Increased Baseline Mean Arterial Pressure Research Articles

Selective Afferent Denervation in Epididymal White Adipose Tissue Reduces Blood Pressure in Obese Male Mice Exposed to Early Life Stress

Visceral adiposity has been largely implicated in increased sympathetic activation, and pathogenesis and target organ damage associated with the development of obesity-induced hypertension. The experimental stimulation of afferent excitatory signals from adipose tissue has been shown to contribute with the increased sympathetic activation associated with obesity-induced hypertension as part of a sympathoexcitatory mechanism called the adipose afferent reflex (AAR). Previous studies from our lab have shown that male mice exposed to maternal separation and early weaning (MSEW), a mouse model of early life stress, display increased mean arterial pressure (MAP) compared to control mice when fed a high fat diet (HF) that correlates with increased neuronal activation in non-endocrine neurons in the posterior paraventricular nucleus of the hypothalamus. Moreover, MSEW-HF males display exacerbated blood pressure responses to the acute stimulation of the AAR in epididymal white adipose tissue (eWAT) compared to controls. Therefore, the aim of this study was to determine the contribution of the afferent sensory nerves innervating the eWAT in the chronically increased blood pressure displayed by obese MSEW male mice. Male C57BL/6J mice pups were separated from the dams from postnatal day (PD) 2 to PD 16 and weaned early on PD 17. Control litters remained undisturbed with the dams and were weaned on PD 21. After weaning, Control and MSEW mice were placed on HF (60% Kcal from fat) for four months. At 14 weeks, mice (n=8) were implanted with radiotelemetry for chronic MAP measurements. After two weeks of recovery, MAP baseline was recorded during five consecutive days followed by eWAT denervation. For this procedure, eWAT was exposed and infused with vehicle (0.6% ethanol in saline; 4ul; 8 sites; bilateral) or resiniferatoxin (RTX, 10 pmol/ul, 8 sites, 4ul; bilateral). Then, MAP was continuously recorded for 4 days. RTX procedure in eWAT was validated using a GFP reporter mouse for sensory neurons, B6.129P2(Cg)-Calcatm1.1(EGFP/HBEGF)Mjz/Mmnc, and the absence of a green signal in the RTX-treated areas confirmed the afferent denervation. Sympathetic index was evaluated before and after denervations by a single injection of a ganglion blocker (mecamylamine; 5mg/kg; ip). MSEW mice fed a HF displayed significantly increased baseline MAP compared to controls (120±2 vs. 110±2 mmHg respectively, p<0.05). Ganglion blockade induced a greater MAP reduction in MSEW compared to controls (Delta MAP: -20±2 vs. -12±2 mmHg, respectively, p<0.05). Sham denervation did not change MAP from baseline in both groups. However, RTX denervation only decrease MAP in MSEW-HF mice (baseline: 120±2 vs. RTX: 112±2 mmHg, p<0.05) and abolished the baseline MAP differences between groups. MAP decrease in response to ganglion blockade after RTX denervation was similar in both groups (MSEW: -14±4 vs. Control: -15±2 mmHg). Taken together these results show that afferent sensory nerves from epididymal white adipose tissue contribute to the exacerbated obesity-induced hypertension in male mice exposed to early life stress.

Abstract 106: Central Angiotensin II Mediates Neurogenic Hypertension Through Post-Translational Regulation of Neuronal Nitric Oxide Synthase in the Paraventricular Nucleus of the Hypothalamus

Activation of renin-angiotensin- system, nitric oxide (NO) bioavailability and subsequent sympathoexcitation plays a pivotal role in the pathogenesis of many cardiovascular diseases including hypertension. Previously we have shown Ang II enhances PIN (a protein inhibitor of nNOS: neuronal nitric oxide synthase, known to dissociate nNOS dimers into monomers) mediated ubiquitination of nNOS in the PVN of rats with heart failure. To further elucidate the mechanism by which Ang II increases sympathetic outflow by regulating PIN mediated nNOS ubiquitination, we used Sprague-Dawley rats (250-300 g) subjected to intracerebroventricular infusion of Ang II (20 ng/min, 14days, 0.5μl/h) through osmotic mini-pumps and NG108-15 hybrid neuronal cell line treated with Ang II as an in vitro model. Ang II infusion increased baseline mean arterial pressure (126 ± 9* vs. 84 ± 4 mmHg) and renal sympathetic nerve activity (20.5 ± 2.3* vs. 6.4 ± 1.9 % of Max. Activity). Ang II infusion increased the expression of PIN (1.36 ± 0.04* Ang II vs. 0.81 ± 0.02 Veh) with a concomitant 52% decrease in dimeric nNOS and PIN-Ub conjugates (0.73 ± 0.04* Ang II vs. 1 ± 0.03 Veh) in the PVN. Further, Ang II-mediated increase in PIN expression(0.67±0.06* CHX AngII vs. 0.40±0.08 CHX 0h) was independent of CHX (protein synthesis inhibitor) mediated decrease in PIN expression (0.41±0.06* CHX AngII vs. 0.19±0.04 CHX 4h) suggesting Ang II-mediated post-translational stabilization of PIN. Substrate-dependent ligase assay in cells transfected with pCMV-(HA-Ub)8 vector revealed a reduction of HA-Ub-PIN conjugates after Ang II and proteasome inhibitor lactacystin (LC) treatment (4.5 ± 0.6* LC Ang II vs. 9.2 ± 2.2 LC). TUBE (Tandem Ubiquitin-Binding Entities) assay showed decrease PIN-Ub conjugates in Ang II-treated cells (0.82 ± 0.12* LC Ang II vs. 1.23 ± 0.05 LC) while AT1R blocker Losartan(Los) treatment diminishes the Ang II-mediated stabilization of PIN (1.21 ± 0.07 LC Los vs. 1.14 ± 0.04* LC AngII Los). Taken together, our studies suggest that increased central levels of Ang II contribute to the enhanced expression of PIN leading to reduced expression of the dimeric form of nNOS, thus diminishing the inhibitory action of NO on pre-autonomic neurons in the PVN resulting in an increase in sympathetic outflow.

