Renal tissue NO and intrarenal haemodynamics during experimental variations of NO content in anaesthetised rats.

Direct renal nitric oxide (NO) measurements were infrequent and no simultaneous measurements of renal cortical and medullary NO and local perfusion. Large-surface NO electrodes were placed in renal cortex and medulla of anaesthetised rats; simultaneously, renal blood flow (RBF, index of cortical perfusion) and medullary laser-Doppler flux (MBF) were determined. NO synthase inhibitors: nonselective (L-NAME) or selective for neuronal NOS (nNOS) (S-methyl-thiocitrulline, SMTC), and NO donor (SNAP), were used to manipulate tissue NO. Baseline tissue NO was significantly higher in medulla (703+/-49 NM) than in cortex (231+/-17 nM). Minimal cortical and medullary NO current measured after maximal L-NAME dose (2.4 mg kg(-1) i.v.) was taken as tissue NO zero kevel. This dose decreased RBF and MBF significantly (-43%). SMTC, 1.2 mg kg(-1) h(-1) i.v., significantly decreased tissue NO by 105+/-32 nM in cortex and 546+/-64 nM in medulla, RBF and MBF decreased 30% and 20%, respectively. Renal artery infusion of SNAP, 0.24 mg kg(-1) min(-1) significantly increased tissue NO by 139+/-18 nM in cortex and 948+/-110 nM in medulla. Since inhibition of nNOS decreased medullary NO by 80% and MBF by 20% only, this isoform has probably minor role in the maintenance of medullary perfusion.     

Renal cortical and medullary blood flow responses to L-NAME and ANG II in wild-type, nNOS null mutant, and eNOS null mutant mice

Experiments in wild-type (WT; C57BL/6J) mice, endothelial nitric oxide synthase null mutant [eNOS(-/-)] mice, and neuronal NOS null mutant [nNOS(-/-)] mice were performed to determine which NOS isoform regulates renal cortical and medullary blood flow under basal conditions and during the infusion of ANG II. Inhibition of NOS with N(omega)-nitro-l-arginine methyl ester (l-NAME; 50 mg/kg iv) in Inactin-anesthetized WT and nNOS(-/-) mice increased arterial blood pressure by 28-31 mmHg and significantly decreased blood flow in the renal cortex (18-24%) and the renal medulla (13-18%). In contrast, blood pressure and renal cortical and medullary blood flow were unaltered after l-NAME administration to eNOS(-/-) mice, indicating that NO derived from eNOS regulates baseline vascular resistance in mice. In subsequent experiments, intravenous ANG II (20 ng x kg(-1) x min(-1)) significantly decreased renal cortical blood flow (by 15-25%) in WT, eNOS(-/-), nNOS(-/-), and WT mice treated with l-NAME. The infusion of ANG II, however, led to a significant increase in medullary blood flow (12-15%) in WT and eNOS(-/-) mice. The increase in medullary blood flow following ANG II infusion was not observed in nNOS(-/-) mice, in WT or eNOS(-/-) mice pretreated with l-NAME, or in WT mice administered the nNOS inhibitor 5-(1-imino-3-butenyl)-l-ornithine (1 mg x kg(-1) x h(-1)). These data demonstrate that NO from eNOS regulates baseline blood flow in the mouse renal cortex and medulla, while NO produced by nNOS mediates an increase in medullary blood flow in response to ANG II.     

L-Arginine uptake affects nitric oxide production and blood flow in the renal medulla

Experiments were performed to determine whether L-arginine transport regulates nitric oxide (NO) production and hemodynamics in the renal medulla. The effects of renal medullary interstitial infusion of cationic amino acids, which compete with L-arginine for cellular uptake, on NO levels and blood flow in the medulla were examined in anesthetized rats. NO concentration in the renal inner medulla, measured with a microdialysis-oxyhemoglobin trapping technique, was significantly decreased by 26-44% and renal medullary blood flow, measured by laser Doppler flowmetry, was significantly reduced by 20-24% during the acute renal medullary interstitial infusion of L-ornithine, L-lysine, and L-homoarginine (1 micromol.kg(-1).min(-1) each; n = 6-8/group). In contrast, intramedullary infusion of L-arginine increased NO concentration and medullary blood flow. Flow cytometry experiments with 4-amino-5-methylamino-2',7'-difluorescein diacetate, a fluorophore reactive to intracellular NO, demonstrated that L-ornithine, L-lysine, and L-homoarginine decreased NO by 54-57% of control, whereas L-arginine increased NO by 21% in freshly isolated inner medullary cells (1 mmol/l each, n > 1,000 cells/experiment). The mRNA for the cationic amino acid transporter-1 was predominantly expressed in the inner medulla, and cationic amino acid transporter-1 protein was localized by immunohistochemistry to the collecting ducts and vasa recta in the inner medulla. These results suggest that L-arginine transport by cationic amino acid transport mechanisms is important in the production of NO and maintenance of blood flow in the renal medulla.     

