggerated sympathoexcitation in diabetes.     

Acute inhibition of the hypothalamic paraventricular nucleus decreases renal sympathetic nerve activity and arterial blood pressure in water-deprived rats

The present study was performed to determine whether sympathetic outflow and arterial blood pressure in water-deprived rats are dependent on the ongoing neuronal activity of the hypothalamic paraventricular nucleus (PVN). Renal sympathetic nerve activity (RSNA), mean arterial blood pressure (MAP), and heart rate were recorded in urethane-alpha-chloralose-anesthetized rats that were deprived of water but not food for 48 h before experiments. Acute inhibition of the PVN by bilateral microinjection of the GABA(A) agonist muscimol (100 pmol/side) significantly decreased RSNA in water-deprived rats (-26.7 +/- 4.7%, n = 7) but was without effect in control rats (1.3 +/- 6.3%, n = 7). Similarly, injection of muscimol produced a greater decrease in MAP in water-deprived rats than in control rats (-46 +/- 3 vs. -16 +/- 3 mmHg, respectively), although baseline MAP was not different between groups (105 +/- 4 vs. 107 +/- 4 mmHg, respectively). Neither bilateral microinjection of isotonic saline vehicle (100 nl/side) into the PVN nor muscimol (100 pmol/side) outside the PVN altered RSNA or MAP in either group. In addition, ganglionic blockade with hexamethonium (30 mg/kg i.v.) significantly decreased MAP in both groups; however, the decrease in MAP was significantly greater in water-deprived rats than in control rats (62 +/- 2 vs. 48 +/- 2 mmHg, respectively). Collectively, these findings suggest that sympathetic outflow contributes more to the maintenance of blood pressure in the water-deprived rat, and this depends, at least partly, on the ongoing activity of PVN neurons.     

Plasticity of pre- and postsynaptic GABAB receptor function in the paraventricular nucleus in spontaneously hypertensive rats

GABA(B) receptor function is upregulated in the paraventricular nucleus (PVN) of the hypothalamus in spontaneously hypertensive rats (SHR), but it is unclear whether this upregulation occurs pre- or postsynaptically. We therefore determined pre- and postsynaptic GABA(B) receptor function in retrogradely labeled spinally projecting PVN neurons using whole cell patch-clamp recording in brain slices in SHR and Wistar-Kyoto (WKY) rats. Bath application of the GABA(B) receptor agonist baclofen significantly decreased the spontaneous firing activity of labeled PVN neurons in both SHR and WKY rats. However, the magnitude of reduction in the firing rate was significantly greater in SHR than in WKY rats. Furthermore, baclofen produced larger membrane hyperpolarization and outward currents in labeled PVN neurons in SHR than in WKY rats. The baclofen-induced current was abolished by either including G protein inhibitor GDPbetaS in the pipette solution or bath application of the GABA(B) receptor antagonist in both SHR and WKY rats. Blocking N-methyl-d-aspartic acid receptors had no significant effect on baclofen-elicited outward currents in SHR. In addition, baclofen caused significantly greater inhibition of glutamatergic excitatory postsynaptic currents (EPSCs) in labeled PVN neurons in brain slices from SHR than WKY rats. By contrast, baclofen produced significantly less inhibition of GABAergic inhibitory postsynaptic currents (IPSCs) in labeled PVN neurons in SHR than in WKY rats. Although microinjection of the GABA(B) antagonist into the PVN increases sympathetic vasomotor tone in SHR, the GABA(B) antagonist did not affect EPSCs and IPSCs of the PVN neurons in vitro. These findings suggest that postsynaptic GABA(B) receptor function is upregulated in PVN presympathetic neurons in SHR. Whereas presynaptic GABA(B) receptor control of glutamatergic synaptic inputs is enhanced, presynaptic GABA(B) receptor control of GABAergic inputs in the PVN is attenuated in SHR. Changes in both pre- and postsynaptic GABA(B) receptors in the PVN may contribute to the control of sympathetic outflow in hypertension.     

