Role of the hypothalamic PVN in the reflex reduction in mesenteric blood flow elicited by hyperthermia

The hypothalamic paraventricular nucleus (PVN) is an important integrative center in the brain. In the present study, we investigated whether the PVN is a key region in the mesenteric vasoconstriction that normally accompanies an increase in core body temperature. Anesthetized rats were monitored for blood pressure, heart rate, mesenteric blood flow, and vascular conductance. In control rats, elevation of core body temperature to 41 degrees C had no significant effect on blood pressure, increased heart rate, and reduced mesenteric blood flow by 21%. In a separate group of rats, muscimol was microinjected bilaterally (1 nmol/side) into the PVN. Compared with the control group, there was no significant difference in the blood pressure and heart rate responses elicited by the increase in core body temperature. In contrast to control animals, however, mesenteric blood flow did not fall in the muscimol-treated rats in response to the elevation in core body temperature. In a separate group, in which muscimol was microinjected into regions outside the PVN, elevating core body temperature elicited the normal reduction in mesenteric blood flow. The results suggest that the PVN may play a key role in the reflex decrease in mesenteric blood flow elicited by hyperthermia.     

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.     

Renal effects of angiotensin II receptor subtype 1 and 2-selective ligands injected into the paraventricular nucleus of conscious rats.

We determined the effects of losartan and CGP42112A (selective ligands of the AT1 and AT2 angiotensin receptors, respectively) and salarasin (a relatively nonselective angiotensin receptor antagonist) on urinary volume and urinary sodium and potassium excretion induced by administration of angiotensin II (ANG II) into the paraventricular nucleus (PVN) of conscious rats. Both the AT1 and AT2 ligands and salarasin administered in the presence of ANG II elicited a concentration-dependent inhibition of urine excretion, but losartan inhibited only 75% of this response. The IC50 for salarasin, CGP42112A, and losartan was 0.01, 0.05, and 6 nM, respectively. Previous treatment with saralasin, CGP42112A and losartan competitively antagonized the natriuretic responses to PVN administration of ANG II, and the IC50 values were 0.09, 0.48, and 10 nM, respectively. The maximum response to losartan was 65% of that obtained with saralasin. Pretreatment with saralasin, losartan, and CGP42112A injected into the PVN caused shifts to the right of the concentration-response curves, but the losartan concentrations were disproportionately greater compared with salarasin or CGP42112A. The IC50 values were 0.06, 0.5, and 7.0 for salarasin, CGP42112A, and losartan, respectively. These results suggest that both AT1 and AT2 receptor subtypes in the PVN are involved in ANG II-related urine, sodium, and potassium excretion, and that the inhibitory responses to AT2 blockade are predominant.     

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.     

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.     

ANG II in the paraventricular nucleus potentiates the cardiac sympathetic afferent reflex in rats with heart failure.

Chronic heart failure (CHF) is characterized by sympathoexcitation, and the cardiac sympathetic afferent reflex (CSAR) is a sympathoexcitatory reflex. Our previous studies have shown that the CSAR was enhanced in CHF. In addition, central angiotensin II (ANG II) is an important modulator of this reflex. This study was performed to determine whether the CSAR evoked by stimulation of cardiac sympathetic afferent nerves (CSAN) in rats with coronary ligation-induced CHF is enhanced by ANG II in the paraventricular nucleus (PVN). Under alpha-chloralose and urethane anesthesia, renal sympathetic nerve activity (RSNA) was recorded. The RSNA responses to electrical stimulation (5, 10, 20, and 30 Hz) of the CSAN were evaluated. Bilateral microinjection of the AT1-receptor antagonist losartan (50 nmol) into the PVN had no significant effects in the sham group, but it abolished the enhanced RSNA response to stimulation in the CHF group. Unilateral microinjection of three doses of ANG II (0.03, 0.3, and 3 nmol) into the PVN resulted in dose-related increases in the RSNA responses to stimulation. Although ANG II also potentiated the RSNA response to electrical stimulation in sham rats, the RSNA responses to stimulation after ANG II into the PVN in rats with CHF were much greater than in sham rats. The effects of ANG II were prevented by pretreatment with losartan into the PVN in CHF rats. These results suggest that the central gain of the CSAR is enhanced in rats with coronary ligation-induced CHF and that ANG II in the PVN augments the CSAR evoked by CSAN, which is mediated by the central angiotensin AT1 receptors in rats with CHF.     