Renal medullary nitric oxide deficit of Dahl S rats enhances hypertensive actions of angiotensin II

Studies were designed to examine the hypothesis that the renal medulla of Dahl salt-sensitive (Dahl S) rats has a reduced capacity to generate nitric oxide (NO), which diminishes the ability to buffer against the chronic hypertensive effects of small elevations of circulating ANG II. NO synthase (NOS) activity in the outer medulla of Dahl S rats (arginine-citrulline conversion assay) was significantly reduced. This decrease in NOS activity was associated with the downregulation of protein expression of NOS I, NOS II, and NOS III isoforms in this region as determined by Western blot analysis. In anesthetized Dahl S rats, we observed that a low subpressor intravenous infusion of ANG II (5 ng. kg(-1). min(-1)) did not increase the concentration of NO in the renal medulla as measured by a microdialysis with oxyhemoglobin trapping technique. In contrast, ANG II produced a 38% increase in the concentration of NO (87 +/- 8 to 117 +/- 8 nmol/l) in the outer medulla of Brown-Norway (BN) rats. The same intravenous dose of ANG II reduced renal medullary blood flow as determined by laser-Doppler flowmetry in Dahl S, but not in BN rats. A 7-day intravenous ANG II infusion at a dose of 3 ng. kg(-1). min(-1) did not change mean arterial pressure (MAP) in the BN rats but increased MAP in Dahl S rats from 120 +/- 2 to 138 +/- 2 mmHg (P < 0.05). ANG II failed to increase MAP after NO substrate was provided by infusion of L-arginine (300 microg. kg(-1). min(-1)) into the renal medulla of Dahl S rats. Intravenous infusion of L-arginine at the same dose had no effect on the ANG II-induced hypertension. These results indicate that an impaired NO counterregulatory system in the outer medulla of Dahl S rats makes them more susceptible to the hypertensive actions of small elevations of ANG II.     

Mitochondrial function and nitric oxide metabolism are modified by enalapril treatment in rat kidney

The renal and cardiac benefits of renin-angiotensin system (RAS) inhibition in hypertension exceed those attributable to blood pressure reduction, and seem to involve mitochondrial function changes. To investigate whether mitochondrial changes associated with RAS inhibition are related to changes in nitric oxide (NO) metabolism, four groups of male Wistar rats were treated during 2 wk with a RAS inhibitor, enalapril (10 mg x kg(-1) x day(-1); Enal), or a NO synthase (NOS) inhibitor, N(omega)-nitro-L-arginine methyl ester (L-NAME) (1 mg x kg(-1) x day(-1)), or both (Enal+L-NAME), or were untreated (control). Blood pressure and body weight were lower in Enal than in control. Electron transfer through complexes I to III and cytochrome oxidase activity were significantly lower, and uncoupling protein-2 content was significantly higher in kidney mitochondria isolated from Enal than in those from control. All of these changes were prevented by L-NAME cotreatment and were accompanied by a higher production/bioavailability of kidney NO. L-NAME abolished mitochondrial NOS activity but failed to inhibit extra-mitochondrial kidney NOS, underscoring the relevance of mitochondrial NO in those effects of enalapril that were suppressed by L-NAME cotreatment. In Enal, kidney mitochondria H(2)O(2) production rate and MnSOD activity were significantly lower than in control, and these effects were not prevented by L-NAME cotreatment. These findings may clarify the role of NO in the interactions between RAS and mitochondrial metabolism and can help to unravel the mechanisms involved in renal protection by RAS inhibitors.     