Altered nitric oxide mechanism within the paraventricular nucleus contributes to the augmented carotid body chemoreflex in heart failure

Our previous study demonstrated a contribution of the paraventricular nucleus (PVN) of the hypothalamus in the processing of the carotid body (CB) chemoreflex. Nitric oxide (NO) (within the PVN), known to modulate autonomic function, is altered in rats with heart failure (HF). Therefore, the goal of the present study was to examine the influence of endogenous and exogenous NO within the PVN on the sympathoexcitatory component of the peripheral chemoreflex in normal and HF states. We measured mean arterial blood pressure, heart rate, renal sympathetic nerve activity (RSNA), and phrenic nerve activity (PNA) in sham-operated and HF rats (6-8 wk after coronary artery ligation) after incremental doses of potassium cyanide (25-100 mug/kg iv). There was potentiation of the reflex responses in HF compared with sham-operated rats. Bilateral microinjection of an inhibitor of NO synthase, N(G)-monomethyl-l-arginine (50 pmol), into the PVN augmented the RSNA and PNA response to peripheral chemoreceptor stimulation in sham-operated rats but had no effect in HF rats. Conversely, bilateral microinjection of a NO donor, sodium nitroprusside (50 nmol), into the PVN attenuated the RSNA response of the peripheral chemoreflex in sham-operated rats but to a smaller extent in HF rats. These data indicate that 1) NO within the PVN plays an important role in the processing of the CB chemoreflex and 2) there is an impairment of the NO function within the PVN of HF rats, which contributes to an augmented peripheral chemoreflex and subsequent elevation of sympathetic activity in HF.     

Role of paraventricular nucleus in the cardiogenic sympathetic reflex in rats

Myocardial ischemia stimulates cardiac spinal afferents to initiate a sympathoexcitatory reflex. However, the pathways responsible for generation of increased sympathetic outflow in this reflex are not fully known. In this study, we determined the role of the paraventricular nucleus (PVN) in the cardiogenic sympathetic reflex. Renal sympathetic nerve activity (RSNA) and blood pressure were recorded in anesthetized rats during epicardial application of 10 microg/ml bradykinin. Bilateral microinjection of muscimol (0.5 nmol), a GABA(A) receptor agonist, was performed to inhibit the PVN. In 10 vehicle-injected rats, epicardial bradykinin significantly increased RSNA 178.4 +/- 48.5% from baseline, and mean arterial pressure from 76.9 +/- 2.0 to 102.3 +/- 3.3 mmHg. Microinjection of muscimol into the PVN significantly reduced the basal blood pressure and RSNA (n = 12). After muscimol injection, the bradykinin-induced increases in RSNA (111.6 +/- 35.9% from baseline) and mean arterial pressure (61.2 +/- 1.3 to 74.5 +/- 2.7 mmHg) were significantly reduced compared with control responses. The response remained attenuated even when the basal blood pressure was restored to the control. In a separate group of rats (n = 9), bilateral microinjection of the ionotropic glutamate antagonist kynurenic acid (4.82 or 48.2 nmol in 50 nl) had no significant effect on the RSNA and blood pressure responses to bradykinin compared with controls. These results suggest that the tonic PVN activity is important for the full manifestation of the cardiogenic sympathoexcitatory response. However, ionotropic glutamate receptors in the PVN are not directly involved in this reflex response.     

Plasticity of GABAergic control of hypothalamic presympathetic neurons in hypertension

Increased sympathetic outflow contributes to the pathogenesis of hypertension. However, the mechanisms of increased sympathetic drive in hypertension remain unclear. We examined the tonic GABAergic inhibition in control of the excitability of paraventricular (PVN) presympathetic neurons in spontaneously hypertensive rats (SHR) and normotensive controls, including Sprague-Dawley (SD) and Wistar-Kyoto (WKY) rats. Whole cell patch-clamp recordings were performed on retrogradely labeled PVN neurons projecting to the rostral ventrolateral medulla (RVLM) in brain slices. The basal firing rate of PVN neurons was significantly decreased in 13-wk-old SD and WKY rats but increased in 13-wk-old SHR, compared with their respective 6-wk-old controls. The GABA(A) antagonist bicuculline consistently increased the firing of PVN neurons in normotensive controls. Surprisingly, bicuculline either decreased the firing or had no effect in 59.3% of labeled cells in 13-wk-old SHR. In contrast, the GABA(B) antagonist CGP-55845 had no effect on the firing of PVN neurons in normotensive controls but significantly increased the firing of 75% of cells studied in 13-wk-old SHR. Furthermore, the evoked GABA(A) current decreased significantly in labeled PVN neurons of 13-wk-old SHR compared with that in normotensive controls. Both the frequency and amplitude of GABAergic spontaneously inhibitory postsynaptic currents were also reduced in 13-wk-old SHR. This study demonstrates an unexpected functional change in GABA(A) and GABA(B) receptors in regulation of the firing activity of PVN-RVLM neurons in SHR. This change in GABA(A) receptor function and GABAergic inputs to PVN output neurons may contribute to increased sympathetic outflow in hypertension.     