Central Angiotensin II regulates Protein Inhibitor of Neuronal Nitric Oxide Synthase through post‐translational mechanisms in the Paraventricular Nucleus resulting in increased Sympathetic outflow

Increased activation of Renin‐Angiotensin System in the paraventricular nucleus (PVN) of the hypothalamus has been associated with the increased sympathetic outflow observed in hypertension and chronic heart failure (CHF). Previously we have shown increased protein expression of the PIN (a protein inhibitor of nNOS: neuronal nitric oxide synthase, known to dissociate nNOS dimers into monomers) with concomitantly reduced levels of catalytically active dimers of nNOS in the PVN of rats with CHF. However, PIN transcripts levels remain unchanged. To elucidate the molecular mechanism of increased PIN expression post‐transcriptionally, we used Sprague‐Dawley rats (250–300 g) subjected to intracerebroventricular infusion of Ang II (20 ng/min, 14 days, 0.5 μl/h) through osmotic mini‐pumps and NG108‐15 hybrid neuronal cell line treated with Ang II as an in vitro model. Ang II infusion significantly increased baseline mean arterial pressure (126 ± 9 vs. 84 ± 4 mmHg) and baseline renal sympathetic nerve activity (20.5 ± 2.3 vs. 6.4 ± 1.9 % of Max activity). Ang II infusion increased the expression of the PIN (1.36 ± 0.04* Ang II vs. 0.81 ± 0.02 Veh) a with concomitant decrease in PIN‐Ub conjugates (0.73 ± 0.04* Ang II vs. 1 ± 0.03 Veh) in the PVN. Further, PIN translation was inhibited in vitro using protein synthesis inhibitor cycloheximide (CHX) for 0–4h after 20h of pretreatment with Ang II. Ang II‐mediated increase in PIN expression(0.67±0.06* CHX AngII vs. 0.40±0.08 CHX 0h) was independent of CHX mediated decrease in PIN expression (0.41±0.06* CHX AngII vs. 0.19±0.04 CHX 4h) suggesting Ang II‐mediated post‐translational stabilization of PIN. Ubiquitination assay in cells transfected with pCMV‐(HA‐Ub) 8 vector revealed a reduction of HA‐Ub‐PIN conjugates after Ang II and proteasome inhibitor lactacystin (LC) treatment (4.5 ± 0.6* LC Ang II vs. 9.2 ± 2.2 LC). TUBE (Tandem Ubiquitin‐Binding Entities) assay showed decrease PIN‐Ub conjugates in Ang II‐treated cells (0.82 ± 0.12* LC Ang II vs. 1.23 ± 0.05 LC) while AT1R blocker Losartan(Los) treatment diminishes the Ang II‐mediated stabilization of PIN (1.21 ± 0.07 LC Los vs. 1.14 ± 0.04* LC AngII Los). These results suggest that PIN is targeted for rapid degradation by the ubiquitin‐proteasome and Ang II via AT1R delays the rate of degradation resulting in accumulation of PIN. It of interest to note that previously we observed that there was a decreased accumulation of PIN‐Ub conjugates in the PVN of CHF rats compared to Sham (0.63 ± 0.05* CHF vs. 1.00 ± 0.10 Sham). Taken together, our studies suggest that increased central levels of Ang II contribute to the upregulation of PIN expression in the PVN via decreasing the ubiquitination which may be involved in reducing the expression of dimeric nNOS of pre‐autonomic neurons in the PVN resulting in the increased sympathetic outflow.Support or Funding InformationSupported by AHA ‐ 14SDG19980007 and NIH grants HL124104 &amp; HL62222.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Role of nucleus tractus solitarii (nTS) hydrogen sulfide (H2S) in hindlimb unloaded rats