N-acetyl-L-cysteine improves renal medullary hypoperfusion in acute renal failure

This study evaluated the effects of N-acetyl-L-cysteine (NAC), a free radical scavenger, and N(omega)-nitro-L-arginine methyl ester (L-NAME), a nitric oxide (NO) synthesis inhibitor, on the changes in renal function, intrarenal blood flow distribution (laser-Doppler flowmetry), and plasma peroxynitrite levels during the acute renal failure (ARF) produced by inferior vena cava occlusion (IVCO; 45 min) in anesthetized rats. Renal blood flow fell on reperfusion (whole kidney by -45.7%; cortex -58.7%, outer medulla -62.8%, and papilla -47.7%); glomerular filtration rate (GRF) also decreased (-68.6%), whereas fractional sodium excretion (FE(Na%)) and peroxynitrite and NO/NO plasma levels increased (189.5, 46.5, and 390%, respectively) after ischemia. Pretreatment with L-NAME (10 microg. kg(-1). min(-1)) aggravated the fall in renal blood flow seen during reperfusion (-60%). Pretreatment with NAC (150 mg/kg bolus + 715 microg. kg(-1). min(-1) iv) partially prevented those changes in renal function (GFR only fell by -29.2%, and FE(Na%) increased 119.4%) and laser-Doppler blood flow, especially in the outer medulla, where blood flow recovered to near control levels during reperfusion. These beneficial effects seen in rats given NAC seem to be dependent on the presence of NO, because they were abolished in rats pretreated with L-NAME. Also, the antioxidant effects of NAC prevented the increase in plasma peroxynitrite after ischemia. In conclusion, NAC ameliorates the renal failure and the outer medullary vasoconstriction induced by ICVO, effects that seem to be dependent on the presence of NO and the scavenging of peroxynitrite.     

肾髓质诱导型一氧化氮合酶在动脉血压调控中的作用

本文通过慢性血液动力学实验,观察了肾髓质局部输入诱导型一氧化酶（ｉＮＯＳ）抑制剂ＡＧ（ａｍｉｎｏｇｕａｎｉｄｉｎｅ）对Ｄａｈｌ盐敏感大鼠（ＤＳ）、Ｄａｈｌ盐抵抗大鼠（ＤＲ）及ＳＤ（Ｓｐｒａｇｕｅ Ｄａｗｌｅｙ）大鼠动脉血压的影响,并测定了一氧化氮（ＮＯ）代谢终产物ＮＯ２及ＮＯ３含量（ＵＮＯＸ）、ｉＮＯＳ活性、肾功能以及血浆肾素活性（ＰＲＡ）。结果表明：ＡＧ能明显放大高盐（８％）引起的ＤＳ及ＳＤ大鼠     

Interactions between angiotensin II and nitric oxide during exercise in normal and heart failure rats.

We hypothesized that nitric oxide (NO) opposes ANG II-induced increases in arterial pressure and reductions in renal, splanchnic, and skeletal muscle vascular conductance during dynamic exercise in normal and heart failure rats. Regional blood flow and vascular conductance were measured during treadmill running before (unblocked exercise) and after 1) ANG II AT(1)-receptor blockade (losartan, 20 mg/kg ia), 2) NO synthase (NOS) inhibition [N(G)-nitro-L-arginine methyl ester (L-NAME); 10 mg/kg ia], or 3) ANG II AT(1)-receptor blockade + NOS inhibition (combined blockade). Renal conductance during unblocked exercise (4.79 +/- 0.31 ml x 100 g(-1) x min(-1) x mmHg(-1)) was increased after ANG II AT(1)-receptor blockade (6.53 +/- 0.51 ml x 100 g(-1) x min(-1) x mmHg(-1)) and decreased by NOS inhibition (2.12 +/- 0.20 ml x 100 g(-1) x min(-1) x mmHg(-1)) and combined inhibition (3.96 +/- 0.57 ml x 100 g(-1) x min(-1) x mmHg(-1); all P < 0.05 vs. unblocked). In heart failure rats, renal conductance during unblocked exercise (5.50 +/- 0.66 ml x 100 g(-1) x min(-1) x mmHg(-1)) was increased by ANG II AT(1)-receptor blockade (8.48 +/- 0.83 ml x 100 g(-1) x min(-1) x mmHg(-1)) and decreased by NOS inhibition (2.68 +/- 0.22 ml x 100 g(-1) x min(-1) x mmHg(-1); both P < 0.05 vs. unblocked), but it was unaltered during combined inhibition (4.65 +/- 0.51 ml x 100 g(-1) x min(-1) x mmHg(-1)). Because our findings during combined blockade could be predicted from the independent actions of NO and ANG II, no interaction was apparent between these two substances in control or heart failure animals. In skeletal muscle, L-NAME-induced reductions in conductance, compared with unblocked exercise (P < 0.05), were abolished during combined inhibition in heart failure but not in control rats. These observations suggest that ANG II causes vasoconstriction in skeletal muscle that is masked by NO-evoked dilation in animals with heart failure. Because reductions in vascular conductance between unblocked exercise and combined inhibition were less than would be predicted from the independent actions of NO and ANG II, an interaction exists between these two substances in heart failure rats. L-NAME-induced increases in arterial pressure during treadmill running were attenuated (P < 0.05) similarly in both groups by combined inhibition. These findings indicate that NO opposes ANG II-induced increases in arterial pressure and in renal and skeletal muscle resistance during dynamic exercise.     