Microinjection of ANG II into paraventricular nucleus enhances cardiac sympathetic afferent reflex in rats

The aims of present study were to determine whether angiotensin II (ANG II) in the paraventricular nucleus (PVN) is involved in the central integration of the cardiac sympathetic afferent reflex and whether this effect is mediated by the ANG type 1 (AT(1)) receptor. While the animals were under alpha-chloralose and urethane anesthesia, mean arterial pressure, heart rate, and renal sympathetic nerve activity (RSNA) were recorded in sinoaortic-denervated and cervical-vagotomized rats. A cannula was inserted into the left PVN for microinjection of ANG II. The cardiac sympathetic afferent reflex was tested by electrical stimulation (5, 10, 20, and 30 Hz in 10 V and 1 ms) of the afferent cardiac sympathetic nerves or epicardial application of bradykinin (BK) (0.04 and 0.4 microg in 2 microl). Microinjection of ANG II (0.03, 0.3, and 3 nmol) into the PVN resulted in dose-related increases in the RSNA responses to electrical stimulation. The percent change of RSNA response to 20- and 30-Hz stimulation increased significantly at the highest dose of ANG II (3 nmol). The effects of ANG II were prevented by pretreatment with losartan (50 nmol) into the PVN. Microinjection of ANG II (0.3 nmol) into the PVN significantly enhanced the RSNA responses to epicardial application of BK, which was abolished by pretreatment with losartan (50 nmol) into the PVN. These results suggest that exogenous ANG II in the PVN augments the cardiac sympathetic afferent reflex evoked by both electrical stimulation of cardiac sympathetic afferent nerves and epicardial application of BK. These central effects of ANG II are mediated by AT(1) receptors.     

Angiotensin depolarizes parvocellular neurons in paraventricular nucleus through modulation of putative nonselective cationic and potassium conductances

Neurosecretory parvocellular neurons in the hypothalamic paraventricular nucleus (PVN) exercise considerable influence over the adenohypophysis and thus play a critical role in neuroendocrine regulation. ANG II has been demonstrated to act as a neurotransmitter in PVN, exerting significant impact on neuronal excitability and also influencing corticotrophin-releasing hormone secretion from the median eminence and, therefore, release of ACTH from the pituitary. We have used whole cell patch-clamp techniques in hypothalamic slices to examine the effects of ANG II on the excitability of neurosecretory parvocellular neurons. ANG II application resulted in a dose-dependent depolarization of neurosecretory neurons, a response that was maintained in tetrodotoxin (TTX), suggesting a direct mechanism of action. The depolarizing actions of this peptide were abolished by losartan, demonstrating these effects are AT(1) receptor mediated. Voltage-clamp analysis using slow voltage ramps revealed that ANG II activates a voltage-independent conductance with a reversal potential of -37.8 +/- 3.8 mV, suggesting ANG II effects on a nonselective cationic current. Further, a sustained potassium current characteristic of I(K) was significantly reduced (29.1 +/- 4.7%) by ANG II. These studies identify multiple postsynaptic modulatory sites through which ANG II can influence the excitability of neurosecretory parvocellular PVN neurons and, as a consequence of such actions, control hormonal secretion from the anterior pituitary.     

p22phox in the macula densa regulates single nephron GFR during angiotensin II infusion in rats