Central neural control of sympathetic nerve activity in heart failure following exercise training

Typical characteristics of chronic congestive heart failure (HF) are increased sympathetic drive, altered autonomic reflexes, and altered body fluid regulation. These abnormalities lead to an increased risk of mortality, particularly in the late stage of chronic HF. Recent evidence suggests that central nervous system (CNS) mechanisms may be important in these abnormalities during HF. Exercise training (ExT) has emerged as a nonpharmacological therapeutic strategy substitute with significant benefit to patients with HF. Regular ExT improves functional capacity as well as quality of life and perhaps prognosis in chronic HF patients. The mechanism(s) by which ExT improves the clinical status of HF patients is not fully known. Recent studies have provided convincing evidence that ExT significantly alleviates the increased sympathetic drive, altered autonomic reflexes, and altered body fluid regulation in HF. This review describes and highlights the studies that examine various central pathways involved in autonomic outflow that are altered in HF and are improved following ExT. The increased sympathoexcitation is due to an imbalance between inhibitory and excitatory mechanisms within specific areas in the CNS such as the paraventricular nucleus (PVN) of the hypothalamus. Studies summarized here have revealed that ExT improves the altered inhibitory pathway utilizing nitric oxide and GABA mechanisms within the PVN in HF. ExT alleviates elevated sympathetic outflow in HF through normalization of excitatory glutamatergic and angiotensinergic mechanisms within the PVN. ExT also improves volume reflex function and thus fluid balance in HF. Preliminary observations also suggest that ExT induces structural neuroplasticity in the brain of rats with HF. We conclude that improvement of the enhanced CNS-mediated increase in sympathetic outflow, specifically to the kidneys related to fluid balance, contributes to the beneficial effects of ExT in HF.     

GABA in Paraventricular Nucleus Regulates Adipose Afferent Reflex in Rats

Background

Chemical stimulation of white adipose tissue (WAT) induces adipose afferent reflex (AAR), and thereby causes a general sympathetic activation. Paraventricular nucleus (PVN) is important in control of sympathetic outflow. This study was designed to investigate the role of γ-aminobutyric acid (GABA) in PVN in regulating the AAR.

Methodology/Principal Findings

Experiments were carried out in anesthetized rats. Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were continuously recorded. AAR was evaluated by the RSNA and MAP responses to electrical stimulation of the right epididymal WAT (eWAT) afferent nerve. Electrical stimulation of eWAT afferent nerve increase RSNA. Bilateral microinjection of the GABA_A receptor agonist isoguvacine or the GABA_B receptor agonist baclofen attenuated the AAR. The effect of isoguvacine on the AAR was greater than that of baclofen. The GABA_A receptor antagonist gabazine enhanced the AAR, while the GABA_B receptor antagonist CGP-35348 had no significant effect on the AAR. Bilateral PVN microinjection of vigabatrin, a selective GABA-transaminase inhibitor, to increase endogenous GABA levels in the PVN abolished the AAR. The inhibitory effect of vigabatrin on the AAR was attenuated by the pretreatment with gabazine or CGP-35348. Pretreatment with combined gabazine and CGP-35348 abolished the effects of vigabatrin.

Conclusions

Activation of GABA_A or GABA_B receptors in the PVN inhibits the AAR. Blockade of GABA_A receptors in the PVN enhances the AAR. Endogenous GABA in the PVN plays an important role in regulating the AAR.     