Adenosine A2A receptor activation attenuates tubuloglomerular feedback responses by stimulation of endothelial nitric oxide synthase

Adenosine A(2) receptors have been suggested to modulate tubuloglomerular feedback (TGF) responses by counteracting adenosine A(1) receptor-mediated vasoconstriction, but the mechanisms are unclear. We tested the hypothesis that A(2A) receptor activation blunts TGF by release of nitric oxide in the juxtaglomerular apparatus (JGA). Maximal TGF responses were measured in male Sprague-Dawley rats as changes in proximal stop-flow pressure (ΔP(SF)) in response to increased perfusion of the loop of Henle (0 to 40 nl/min) with artificial tubular fluid (ATF). The maximal TGF response was studied after 5 min intratubular perfusion (10 nl/min) with ATF or ATF + A(2A) receptor agonist (CGS-21680; 10(-7) mol/l). The interaction with nitric oxide synthase (NOS) isoforms was tested by perfusion with a nonselective NOS inhibitor [N(ω)-nitro-L-arginine methyl ester hydrochloride (L-NAME); 10(-3) mol/l] or a selective neuronal NOS (nNOS) inhibitor [N(ω)-propyl-L-arginine (L-NPA); 10(-6) mol/l] alone, and with the A(2A) agonist. Blood pressure, urine flow, and P(SF) at 0 nl/min were similar among the groups. The maximal TGF response (ΔP(SF)) with ATF alone (12.3 ± 0.6 mmHg) was attenuated by selective A(2A) stimulation (9.5 ± 0.4 mmHg). L-NAME enhanced maximal TGF responses (18.9 ± 0.4 mmHg) significantly more than L-NPA (15.2 ± 0.7 mmHg). Stimulation of A(2A) receptors did not influence maximal TGF response during nonselective NOS inhibition (19.0 ± 0.4) but attenuated responses during nNOS inhibition (10.3 ± 0.4 mmHg). In conclusion, adenosine A(2A) receptor activation attenuated TGF responses by stimulation of endothelial NOS (eNOS), presumably in the afferent arteriole. Moreover, NO derived from both eNOS and nNOS in the JGA may blunt TGF responses.     

Blood flow of the right and left submandibular gland during unilateral carotid artery occlusion in rat: role of nitric oxide

The aim of the present study was to investigate the effect of unilateral carotid artery occlusion on the blood flow of submandibular gland in anesthetized rats and identify the role of nitric oxide (NO) in blood flow changes after the artery occlusion. L-NAME (N omega-nitro-L-arginine-methyl-ester; 10 mg/kg/day, per os) dissolved in tap water was used to block nitric oxide synthase. Glandular blood flow was measured using Sapirstein's indicator (86Rb) distribution technique. In the control animals the blood flow of left (ligated side) submandibular gland was lower than in the right (unligated side) one (right: 76.4+/-15.4 ml/min/100 g, 64.1+/-13.4 ml/min/100 g, p<0.01). The blood flow of submandibular glands decreased in NOS blocked group versus control. The vascular resistance after L-NAME treatment was elevated (control: 11+/-2.3 R/kg, L-NAME: 17.5+/-4.1 R/kg, p<0.001). In L-NAME group the difference between blood flow value of the left and right submandibular gland was significantly lower than in the control group (control: -16%, NAME: -8%, p<0.01). Conclusion: The maintenance of the blood flow in the left submandibular gland during ligation of the left common carotid artery could be due to the good vascular anastomotic system at these regions and adaptation of the submandibular vessels to the decreased perfusion pressure. Nitric oxide may have a role in the regulation of blood flow tinder this condition.     