Angiotensin II (ANG II) infusion increases renal superoxide (O(2)(-)) and enhances renal vasoconstriction via macula densa (MD) regulation of tubuloglomerular feedback, but the mechanism is unclear. We targeted the p22(phox) subunit of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) with small-interfering RNA (siRNA) to reduce NADPH oxidase activity and blood pressure response to ANG II in rats. We compared single nephron glomerular filtration rate (SNGFR) in samples collected from the proximal tubule (PT), which interrupts delivery to the MD, and from the distal tubule (DT), which maintains delivery to the MD, to assess MD regulation of GFR. SNGFR was measured in control and ANG II-infused rats (200 ng.kg(-1).min(-1) for 7 days) 2 days after intravenous injection of vehicle or siRNA directed to p22(phox) to test the hypothesis that p22(phox) mediates MD regulation of SNGFR during ANG II. The regulation of SNGFR by MD, determined by PT SNGFR-DT SNGFR, was not altered by siRNA in control rats (control + vehicle, 13 +/- 1, n = 8; control + siRNA, 12 +/- 2 nl/min, n = 8; not significant) but was reduced by siRNA in ANG II-treated rats (ANG II + vehicle, 13 +/- 2, n = 7; ANG II + siRNA, 7 +/- 1 nl/min, n = 8; P < 0.05). We conclude that p22(phox) and NADPH oxidase regulate the SNGFR during ANG II infusion via MD-dependent mechanisms.     

Angiotensin II attenuates myocardial interstitial acetylcholine release in response to vagal stimulation

Although ANG II exerts a variety of effects on the cardiovascular system, its effects on the peripheral parasympathetic neurotransmission have only been evaluated by changes in heart rate (an effect on the sinus node). To elucidate the effect of ANG II on the parasympathetic neurotransmission in the left ventricle, we measured myocardial interstitial ACh release in response to vagal stimulation (1 ms, 10 V, 20 Hz) using cardiac microdialysis in anesthetized cats. In a control group (n = 6), vagal stimulation increased the ACh level from 0.85 +/- 0.03 to 10.7 +/- 1.0 (SE) nM. Intravenous administration of ANG II at 10 microg x kg(-1) x h(-1) suppressed the stimulation-induced ACh release to 7.5 +/- 0.6 nM (P < 0.01). In a group with pretreatment of intravenous ANG II receptor subtype 1 (AT(1) receptor) blocker losartan (10 mg/kg, n = 6), ANG II was unable to inhibit the stimulation-induced ACh release (8.6 +/- 1.5 vs. 8.4 +/- 1.7 nM). In contrast, in a group with local administration of losartan (10 mM, n = 6) through the dialysis probe, ANG II inhibited the stimulation-induced ACh release (8.0 +/- 0.8 vs. 5.8 +/- 1.0 nM, P < 0.05). In conclusion, intravenous ANG II significantly inhibited the parasympathetic neurotransmission through AT(1) receptors. The failure of local losartan administration to nullify the inhibitory effect of ANG II on the stimulation-induced ACh release indicates that the site of this inhibitory action is likely at parasympathetic ganglia rather than at postganglionic vagal nerve terminals.     

Role of angiotensin in body fluid homeostasis of mice: effect of losartan on water and NaCl intakes

It is known that mice injected peripherally with ANG II do not show a drinking response but that cFos immunoreactivity (ir) is induced in brain regions similar to those in rats. We now show in Crl:CD1(ICR) mice that peripheral injection of the ANG II type 1 receptor antagonist losartan was sufficient to prevent this induction of Fos-ir in the subfornical organ (SFO). Injection of ANG II into the lateral cerebral ventricle produced a robust water intake in mice and induced Fos-ir in SFO, as well as in median preoptic (MnPO) and paraventricular (PVN) nuclei. Peripheral injection of losartan blocked this drinking response and prevented the induction of Fos-ir in each of these brain regions. Hypovolemia produced by polyethylene glycol (PEG) produced a robust water intake but no evidence of sodium appetite, and it induced Fos-ir in SFO, MnPO, and PVN. Peripheral injection of losartan did not affect this drinking response. Fos-ir induced by PEG in SFO and MnPO was reduced by treatment with losartan, while that induced in the PVN was further increased by losartan. Sodium depletion with furosemide and low-sodium diet produced a strong sodium appetite and induced Fos-ir in SFO and MnPO. Treatment with losartan completely blocked the sodium appetite, as well as the induction of Fos-ir in these brain regions. These data indicate that endogenous production of ANG II and action at forebrain receptors is critically involved in depletion-related sodium appetite in mice. The absence of an effect of losartan on PEG-induced drinking suggests the critical involvement of other factor(s) such as arterial or venous baroreceptor input, and we discuss how this factor could also explain why peripheral ANG II is not dipsogenic in mice.     