Cardiovascular deconditioning alters central cardiorespiratory regulation and is mimicked by hindlimb unloading (HU). Preliminary data suggest the ventilatory response to hypoxia is augmented in HU rats. Furthermore, H2S from cystathionine beta synthase (CBS) augments excitatory neurotransmission in nTS, the initial site for cardiorespiratory sensory integration. Preliminary studies suggest nTS H2S augments phrenic (PNA) and splanchnic sympathetic (sSNA) nerve activity during acute hypoxia (AH) but its influence in HU rats is unknown. To test the hypothesis that increased endogenous H2S from CBS in the nTS enhances cardiorespiratory responses to AH after HU, we examined mean arterial pressure (MAP), PNA and sSNA responses to AH (2 min; 10% O2). Baseline cardiorespiratory parameters were similar in anesthetized control (C) and HU rats. Chemoreflex activation with AH increased PNA and sSNA and decreased MAP in both groups; the depressor response was less in HU rats. Bilateral nTS microinjection of the CBS inhibitor aminooxyacetate (AOA; 1 mM, 90 nl), to decrease H2S, increased baseline MAP and sSNA in both groups. In the presence of AOA, the increase in PNA amplitude to AH tended to be blunted in CC rats; this attenuation appeared to be less in HU. AOA did not alter the sSNA response to AH in CC rats, but reduced sSNA during AH after HU. These preliminary data suggest that CBS‐derived H2S in nTS augments cardiorespiratory responses to hypoxia and this modulation is diminished after HU. HL55306

Blunted endogenous GABA-mediated inhibition in the hypothalamic paraventricular nucleus of rats with streptozotocin-induced diabetes

The hypothalamic paraventricular nucleus (PVN) is involved in the regulation of sympathetic outflow and particularly affects the heart. This study sets out to determine the role of GABA of the paraventricular nucleus (PVN) in cardiovascular regulation in streptozotocin-induced diabetic rats. Pharmacological stimulation of glutamatergic receptors with DL-Homocysteic acid (200 mM in 100 nL) in the PVN region showed a significant depression in both mean arterial pressure (MAP) and heart rate (HR) of diabetic rats (Diabetic vs. non-diabetic: MAP 15.0 ± 1.5 vs. 35.8 ± 2.8 mmHg; HR 3.0 ± 2.0 vs. 30.0 ± 6.0 bpm, P < 0.05). Microinjection of bicuculline methiodide (1 mM in 100 nL), a GABAA receptor antagonist, produced an increase in baseline MAP and HR in both non-diabetic and diabetic rats. In the diabetic rats, bicuculline injection into the PVN reduced the pressor and HR responses (Diabetic vs. non-diabetic: MAP 6.2 ± 0.8 vs. 25.1 ± 2.2 mmHg; HR 1.8 ± 1.1 vs. 25.4 ± 6.2 bpm, P < 0.05). A microinjection of muscimol (2 mM in 100 nL), which is a GABAA receptor agonist, in the PVN elicited decreases in MAP and HR in both groups. The diabetic group showed a significantly blunted reduction in HR, but not MAP (Diabetic vs. non-diabetic: MAP -15.7 ± 4.0 vs. -25.0 ± 3.8 mmHg; HR -5.2 ± 2.1 vs. -39.1 ± 7.9 bpm). The blunted vasopressor and tachycardic responses to bicuculline microinjection in the diabetic rats are likely to result from decreased GABAergic inputs, attenuated release of endogenous GABA or alterations in GABAA receptors within the PVN.

Salt Intake Augments Hypotensive Effects of Transient Receptor Potential Vanilloid 4

To test the hypothesis that activation of the transient receptor potential vanilloid 4 (TRPV4) channel conveys a hypotensive effect that is enhanced during salt load, male Wistar rats fed a normal-sodium (0.5%) or high-sodium (HS; 4%) diet for 3 weeks were given 4α-phorbol 12,13-didecanoate (4α-PDD), a specific TRPV4 activator, in the presence or absence of capsazepine, a selective TRPV1 blocker, ruthenium red, a TRPV4 blocker, or TRPV4 small hairpin RNA that selectively knockdowns TRPV4. 4α-PDD (1, 2.5, or 5 mg/kg IV) dose-dependently decreased mean arterial pressure ( P &lt;0.05). HS enhanced 4α-PDD–induced depressor effects as well as 4α-PDD–mediated release of calcitonin gene–related peptide and substance P ( P &lt;0.001). Ruthenium red markedly blunted ( P &lt;0.001), whereas capsazepine slightly attenuated ( P &lt;0.05) 4α-PDD–induced depressor effects in HS and normal-sodium diet rats. Ruthenium red alone increased baseline mean arterial pressure in both HS and normal-sodium diet rats with a greater magnitude in the former ( P &lt;0.05). Western blot analysis showed that HS increased TRPV4 expression in dorsal root ganglia and mesenteric arteries ( P &lt;0.05) but not the renal cortex and medulla. Gene-silencing approach revealed that TRPV4 small hairpin RNA downregulated TRPV4 expression leading to blunted 4α-PDD–induced hypotension ( P &lt;0.05). Thus, TRPV4 activation decreases blood pressure in rats given a normal-sodium diet. HS enhances TRPV4 expression in sensory nerves/mesenteric arteries and TRPV4-mediated depressor effects and calcitonin gene–related peptide/substance P release such that HS causes a greater increase in blood pressure when TRPV4 is blocked. Our data indicate that TRPV4 activation may constitute a compensatory mechanism in preventing salt-induced increases in blood pressure.