Effect of desflurane-induced preconditioning following ischemia-reperfusion on nitric oxide release in rabbits

Nitric oxide (NO) is the mediator of ischemic preconditioning against myocardial infarction. Desflurane produces anesthetic preconditioning to protect the myocardium against infarction. In the model of myocardial ischemia-reperfusion injury in rabbits, we evaluated desflurane-induced ischemic preconditioning and studied its mechanism of NO synthesis. Thirty-two male adult New Zealand white rabbits were anesthetized with intravenous (IV) 30 mg/kg pentobarbital followed by 5 mg/kg/hr infusion. All rabbits were subjected to 30 minutes (min) long lasting left anterior descending coronary artery (LAD) occlusion and three hours (hr) of subsequent reperfusion. Before LAD occlusion, the rabbits were randomly allocated into four groups for preconditioning treatment (eight for each group). The control group did not receive any preconditioning treatment. The desflurane group received inhaled desflurane 1.0 MAC (minimal end-tidal alveolar concentration) for 30 min that was followed by a 15 min washout period. The L-NAME-desflurane group received L-NAME (NG-nitro-L-arginine methyl ester; non-selective Nitric Oxide Synthetase (NOS) inhibitor) 1 mg/kg IV 15 min before 1.0 MAC inhaled desflurane for 30 min. The L-NAME group received L-NAME 1 mg/kg IV. Infarct volume, ventricular arrhythmia, plasma lactate dehydrogenase (LDH), creatine kinase (CK) activity and myocardial perfusion were recorded simultaneously. We have found that hemodynamic values of the coronary blood flow before, during, and after LAD occlusion were not significantly different among these four groups. For the myocardial ischemia-reperfusion injury animals, the infarction size (mean +/- SEM) in the desflurane group was significantly reduced to 18 +/- 3% in the area at risk as compared with 42 +/- 7% in the control group, 35 +/- 6 in the L-NAME group, and 34 +/- 4% in the L-NAME-desflurane group. The plasma LDH, CK levels, and duration of ventricular arrhythmia were also significantly decreased in the desflurane group during ischemia-reperfusion injury. Our results indicate that desflurane is an anesthetic preconditioning agent, which could protect the myocardium against the ischemia-reperfusion injury. This beneficial effect of desflurane on the ischemic preconditioning is probably through NO release since L-NAME abrogates the desflurane preconditioning effect.     

Role of nitric oxide in the vascular effects of local warming of the skin in humans.

Local warming of skin induces vasodilation by unknown mechanisms. To test whether nitric oxide (NO) is involved, we examined effects of NO synthase (NOS) inhibition with NG-nitro-L-arginine methyl ester (L-NAME) on vasodilation induced by local warming of skin in six subjects. Two adjacent sites on the forearm were instrumented with intradermal microdialysis probes for delivery of L-NAME and sodium nitroprusside. Skin blood flow was monitored by laser-Doppler flowmetry (LDF) at microdialysis sites. Local temperature (Tloc) of the skin at both sites was controlled with special LDF probe holders. Mean arterial pressure (MAP; Finapres) was measured and cutaneous vascular conductance calculated (CVC = LDF/MAP = mV/mmHg). Data collection began with a control period (Tloc at both sites = 34 degrees C). One site was then warmed to 41 degrees C while the second was maintained at 34 degrees C. Local warming increased CVC from 1.44 +/- 0.41 to 4.28 +/- 0.60 mV/mmHg (P < 0.05). Subsequent L-NAME administration reduced CVC to 2.28 +/- 0.47 mV/mmHg (P < 0.05 vs. heating), despite the continued elevation of Tloc. At a Tloc of 34 degrees C, L-NAME reduced CVC from 1.17 +/- 0.23 to 0.75 +/- 0.11 mV/mmHg (P < 0.05). Administration of sodium nitroprusside increased CVC to levels no different from those induced by local warming. Thus NOS inhibition attenuated, and sodium nitroprusside restored, the cutaneous vasodilation induced by elevation of Tloc; therefore, the mechanism of cutaneous vasodilation by local warming requires NOS generation of NO.     