Competitive antagonism of pressor responses to angiotensin II and angiotensin III by the angiotensin II-1 receptor ligand losartan.

Losartan (DuP 753) and PD123177 are nonpeptide angiotensin (ANG) receptor ligands for subtypes of ANG II receptors ANG II-1 and ANG II-2, respectively. We examined the effects of losartan and PD123177 on dose - mean arterial pressure (MAP) response curves for ANG II and ANG III in eight groups (n = 6 each) of conscious rats. Saline (0.9% NaCl), losartan (1 x 10(-6) and 9 x 10(-6) mol/kg), and PD123177 (2 x 10(-5) mol/kg) were i.v. bolus injected 15 min before the construction of ANG II dose - response curves in groups I, II, III, and IV, respectively. Groups V-VIII were treated similarly to I-IV except that ANG III was given in place of ANG II. Losartan dose dependently shifted the dose-response curves of ANG II and ANG III to the right with similar dissociation constants (-log KI of 6.6 +/- 0.7 and 6.6 +/- 0.1 mol/kg, respectively) and no change in the maxima. PD123177 affected neither maximum MAP nor ED50 values for ANG II or ANG III. Our results show that losartan but not PD123177 is a competitive antagonist of the MAP effects of ANG II and ANG III.     

AT1 receptor-mediated augmentation of angiotensinogen, oxidative stress, and inflammation in ANG II-salt hypertension

Augmentation of intrarenal angiotensinogen (AGT) synthesis, secretion, and excretion is associated with the development of hypertension, renal oxidative stress, and tissue injury during ANG II-dependent hypertension. High salt (HS) exacerbates hypertension and kidney injury, but the mechanisms remain unclear. In this study, we determined the consequences of HS intake alone compared with chronic ANG II infusion and combined HS plus ANG II on the stimulation of urinary AGT (uAGT), renal oxidative stress, and renal injury markers. Sprague-Dawley rats were subjected to 1) a normal-salt diet [NS, n = 5]; 2) HS diet [8% NaCl, n = 5]; 3) ANG II infusion in NS rats [ANG II 80 ng/min, n = 5]; 4) ANG II infusion in HS rats [ANG II+HS, n = 5]; and 5) ANG II infusion in HS rats treated with ANG II type 1 receptor blocker (ARB) [ANG II+HS+ARB, n = 5] for 14 days. Rats fed a HS diet alone did not show changes in systolic blood pressure (SBP), proteinuria, cell proliferation, or uAGT excretion although they did exhibit mesangial expansion, collagen deposition, and had increased NADPH oxidase activity accompanied by increased peroxynitrite formation in the kidneys. Compared with ANG II rats, the combination of ANG II infusion and a HS diet led to exacerbation in SBP (175 ± 10 vs. 221 ± 8 mmHg; P < 0.05), proteinuria (46 ± 7 vs. 127 ± 7 mg/day; P < 0.05), and uAGT (1,109 ± 70 vs.. 7,200 ± 614 ng/day; P < 0.05) associated with greater collagen deposition, mesangial expansion, interstitial cell proliferation, and macrophage infiltration. In both ANG II groups, the O(2)(-) levels were increased due to increased NADPH oxidase activity without concomitant increases in peroxynitrite formation. The responses in ANG II rats were prevented or ameliorated by ARB treatment. The results indicate that HS independently stimulates ROS formation, which may synergize with the effect of ANG II to limit peroxynitrite formation, leading to exacerbation of uAGT and greater injury during ANG II salt hypertension.     