GABA and nitric oxide in the PVN are involved in arterial pressure control but not in the chemoreflex responses in rats

GABAergic, nitrergic and glutamatergic mechanisms in the PVN on the baseline mean arterial pressure (MAP), heart rate (HR) and on the cardiovascular responses to chemoreflex activation in awake rat were evaluated. Chemoreflex was activated with KCN before and after microinjections into the PVN. Bicuculline into the PVN increased baseline MAP (94 ± 3 vs 113 ± 5 mmHg) and HR (350 ± 9 vs 439 ± 18 bpm) but had no effect on the pressor (49 ± 5 vs 47 ± 6 mmHg) or bradicardic (− 213 ± 23 vs − 256 ± 42 bpm) responses ( n = 7). Kynurenic acid into the PVN ( n = 6) produced no significant changes in the MAP (98 ± 3 vs 100 ± 3 mmHg), HR (330 ± 5 vs 339 ± 12 mmHg) or in the pressor (50 ± 4 vs 42 ± 4 mmHg) and bradicardic (− 252 ± 4 vs − 285 ± 16 bpm) responses to chemoreflex. L-NAME into the PVN ( n = 8) produced increase in the MAP (94 ± 3 vs 113 ± 5 mmHg) and HR (350 ± 9 vs 439 ± 18 bpm) but had no effect on the pressor (52 ± 5 vs 47 ± 6 mmHg) or bradicardic (− 253 ± 19 vs − 320 ± 25 bpm) responses to chemoreflex. We conclude that GABA A and nitric oxide in the PVN are involved in the maintenance of the baseline MAP but not in the modulation of the responses to chemoreflex. The results also show that Glutamate receptors in the PVN are not involved in maintenance of the baseline MAP, HR or in the cardiovascular responses to chemoreflex in awake rats.

Involvement of nitric oxide (NO) in the nucleus tractus solitarii (NTS) on respiratory but not in cardiovascular responses to chemoreflex activation in awake rats

There is evidence that NO is a neuromodulator in the NTS. In the present study we evaluated the effect of nitric oxide synthase (NOS) inhibition in the caudal nucleus tractus solitarii (cNTS) on: a) the baseline cardiovascular and respiratory functions, and b) the changes on mean arterial pressure (MAP), heart rate (HR) and respiratory frequency (RF) in response to chemoreflex activation in awake rats. Rats received bilateral guide‐cannulas in the cNTS and the femoral artery and vein were cannulated. RF was evaluated by body‐plethysmography. Chemoreflex was activated with KCN (iv) before and after bilateral microinjections of a non‐selective NOS inhibitor L‐NAME (n=8) or a selective neuronal NOS inhibitor N‐Propyl (n=7) into cNTS. Microinjection of L‐NAME into cNTS produced a significant increase in baseline MAP (100 ± 2 vs 112 ± 3 mmHg) but no changes in HR and RF. Microinjection of N‐Propyl into cNTS did not affect baseline MAP, HR and RF. Microinjection of L‐NAME (138 ± 10 vs 81 ± 11 cpm) or N‐Propyl (159 ± 13 vs 90 ± 17 cpm) into cNTS significantly reduced the tachypneic but had no effect on the magnitude of cardiovascular responses to chemoreflex. The data shows that the activation of the respiratory component of the chemorreflex in the caudal NTS involves nNOS‐derived NO production, while NO does not modulate the cardiovascular responses. Supported by CNPq and FAPESP.

Malnutrition enhances cardiovascular responses to chemoreflex activation in awake rats

Several studies in the literature suggest that low-protein intake is associated with increases in sympathetic efferent activity and cardiovascular disease. Among the possible mechanisms, changes in the neurotransmission of cardiovascular reflexes have been implicated. Therefore, the present study comprised the evaluation of chemoreflex responsiveness in rats subjected to a low-protein diet during the 35 days after weaning. As a result, we observed that malnourished rats presented higher levels of baseline mean arterial pressure and heart rate and exhibited a mild increase in the pressor response to chemoreflex activation. They also exhibited a massive bradycardic response to chemoreflex activation. Interestingly, bilateral ligature of the carotid body arteries further increased baseline mean arterial pressure and heart rate in malnourished animals. The data suggest severe autonomic imbalance and/or change in the central interplay between neural and cardiovascular mechanisms.