Perivascular nitric oxide and superoxide in neonatal cerebral hypoxia-ischemia

Decreased cerebral blood flow (CBF) has been observed following the resuscitation from neonatal hypoxic-ischemic injury, but its mechanism is not known. We address the hypothesis that reduced CBF is due to a change in nitric oxide (NO) and superoxide anion O(2)(-) balance secondary to endothelial NO synthase (eNOS) uncoupling with vascular injury. Wistar rats (7 day old) were subjected to cerebral hypoxia-ischemia by unilateral carotid occlusion under isoflurane anesthesia followed by hypoxia with hyperoxic or normoxic resuscitation. Expired CO(2) was determined during the period of hyperoxic or normoxic resuscitation. Laser-Doppler flowmetry was used with isoflurane anesthesia to monitor CBF, and cerebral perivascular NO and O(2)(-) were determined using fluorescent dyes with fluorescence microscopy. The effect of tetrahydrobiopterin supplementation on each of these measurements and the effect of apocynin and N(omega)-nitro-L-arginine methyl ester (L-NAME) administration on NO and O(2)(-) were determined. As a result, CBF in the ischemic cortex declined following the onset of resuscitation with 100% O(2) (hyperoxic resuscitation) but not room air (normoxic resuscitation). Expired CO(2) was decreased at the onset of resuscitation, but recovery was the same in normoxic and hyperoxic resuscitated groups. Perivascular NO-induced fluorescence intensity declined, and O(2)(-)-induced fluorescence increased in the ischemic cortex after hyperoxic resuscitation up to 24 h postischemia. L-NAME treatment reduced O(2)(-) relative to the nonischemic cortex. Apocynin treatment increased NO and reduced O(2)(-) relative to the nonischemic cortex. The administration of tetrahydrobiopterin following the injury increased perivascular NO, reduced perivascular O(2)(-), and increased CBF during hyperoxic resuscitation. These results demonstrate that reduced CBF follows hyperoxic resuscitation but not normoxic resuscitation after neonatal hypoxic-ischemic injury, accompanied by a reduction in perivascular production of NO and an increase in O(2)(-). The finding that tetrahydrobiopterin, apocynin, and L-NAME normalized radical production suggests that the uncoupling of perivascular NOS, probably eNOS, due to acquired relative tetrahydrobiopterin deficiency occurs after neonatal hypoxic-ischemic brain injury. It appears that both NOS uncoupling and the activation of NADPH oxidase participate in the changes of reactive oxygen concentrations seen in cerebral hypoxic-ischemic injury.     

Sex differences in renal injury and nitric oxide production in renal wrap hypertension

To investigate the faster rate of renal disease progression in men compared with women, we addressed the following questions in the renal wrap (RW) model of hypertension: 1) Do sex differences exist in RW-induced renal injury, which are independent of sex differences in blood pressure? 2) Do sex differences in nitric oxide (NO) production exist in RW hypertension? Male (M) and female (F) rats underwent sham-operated (M-Sham, n = 7; F-Sham, n = 10) or RW (M-RW, n = 13; F-RW, n = 14) surgery for 9 wk. Markers of renal injury, including the glomerulosclerosis index (F-RW, 0.70 +/- 0.1 vs. M-RW, 2.2 +/- 0.6; P < 0.05), mean glomerular volume (F-RW, 1.05 +/- 0.050 x 10(6) vs. M-RW, 1.78 +/- 0.15 x 10(6) microm(3); P < 0.001), and proteinuria (F-RW, 68.7 +/- 15 vs. M-RW, 124 +/- 7.7 mg/day; P < 0.001) were greater in RW males compared with RW females. Endothelial NO synthase protein expression was elevated in the renal cortex (3.2-fold) and medulla (2.2-fold) 9 wk after RW in males, whereas no differences were observed in females. Neuronal NO synthase protein expression was unchanged in the renal cortex in males and in both the renal cortex and medulla in females, whereas in the male medulla, neuronal NOS was decreased by 57%. These data suggest the degree of renal injury is greater in male compared with female rats in RW hypertension despite similar degrees of hypertension and renal function and may involve sex differences in renal NO metabolism.     

Influence of renal denervation on renal effects of acute nitric oxide and ETA/ETB receptor inhibition in conscious normotensive rats.