Paraventricular noradrenergic systems participate in angiotensin II-induced drinking

The present study was designed to examine the role of noradrenergic systems in the hypothalamic paraventricular nucleus (PVN) in the drinking response induced by microinjection of angiotensin II (ANG II) into the subfornical organ (SFO) in the awake rat. Intracerebral microdialysis techniques were utilized to quantify the extracellular concentration of noradrenaline (NA) in the region of the PVN. Injections of ANG II (10⁻⁶ M, 0.2 μl) into the SFO significantly increased NA release in the PVN area. The increase in the NA concentration caused by the ANG II injection was significantly attenuated by water ingestion. In urethane-anesthetized rats, injections of ANG II into the SFO elicited an elevation in mean arterial pressure (MAP). On the other hand, intravenous injections of the -agonist metaraminol (5 μg) slightly decreased the release of NA in the PVN area that accompanied an elevation in MAP. These results show that the noradrenergic system in the PVN area may be involved in the dipsogenic response induced by ANG II acting at the SFO.     

Effects of central infusion of ANG II and losartan on the cardiac baroreflex in rabbits

The effect of chronic activation or inhibition of central ANG II receptors on cardiac baroreflex function in conscious normotensive rabbits was examined. Animals received a fourth ventricular (4V) infusion of ANG II (30 and 100 ng/h), losartan (3 and 30 microg/h), or Ringer solution (2 microl/h) for 2 wk. After 1 and 2 wk, ANG II (100 ng/h) decreased cardiac baroreflex gain by 20 and 37%, respectively (P = 0.015), whereas losartan (30 microg/h) increased baroreflex gain by 24 and 58%, respectively (P = 0.02). Within 1 wk of the end of the infusions, cardiac baroreflex gain had returned to control. Ringer solution or the lower doses of ANG II or losartan did not modify the cardiac baroreflex function. Blood pressure and heart rate were not altered by any treatment, nor was their variability affected. These data demonstrate a novel long-term modulation of cardiac baroreflexes by endogenous ANG II that is independent of blood pressure level.     

ACE and non-ACE mediated effect of angiotensin I on intracellular calcium mobilization in rat glomerular arterioles

Because renin and angiotensin I (ANG I) level are high in the renal circulation, the conversion of ANG I is a critical step in the regulation of glomerular hemodynamics. We studied this conversion by investigating the effect of ANG I on intracellular Ca(2+) concentration ([Ca(2+)](i)) in rat juxtamedullary glomerular afferent and efferent arterioles (AA and EA, respectively). Two types of EA were considered, thin EA and muscular EA, terminating as peritubular capillaries and vasa rectae, respectively. In all arterioles, ANG I elicited [Ca(2+)](i) elevations. Maximal responses of 171 +/- 28 (AA), 183 +/- 7 (muscular EA), and 78 +/- 11 nM (thin EA) (n = 6), similar to those obtained with ANG II, were observed with 100 nM ANG I. The EC(50) values were 20 times higher for ANG I than for ANG II in AA (10.2 vs. 0.5) and muscular EA (6.8 vs. 0.4 nM) and 150 times higher in thin EA (15.2 vs. 0.1 nM). ANG I effect was blocked by losartan, indicating that AT(1) receptors were involved. The ANG-converting enyzme (ACE) inhibitor lisinopril inhibited the maximal response to ANG I in AA and muscular EA by 75 +/- 9% (n = 13) and 70 +/- 7% (n = 13), respectively, but had no effect in thin EA (n = 14). The serine protease inhibitor aprotinin, the chymase inhibitor chymostatin, and the cysteine protease inhibitors E64 and leupeptin had no effect on ANG I action. These data show that ANG I effects are mainly mediated by ACE in AA and muscular EA but not in thin EA. The lisinopril-insensitive response may be related to conversion by unknown enzyme(s) and/or to activation of AT(1) receptors by ANG I.     