Endocannabinoid Regulates Blood Pressure via Activation of the Transient Receptor Potential Vanilloid Type 1 in Wistar Rats Fed a High-Salt Diet

This study was designed to examine the role of the endocannabinoids in blood pressure regulation during high sodium (HS) intake. HS (4% Na+ by weight) intake for 3 weeks increased baseline mean arterial pressure (MAP, mm Hg) compared with normal sodium (NS, 0.4% Na+ by weight)-treated male Wistar rats. Capsazepine (3 mg/kg), a selective transient receptor potential vanilloid type 1 (TRPV1) antagonist, caused a greater increase in MAP (mm Hg) in HS-treated compared with NS-treated rats (13+/-3 versus 4+/-2, p<0.05), whereas calcitonin gene-related peptide (CGRP) dose-dependently decreased MAP in both HS- and NS-treated rats with a more profound effect in the former. HS increased plasma anandamide levels analyzed by liquid chromatography/electrospray tandem mass spectrometry (NS, 2.40+/-0.31 versus HS, 4.05+/-0.47 pmol/ml, p<0.05) and plasma CGRP levels determined by radioimmunoassay (NS, 36.6+/-3.8 versus HS, 55.7+/-6.4 pg/ml, p<0.05). Methanandamide, a metabolically stable analog of anandamide, caused a greater CGRP release in mesenteric arteries isolated from HS-treated compared with NS-treated rats. Western blot showed that expression of receptor activity-modifying protein 1, a subunit of the CGRP receptor, in mesenteric arteries was greater in HS-treated compared with NS-treated rats. These results show that HS intake increases production of anandamide, which may serve as an endovanilloid to activate TRPV1, leading to release of CGRP to blunt salt-induced increases in blood pressure. These data support the notion that TRPV1 may act as a molecular target for salt-induced elevation of endovanilloid compounds to regulate blood pressure.

Midbrain modulation of the cardiac baroreflex involves excitation of lateral parabrachial neurons in the rat

Activation of the dorsal periaqueductal gray (PAG) evokes defense-like behavior including a marked increase in sympathetic drive and resetting of baroreflex function. The goal of this study was to investigate the role of the lateral parabrachial nucleus (LPBN) in mediating dorsal PAG modulation of the arterial baroreflex. Reflex responses were elicited by electrical stimulation of the aortic depressor nerve (ADN) at 5 Hz or 15 Hz in urethane anesthetized rats ( n = 18). Electrical stimulation of the dorsal PAG at 10 Hz did not alter baseline mean arterial pressure (MAP) but did significantly attenuate baroreflex control of heart rate (HR) evoked by low frequency ADN stimulation. Alternatively, 40 Hz dorsal PAG stimulation increased baseline MAP (43 ± 3 mm Hg) and HR (33 ± 3 bpm) and attenuated baroreflex control of HR at both ADN stimulation frequencies. Reflex control of MAP was generally unchanged by dorsal PAG stimulation. Bilateral inhibition of neurons in LPBN area ( n = 6) with muscimol (0.45 nmol per side) reduced dorsal PAG-evoked increases in MAP and HR by 50 ± 4% and 95 ± 4%, respectively, and significantly reduced, but did not completely eliminate dorsal PAG attenuation of the cardiac baroreflex. Bilateral blockade of glutamate receptors in the LPBN area ( n = 6) with kynurenic acid (1.8 nmol) had a similar effect on dorsal PAG-evoked increases in MAP, HR and cardiac baroreflex function. Reflex control of MAP was unchanged with either treatment. These findings suggest that the LPBN area is one of several brainstem regions involved in descending modulation of the cardiac baroreflex function during defensive behavior.

Impaired in vivo venous constriction in conscious obese Zucker rats with metabolic syndrome

The venous system plays a crucial role in regulating cardiac output and blood pressure. Although the relationship between obesity and hypertension is well recognized, little is known about the effect of obesity on venous function. We examined if 16-week-old obese Zucker rats, relative to age-matched lean Zucker rats, had altered in vivo venoconstriction to noradrenaline. The obese rats, compared to the controls, had higher mean arterial pressure (MAP), body weight, and plasma insulin and triglycerides, but reduced pressor and mean circulatory filling pressure (MCFP, index of venous tone) responses to noradrenaline (2.5-30x10(-9) mol/kg/min, i.v.). N(G)-nitro-L-arginine methyl ester (L-NAME, 8 mg/kg, i.v., non-selective inhibitor of nitric oxide synthase) did not alter MCFP in either group, but increased MAP of both groups, though the increase was markedly less in the obese than lean rats. Therefore, obese Zucker rats had increased baseline MAP, but impaired in vivo pressor and MCFP responses to noradrenaline, and reduced pressor response to L-NAME. The increased baseline MAP in the obese rats was not due to increased arterial and venous constriction to noradrenaline but rather to reduced influence of the nitric oxide/L-arginine system.