Paraventricular nucleus bicuculline alters frequency components of sympathetic nerve discharge bursts

Autospectral and coherence analyses were used to determine the effect of paraventricular nucleus (PVN) GABA(A) receptor antagonism [microinfusion or microinjections of bicuculline methiodide (BMI) 100 pmoles] on sympathetic nerve discharge (SND) frequency components (bursting pattern and relationships between discharges in regionally selective nerves) in alpha-chloralose-anesthetized rats. SND was recorded from the renal, splenic, and lumbar nerves. The following observations were made. First, PVN BMI microinjections, but not PVN saline or cortical BMI microinjections, transformed the cardiac-related SND bursting pattern in baroreceptor-innervated rats to one characterized by the presence of low-frequency bursts not synchronized to the cardiac cycle or phrenic nerve discharge bursts. Second, SND pattern changes were similar in the renal, splenic, and lumbar nerves, and peak coherence values relating low-frequency bursts in sympathetic nerve pairs (renal-splenic, renal-lumbar, and splenic-lumbar) were significantly increased from preinjection control after PVN BMI microinjection. Third, PVN BMI microinjections significantly increased the coupling between low-frequency SND bursts in baroreceptor-denervated rats. Finally, PVN BMI-induced changes in the SND bursting pattern were not observed after PVN pretreatment with muscimol (GABA agonist, 1 nmole). We conclude that PVN GABA(A) receptor antagonism profoundly alters the frequency components in sympathetic nerves.     

Exercise training normalizes enhanced glutamate-mediated sympathetic activation from the PVN in heart failure

Exercise training (ExT) normalizes the increased sympathetic outflow in heart failure (HF), but the mechanisms are not known. We hypothesized that ExT would normalize the augmented glutamatergic mechanisms mediated by N-methyl-d-aspartic acid (NMDA) receptors within the paraventricular nucleus (PVN) that occur with HF. Four groups of rats were used: 1) sham-operated (Sham) sedentary (Sed), 2) Sham ExT, 3) HF Sed, and 4) HF ExT. HF was induced by left coronary artery ligation, and ExT consisted of 3 wk of treadmill running. In alpha-chloralose-urethane-anesthetized rats, the increase in renal sympathetic nerve activity in response to the highest dose of NMDA (200 pmol) injected into the PVN in the HF Sed group was approximately twice that of the Sham Sed group. In the HF ExT group the response was not different from the Sham Sed and Sham ExT groups. Relative NMDA NR1 receptor subunit mRNA expression was 63% higher in the HF Sed group compared with the Sham Sed group but in the HF ExT group was not different from the Sham Sed and Sham ExT groups. NR1 receptor subunit protein expression was increased 87% in the HF Sed group compared with the Sham Sed group but in the HF ExT group was not significantly different from the Sham Sed and Sham ExT groups. Thus one mechanism by which ExT alleviates elevated sympathetic outflow in HF may be through normalization of glutamatergic mechanisms within the PVN.     

Blunted nitric oxide-mediated inhibition of sympathetic nerve activity within the paraventricular nucleus in diabetic rats

Recent evidence suggests that a central mechanism may be contributing to the sympathetic abnormality in diabetes. Nitric oxide (NO) has been known as a neurotransmitter in the central nervous system. The goal of this study was to examine the role of the endogenous NO system of the paraventricular nucleus (PVN) in regulation of renal sympathetic nerve activity (RSNA) in streptozotocin (STZ)-induced diabetic rats. The change in number of NADPH-diaphorase-positive neurons [a marker for neuronal NO synthase (nNOS) activity] in the PVN was measured. Diabetic rats were found to have significantly fewer nNOS positive cells in the PVN than in the control group (120 +/- 11 vs. 149 +/- 13, P < 0.05). Using RT PCR, Western blotting and immunofluorescent staining, it was also found that nNOS mRNA expression and protein level in the PVN were significantly decreased in the diabetic rats. Furthermore, using an in vivo microdialysis technique, we found that there was a lower NO(x) release from the PVN perfusates in rats with diabetes compared with the control rats (142 +/- 33 nM vs. 228 +/- 29 nM, P < 0.05). In alpha-chloralose- and urethane-anesthetized rats, an inhibitor of NO synthase, l-NMMA, microinjected into the PVN produced a dose-dependent increase in RSNA, mean arterial pressure (MAP), and heart rate (HR) in both control and diabetic rats. These responses were significantly attenuated in rats with diabetes compared with control rats (RSNA: 11 +/- 3% vs. 35 +/- 3%, P < 0.05). On the other hand, an NO donor, sodium nitroprusside (SNP), microinjected into the PVN produced a dose-dependent decrease in RSNA, MAP, and HR in the control and diabetic rats. RSNA (17 +/- 3%, vs. 41 +/- 6%, P < 0.05) and MAP in response to SNP were significantly blunted in the diabetic group compared with the control group. In conclusion, these data indicate an altered NO mechanism in the PVN of diabetic rats. This altered mechanism may contribute to the increased renal sympathetic neural activity observed in diabetes.     