Responses to GABA-A receptor blockade in the hypothalamic PVN are attenuated by local AT1 receptor antagonism

Blockade of GABA-A receptors in the hypothalamic paraventricular nucleus (PVN) has been repeatedly shown to increase arterial blood pressure (ABP), heart rate (HR), and sympathetic nerve activity (SNA), but the mechanism(s) that underlies this response has not been determined. Here, we tested whether full expression of the response requires activation of local ANG II AT1 receptors. ABP, HR, and renal SNA responses to PVN microinjection of bicuculline methobromide (BIC; 0.1 nmol) were recorded before and after microinjection of vehicle (saline); losartan (or L-158809), to block local AT1 receptors; or PD123319, to block AT2 receptors. After PVN microinjection of vehicle or PD123319 (10 nmol), BIC significantly (P < 0.05) increased mean arterial pressure (MAP), HR, and renal SNA. However, PVN microinjection of 2 and 20 nmol of losartan dose dependently reduced responses to PVN-injected BIC, with the 20-nmol dose nearly abolishing MAP (P < 0.005), HR (P < 0.05), and renal SNA (P < 0.005) responses. Another AT1 receptor antagonist, L-158809 (10 nmol), produced similar effects. Neither losartan nor L-158809 altered baseline parameters. Responses to PVN injection of BIC were unchanged by losartan (20 nmol) given intravenously or into the PVN on the opposite side. MAP, HR, and renal SNA responses to PVN microinjection of l-glutamate (10 nmol) were unaffected by PVN injection of losartan (20 nmol), indicating that effects of losartan were not due to nonspecific depression of neuronal excitability. We conclude that pressor, tachycardic, and renal sympathoexcitatory responses to acute blockade of GABA-A receptors in the PVN depend on activation of local AT1 receptors.     

Sympathetic nerve activity and whole body heat stress in humans

We and others have shown that moderate passive whole body heating (i.e., increased internal temperature ～0.7°C) increases muscle (MSNA) and skin sympathetic nerve activity (SSNA). It is unknown, however, if MSNA and/or SSNA continue to increase with more severe passive whole body heating or whether these responses plateau following moderate heating. The aim of this investigation was to test the hypothesis that MSNA and SSNA continue to increase from a moderate to a more severe heat stress. Thirteen subjects, dressed in a water-perfused suit, underwent at least one passive heat stress that increased internal temperature ～1.3°C, while either MSNA (n = 8) or SSNA (n = 8) was continuously recorded. Heat stress significantly increased mean skin temperature (Δ～5°C, P < 0.001), internal temperature (Δ～1.3°C, P < 0.001), mean body temperature (Δ～2.0°C, P < 0.001), heart rate (Δ～40 beats/min, P < 0.001), and cutaneous vascular conductance [Δ～1.1 arbitrary units (AU)/mmHg, P < 0.001]. Mean arterial blood pressure was well maintained (P = 0.52). Relative to baseline, MSNA increased midway through heat stress (Δ core temperature 0.63 ± 0.01°C) when expressed as burst frequency (26 ± 14 to 45 ± 16 bursts/min, P = 0.001), burst incidence (39 ± 13 to 48 ± 14 bursts/100 cardiac cyles, P = 0.03), or total activity (317 ± 170 to 489 ± 150 units/min, P = 0.02) and continued to increase until the end of heat stress (burst frequency: 61 ± 15 bursts/min, P = 0.01; burst incidence: 56 ± 11 bursts/100 cardiac cyles, P = 0.04; total activity: 648 ± 158 units/min, P = 0.01) relative to the mid-heating stage. Similarly, SSNA (total activity) increased midway through the heat stress (normothermia; 1,486 ± 472 to mid heat stress 6,467 ± 5,256 units/min, P = 0.03) and continued to increase until the end of heat stress (11,217 ± 6,684 units/min, P = 0.002 vs. mid-heat stress). These results indicate that both MSNA and SSNA continue to increase as internal temperature is elevated above previously reported values.     

Increased renin excretion is associated with augmented urinary angiotensin II levels in chronic angiotensin II-infused hypertensive rats