Antihypertensive Responses Elicited by Central Moxonidine in Rats: Possible Role of Nitric Oxide

In the present study, we investigated the effects of pretreatment with NG-nitro-L-arginine methyl ester (L-NAME) (nitric oxide synthase inhibitor) injected intravenously (IV) on the hypotension, bradycardia, and vasodilation produced by moxonidine (alpha2-adrenergic/imidazoline receptor agonist) injected into the fourth brain ventricle (4th V) in rats submitted to acute hypertension that results from baroreflex blockade by bilateral injections of kynurenic acid (kyn, glutamatergic receptor antagonist) into the nucleus of the solitary tract (NTS) or in normotensive rats. Male Wistar rats (n=5 to 7/group) anesthetized with IV urethane (1.0 g kg(-1) of body weight) and alpha-chloralose (60 mg kg(-1) of body weight) were used. Bilateral injections of kyn (2.7 nmol 100 nL(-1)) into the NTS increased baseline mean arterial pressure (148 +/- 11 mm Hg, vs. control: 102 +/- 4 mm Hg) and baseline heart rate (417 +/- 11 bpm, vs. control: 379 +/- 6 bpm). Moxonidine (20 nmol microL(-1)) into the 4th V reduced mean arterial pressure and heart rate to similar levels in rats treated with kyn into the NTS (68 +/- 9 mm Hg and 359 +/- 7 bpm) or in control normotensive rats (66 +/- 7 mm Hg and 362 +/- 8 bpm, respectively). The pretreatment with L-NAME (25 micromol kg, IV) attenuated the hypotension produced by moxonidine into the 4th V in rats treated with kyn (104 +/- 6 mm Hg) or in normotensive rats (95 +/- 8 mm Hg), without changing bradycardia. Moxonidine into the 4th V also reduced renal, mesenteric, and hindquarter vascular resistances in rats treated or not with kyn into the NTS and the pretreatment with L-NAME IV reduced these effects of moxonidine. Therefore, these data indicate that nitric oxide mechanisms are involved in hypotension and mesenteric, renal, and hindquarter vasodilation induced by central moxonidine in normotensive and in acute hypertensive rats.

Chronic glucose infusion significantly decreases 24 hr/day renal blood flow and increases arterial pressure in dogs with reduced renal mass

Chronic insulin + glucose infusion decreases mean arterial pressure (MAP) and causes renal and systemic vasodilation in normal dogs, obese dogs, and dogs with reduced renal mass (RRM) and high Na+ intake, but the depressor response may be due to excessive insulin infusion. This study tested the hypothesis that a 6-day glucose infusion would increase MAP in dogs with RRM (~70%) and high Na+ intake (~ 300 mEq/d). 4 male RRM dogs were instrumented with a Transonic flow probe on the remaining renal artery and housed in metabolic cages, with an electrical swivel for 24 hr/d RBF measurement and a hydraulic swivel for continuous iv. infusion of vehicle or glucose (450 gm/d). MAP was measured with a Data Sciences catheter. MAP averaged 106±1 mmHg during control, 117±5 mmHg by day 3 of glucose, but was back to 104±6 mmHg by day 6 of glucose. RBF, however, averaged 73±2 ml/min during baseline and, surprisingly, decreased progressively to 79±1% of control by day 6, and recovered when glucose was stopped; GFR increased 21% during glucose infusion. Repeating the study with a continuous background iv. AngII infusion (1 ng/kg/min) increased baseline MAP to 115±1 mmHg, but MAP increased progressively during glucose to 125±8 mmHg by day 6. The renal hemodynamic responses were similar to study 1, but cumulative Na+ retention was less. These results reveal mild pressor and significant renal vasoconstrictor responses to glucose infusion in dogs. The novel RBF data provide new evidence suggesting increased filtration fraction may play a role in the Na+ retention response to high glucose, but the effect of AngII on Na+ balance also suggests a role for systemic vasodilation. (Supported by HL 56259 and HL75625.)

Antagonism of glutamate receptors in the intermediate and caudal NTS of awake rats produced no changes in the hypertensive response to chemoreflex activation