Role of small conductance calcium-activated potassium channels expressed in PVN in regulating sympathetic nerve activity and arterial blood pressure in rats

Small conductance Ca(2+)-activated K(+) (SK) channels regulate membrane properties of rostral ventrolateral medulla (RVLM) projecting hypothalamic paraventricular nucleus (PVN) neurons and inhibition of SK channels increases in vitro excitability. Here, we determined in vivo the role of PVN SK channels in regulating sympathetic nerve activity (SNA) and mean arterial pressure (MAP). In anesthetized rats, bilateral PVN microinjection of SK channel blocker with peptide apamin (0, 0.125, 1.25, 3.75, 12.5, and 25 pmol) increased splanchnic SNA (SSNA), renal SNA (RSNA), MAP, and heart rate (HR) in a dose-dependent manner. Maximum increases in SSNA, RSNA, MAP, and HR elicited by apamin (12.5 pmol, n = 7) were 330 ± 40% (P < 0.01), 271 ± 40% (P < 0.01), 29 ± 4 mmHg (P < 0.01), and 34 ± 9 beats/min (P < 0.01), respectively. PVN injection of the nonpeptide SK channel blocker UCL1684 (250 pmol, n = 7) significantly increased SSNA (P < 0.05), RSNA (P < 0.05), MAP (P < 0.05), and HR (P < 0.05). Neither apamin injected outside the PVN (12.5 pmol, n = 6) nor peripheral administration of the same dose of apamin (12.5 pmol, n = 5) evoked any significant changes in the recorded variables. PVN-injected SK channel enhancer 5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one (DCEBIO, 5 nmol, n = 4) or N-cyclohexyl-N-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-4-pyrimidin]amine (CyPPA, 5 nmol, n = 6) did not significantly alter the SSNA, RSNA, MAP, and HR. Western blot and RT-PCR analysis of punched PVN tissue showed abundant expression of SK1-3 channels. We conclude that SK channels expressed in the PVN play an important role in the regulation of sympathetic outflow and cardiovascular function.     

GABA in the paraventricular nucleus tonically suppresses baroreflex function: alterations during pregnancy

It is well established that GABAergic inputs to the paraventricular nucleus of the hypothalamus (PVN) tonically suppress heart rate and the activity of several sympathetic nerves. However, whether GABA similarly inhibits PVN control of baroreflex function has not been previously investigated. To test this hypothesis, it was determined whether microinjection of the GABA(A) antagonist, bicuculline, into the PVN enhances the baroreflex in anesthetized female virgin rats. In addition, because GABAergic inhibition of PVN preautonomic neurons is decreased during pregnancy, it was also determined whether the effects of PVN bicuculline administration on baroreflex function were less in pregnant animals. In virgin rats, PVN microinjection of bicuculline increased (P < 0.05) baroreflex gain and maximum levels of heart rate (gain, from 1.6 ± 0.6 to 3.8 ± 1.3 bpm/mmHg; maximum, from 406 ± 18 to 475 ± 14 bpm) and of lumbar sympathetic nerve activity (gain from 2.6 ± 0.7 to 4.8 ± 1.6%/mmHg; maximum, 149 ± 32 to 273 ± 48%), indicating that PVN GABA normally suppresses baroreflex function. Pregnancy decreased heart rate baroreflex gain (pregnant, 0.9 ± 0.3 bpm/mmHg; virgin, 1.9 ± 0.2 bpm/mmHg; P < 0.05). Following PVN bicuculline administration in pregnant rats, smaller (P < 0.01) increments in baroreflex gain (pregnant, 0.6 ± 0.1 bpm/mmHg; virgin, 2.4 ± 0.9 bpm/mmHg) and maximum (pregnant, 33 ± 7 bpm; virgin, 75 ± 12 bpm; P < 0.05) were produced. Collectively, these data suggest that the PVN normally inhibits the baroreflex via tonic GABAergic inputs and that this inhibition is less during pregnancy.