Renin expression in principal cells of collecting ducts (CD) is upregulated in angiotensin II (ANG II)-dependent hypertensive rats; however, it remains unclear whether increased CD-derived renin undergoes tubular secretion. Accordingly, urinary levels of renin (uRen), angiotensinogen (uAGT), and ANG II (uANG II) were measured in chronic ANG II-infused Sprague-Dawley rats (80 ng/min for 14 days, n = 10) and sham-operated rats (n = 10). Systolic blood pressure increased in the ANG II rats by day 5 and continued to increase throughout the study (day 13; ANG II: 175 ± 10 vs. sham: 116 ± 2 mmHg; P < 0.05). ANG II infusion increased renal cortical and medullary ANG II levels (cortical ANG II: 606 ± 72 vs. 247 ± 43 fmol/g; P < 0.05; medullary ANG II: 2,066 ± 116 vs. 646 ± 36 fmol/g; P < 0.05). Although plasma renin activity (PRA) was suppressed in the ANG II-infused rats (0.3 ± 0.2 vs. 5.5 ± 1.8 ng ANG I·ml(-1)·h(-1); P < 0.05), renin content in renal medulla was increased (12,605 ± 1,343 vs. 7,956 ± 765 ng ANG I·h(-1)·mg(-1); P < 0.05). Excretion of uAGT and uANG II increased in the ANG II rats [uAGT: 1,107 ± 106 vs. 60 ± 26 ng/day; P < 0.0001; uANG II: 3,813 ± 431 vs. 2,080 ± 361 fmol/day; P < 0.05]. By day 13, despite suppression of PRA, urinary prorenin content increased in ANG II rats [15.7 ± 3 vs. 2.6 ± 1 × 10(-3) enzyme units excreted (EUE)/day, P < 0.01] as was the excretion rate of renin (8.6 ± 2 × 10(-6) EUE/day) compared with sham (2.8 ± 1 × 10(-6) EUE/day; P < 0.05). Urinary renin and prorenin protein levels examined by Western blot were augmented ～10-fold in the ANG II-infused rats. Concomitant AT(1) receptor blockade with candesartan prevented the increase. Thus, in ANG II-dependent hypertensive rats with marked PRA suppression, increased urinary levels of renin and prorenin reflect their augmented secretion by CD cells into the luminal fluid. The greater availability of renin and AGT in the urine reflects the capability for intratubular ANG II formation which stimulates sodium reabsorption in distal nephron segments.     

Reciprocal changes in renal ACE/ANG II and ACE2/ANG 1-7 are associated with enhanced collecting duct renin in Goldblatt hypertensive rats

Alterations in the balance between ANG II/ACE and ANG 1-7/ACE2 in ANG II-dependent hypertension could reduce the generation of ANG 1-7 and contribute further to increased intrarenal ANG II. Upregulation of collecting duct (CD) renin may lead to increased ANG II formation during ANG II-dependent hypertension, thus contributing to this imbalance. We measured ANG I, ANG II, and ANG 1-7 contents, angiotensin-converting enzyme (ACE) and ACE2 gene expression, and renin activity in the renal cortex and medulla in the clipped kidneys (CK) and nonclipped kidneys (NCK) of 2K1C rats. After 3 wk of unilateral renal clipping, systolic blood pressure and plasma renin activity increased in 2K1C rats (n = 11) compared with sham rats (n = 9). Renal medullary angiotensin peptide levels were increased in 2K1C rats [ANG I: (CK = 171 ± 4; NCK = 251 ± 8 vs. sham = 55 ± 3 pg/g protein; P < 0.05); ANG II: (CK = 558 ± 79; NCK = 328 ± 18 vs. sham = 94 ± 7 pg/g protein; P < 0.001)]; and ANG 1-7 levels decreased (CK = 18 ± 2; NCK = 19 ± 2 pg/g vs. sham = 63 ± 10 pg/g; P < 0.001). In renal medullas of both kidneys of 2K1C rats, ACE mRNA levels and activity increased but ACE2 decreased. In further studies, we compared renal ACE and ACE2 mRNA levels and their activities from chronic ANG II-infused (n = 6) and sham-operated rats (n = 5). Although the ACE mRNA levels did not differ between ANG II rats and sham rats, the ANG II rats exhibited greater ACE activity and reduced ACE2 mRNA levels and activity. Renal medullary renin activity was similar in the CK and NCK of 2K1C rats but higher compared with sham. Thus, the differential regulation of ACE and ACE2 along with the upregulation of CD renin in both the CK and NCK in 2K1C hypertensive rats indicates that they are independent of perfusion pressure and contribute to the altered content of intrarenal ANG II and ANG 1-7.