The role of excitatory amino acid (EAA) receptors in the neurotransmission of the sympathoexcitatory component of the chemoreflex (pressor response) in the intermediate and caudal aspects of the commissural nucleus tractus solitarii (NTS) of awake rats was evaluated. Microinjection of kynurenic acid, a non-selective antagonist of EAA receptors, into the intermediate and caudal commissural NTS produced a large increase in the baseline mean arterial pressure (MAP), which may reduce the magnitude of the pressor response to chemoreflex activation. To avoid this problem sodium nitroprusside (SNP) was infused (i.v.) after microinjections of kynurenic acid (2 nmol/50 nl) into the NTS, in order to normalize the MAP and then the chemoreflex was activated and the magnitude of the pressor response evaluated. Microinjection of kynurenic acid into the intermediate (bilaterally) and caudal (midline) commissural NTS ( n=6) produced a significant increase in baseline MAP (103±5 vs. 137±6 mm Hg) normalized by SNP infusion (107±4 mm Hg) and under this experimental condition the pressor response to chemoreflex activation was not statistically different in relation to the control (37±7 vs. 44±6 mm Hg). Bilateral microinjections of kynurenic acid into the caudal NTS ( n=8) also produced a significant increase in baseline MAP (109±4 vs. 145±6 mm Hg) normalized by SNP infusion (109±6 mm Hg). After normalization of MAP, the pressor response to chemoreflex activation at 3 (34±6 mm Hg) and 10 min (37±6 mm Hg) was also not different in relation to the control (46±5 mm Hg). These data indicate that the antagonism of EAA receptors simultaneously in the intermediate (bilateral) and caudal (midline) commissural NTS or only in the caudal commissural NTS (bilateral) of awake rats had no effect on the hypertensive response to chemoreflex activation. We suggest that neurotransmitter other than l-glutamate may take part in the neurotransmission of the sympathoexcitatory component of the chemoreflex at the NTS level.

Pressor response to chemoreflex activation before and after microinjection of glycine into the NTS of awake rats.

Microinjection of glycine into the rostral (bilateral) and caudal (midline) commissural nucleus of the solitary tract (NTS) using three guide cannulas implanted in the direction of these sites produced an increase in mean arterial pressure (MAP) and abolished the pressor response to chemoreflex activation [potassium cyanide (n = 7)]. Strychnine, a glycine receptor antagonist, attenuated the increase in MAP, and in this new experimental condition (n = 5) the pressor response to chemoreflex activation was not altered. Considering that the effect of glycine on the attenuation of the pressor response to chemoreflex activation could be secondary to the increase in baseline MAP, in a third group of rats (n = 5) sodium nitroprusside infusion (intravenous) after microinjections of glycine into the NTS normalizes MAP. In this case, the pressor response to chemoreflex activation was similar to the control. These data show that glycine when microinjected bilaterally into the lateral commissural NTS as well as into the medial commissural NTS plays no major inhibitory role in the processing of the neurotransmission of the sympathoexcitatory component of the chemoreflex.

Role of sensory nervous system vasoactive peptides in hypertension.

The goal of the present research was to elucidate the roles and mechanisms by which the sensory nervous system, through the actions of potent vasodilator neuropeptides, regulates cardiovascular function in both the normal state and in the pathophysiology of hypertension. The animal models of acquired hypertension studied were deoxycorticosterone-salt (DOC-salt), subtotal nephrectomy-salt (SN-salt), and Nomega-nitro-L-arginine methyl ester (L-NAME)-induced hypertension during pregnancy in rats. The genetic model was the spontaneously hypertensive rat (SHR). Calcitonin gene-related peptide (CGRP) and substance P (SP) are potent vasodilating neuropeptides. In the acquired models of hypertension, CGRP and SP play compensatory roles to buffer the blood pressure (BP) increase. Their synthesis and release are increased in the DOC-salt model but not in the SN-salt model. This suggests that the mechanism by which both models lower BP in SN-salt rats is by increased vascular sensitivity. CGRP functions in a similar manner in the L-NAME model. In the SHR, synthesis of CGRP and SP is decreased. This could contribute to the BP elevation in this model. The CGRP gene knockout mouse has increased baseline mean arterial pressure. The long-term synthesis and release of CGRP is increased by nerve growth factor, bradykinin, and prostaglandins and is decreased by alpha2-adrenoreceptor agonists and glucocorticoids. In several animal models, sensory nervous system vasoactive peptides play a role in chronic BP elevation. In the acquired models, they play a compensatory role. In the genetic model, their decreased levels may contribute to the elevated BP. The roles of CGRP and SP in human hypertension are yet to be clarified.

Antagonism of adenosine A(1) receptors in the NTS does not affect the chemoreflex in awake rats.

The possible involvement of adenosine A(1) receptors in neurotransmission of the sympathoexcitatory component of the chemoreflex in the nucleus tractus solitarii (NTS) of awake rats was evaluated. Unilateral microinjection of increasing doses of adenosine (0.01, 0.06, 0.12, 1.25, 2.5, and 5.0 nmol/50 nl) into the lateral aspect of the commissural NTS produced a long-lasting increase in baseline mean arterial pressure (MAP) and no changes in baseline heart rate (HR). Microinjection of adenosine at 1.25 nmol/50 nl (ED(50)) into the NTS (n = 9) produced a significant increase in baseline MAP (119 +/- 3, 122 +/- 4, and 117 +/- 4 mmHg at 30 s, 1 min, and 2 min, respectively) compared with control (102 +/- 3 mmHg) but no significant changes after previous microinjection of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), an adenosine A(1) receptor antagonist (107 +/- 3, 107 +/- 3, and 106 +/- 3 mmHg at 30 s, 1 min, and 2 min, respectively) compared with control (102 +/- 3 mmHg). Microinjection of adenosine before and after DPCPX into the same site of the lateral commissural NTS produced no changes in baseline HR. In another group of rats (n = 8), microinjection of DPCPX (0.285 nmol/50 nl) into lateral and midline aspects of the commissural NTS produced no significant changes in pressor (+46 +/- 4 vs. +47 +/- 2 mmHg) or bradycardic responses (-216 +/- 9 vs. -226 +/- 12 beats/min) to chemoreflex activation with intravenous potassium cyanide compared with control responses. These data show that microinjection of adenosine into the NTS produced a small and long-lasting pressor response by activating A(1) receptors and that blockade of these receptors produced no changes in cardiovascular responses to chemoreflex activation. We conclude that adenosine A(1) receptors are not involved in processing of the chemoreflex afferents at the NTS level.

Nitric oxide modulates sympathoexcitatory cardiac-cardiovascular reflexes elicited by bradykinin.

A number of studies have demonstrated an important role for nitric oxide (NO) in central and peripheral neural modulation of sympathetic activity. To assess the interaction and integrative effects of NO release and sympathetic reflex actions, we investigated the influence of inhibition of NO on cardiac-cardiovascular reflexes. In anesthetized, sinoaortic-denervated and vagotomized cats, transient reflex increases in arterial blood pressure (BP) were induced by application of bradykinin (BK, 0.1-10 microg/ml) to the epicardial surface of the heart. The nonspecific NO synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA, 10 mg/kg iv) was then administered and stimulation was repeated. L-NMMA increased baseline mean arterial pressure (MAP) from 129 +/- 8 to 152 +/- 9 mmHg and enhanced the change in MAP in response to BK from 32 +/- 3 to 39 +/- 5 mmHg (n = 9, P < 0.05). Pulse pressure was significantly enhanced during the reflex response from 6 +/- 4 to 27 +/- 6 mmHg after L-NMMA injection due to relatively greater potentiation of the rise in systolic BP. Both the increase in baseline BP and the enhanced pressor reflex were reversed by L-arginine (30 mg/kg iv). Because L-NMMA can inhibit both brain and endothelial NOS, the effects of 7-nitroindazole (7-NI, 25 mg/kg ip), a selective brain NOS inhibitor, on the BK-induced cardiac-cardiovascular pressor reflex also were examined. In contrast to L-NMMA, we observed significant reduction of the pressor response to BK from 37 +/- 5 to 18 +/- 3 mmHg 30 min after the administration of 7-NI (n = 9, P < 0.05), an effect that was reversed by L-arginine (300 mg/kg iv, n = 7). In a vehicle control group for 7-NI (10 ml of peanut oil ip), the pressor response to BK remained unchanged (n = 6, P > 0.05). In conclusion, neuronal NOS facilitates, whereas endothelial NOS modulates, the excitatory cardiovascular reflex elicited by chemical stimulation of sympathetic cardiac afferents.

AT1 receptor blockers prevent sympathetic hyperactivity and hypertension by chronic ouabain and hypertonic saline

Sympathetic hyperactivity and hypertension caused by chronic treatment with ouabain or sodium-rich artificial cerebrospinal fluid (aCSF) can be prevented by central administration of an angiotensin type 1 (AT(1)) receptor blocker. In the present study, we assessed whether, in Wistar rats, chronic peripheral treatment with the AT(1) receptor blockers losartan and embusartan can exert sufficient central effects to prevent these central effects of ouabain and sodium. Losartan or embusartan (both at 100 mg x kg(-1) x day(-1)) were given subcutaneously once daily. Ouabain (50 microg/day) was infused subcutaneously, and sodium-rich aCSF (1.2 M Na(+), 5 microl/h) was infused intracerebroventricularly, both by osmotic minipump for 13-14 days. The mean arterial pressure (MAP) at rest and in response to air stress and intracerebroventricularly injection of guanabenz (75 microg/7.5 microl), ANG II (30 ng/3 microl), and ouabain (0.5 microg/2 microl) were then measured. In control rats, chronic treatment with ouabain subcutaneously and hypertonic saline intracerebroventricularly both increased baseline MAP by 20-25 mmHg and enhanced twofold the pressor responses to air stress and depressor responses to the alpha(2)-adrenoceptor agonist guanabenz. Simultaneous treatment with losartan or embusartan fully prevented hypertension, maintained normal responses to air stress and guanabenz, and attenuated pressor responses to acute intracerebroventricular injection of ANG II and ouabain. We concluded that peripheral administration of losartan as well as embusartan can cause sufficient central effects to prevent the sympathetic hyperactivity and hypertension induced by chronic peripheral ouabain and central sodium.