The role of renal nerves in the effects of concomitant NO synthase and non-selective ET(A/)ET(B) receptor inhibition on renal function was investigated in conscious normotensive Wistar rats. NO synthase inhibition alone (10 mg/kg b. w. i.v. L-NAME) in sham-operated rats with intact renal nerves induced an increase in systolic, diastolic and mean arterial pressure, urine flow rate, sodium, chloride and calcium excretion (p<0.05). The effect of L-NAME was markedly reduced by bosentan (10 mg/kg b.w. i.v.) and the values of urine flow rate, sodium, chloride and calcium excretions returned to control level (p<0.05). L-NAME administration one week after a bilateral renal denervation increased blood pressure to a similar extent as in sham-operated rats but decreased urine flow rate (p<0.05) and did not change electrolyte excretion. ET(A/)ET(B) receptor inhibition with bosentan during NO synthase inhibition in the renal denervated rats did not produce changes in urine flow rate or electrolyte excretion. NO synthase inhibition as well as concurrent NO synthase and ET(A/)ET(B) receptor inhibition did not change clearance of inulin or paraaminohippuric acid in sham-operated or renal denervated rats. These results indicate that renal sympathetic nerves play an important modulatory role in NO and endothelin induced effects on renal excretory function.     

Role of nitric oxide in the nicotine-induced pituitary-adrenocortical response.

Nitric oxide (NO) is a major signaling molecule and biological mediator of the hypothalamic-pituitary-adrenal (HPA) axis. We investigated the role of NO formed by endothelial (e), neuronal (n) and inducible (i) nitric oxide synthase (NOS) in the stimulatory effect of nicotine on the HPA axis in rats under basal conditions. Also possible interaction of NOS systems with endogenous prostaglandins (PG) in that stimulation was assessed. NOS and cyclooxygenase inhibitors were administered i.p. 15 min prior to nicotine (2, 5 mg/kg i.p.). Plasma ACTH and serum corticosterone levels were measured 1 h after nicotine injection. NOS blockers given alone did not markedly affect the resting ACTH and corticosterone levels. L-NAME (2-10 mg/kg), a broad spectrum NOS inhibitor considerably and dose dependently enhanced the nicotine-induced ACTH and corticosterone secretion. L-NNA (2 mg/kg) and 7-nitroindazole (7-NI 20 mg/kg), neuronal NOS inhibitors in vivo also significantly augmented the nicotine-induced ACTH and corticosterone levels. L-arginine greatly impaired the nicotine-induced hormone responses and reversed the L-NNA elicited enhancement of the nicotine-evoked ACTH and corticosterone response. In contrast to the constitutive eNOS and nNOS antagonists, an inducible NOS antagonist guanethidine (50-100 mg/kg i.p.) did not substantially affect the nicotine-elicited pituitary-adrenocortical responses. Indomethacin (2 mg/kg i.p.), a non-selective cyclooxygenase blocker abolished the L-NAME and L-NNA-induced enhancement of the nicotine-evoked ACTH and corticosterone response. These results indicate that NO is an inhibitory mediator in the HPA axis activity. Inhibition of its generation by eNOS and nNOS significantly enhances the nicotine-induced HPA response. Under basal conditions iNOS is not involved in the nicotine-induced ACTH and corticosterone secretion. Prostaglandins play an obligatory role in the response of HPA axis to systemic nicotine administration.     

Vascular and renal effects of dopamine during extracellular volume expansion: Role of nitric oxide pathway

OBJECTIVE: The aim of the study was to determine the possible role of NO-system activation in vascular and renal effects of the dopaminergic system and the probable interaction between both systems during acute volume expansion in rats. DESIGN AND METHODS: Expanded (10% bw) and non-expanded anaesthetized male Wistar rats were treated with haloperidol, a DA receptor antagonist (3 mg/kg bw, ip). Mean arterial pressure, diuresis, natriuresis, renal plasma flow, glomerular filtration rate, nitrites and nitrates excretion (NOx) were determined. NADPH diaphorase activity was measured using a histochemistry technique in kidney, aorta and renal arteries. NOS activity in kidney and aorta from expanded and non-expanded animals was determined with L-[U14C]-arginine substrate, in basal conditions and after DA (1 microM) administration. RESULTS: The hypotensive effect of L-arg and hypertension induced by L-NAME were not modified by haloperidol. This blocker reverted the increase in diuresis, natriuresis and RPF induced by L-arg in both groups. Dopaminergic blockade induced a decrease in NOx excretion and in NADPH-diaphorase activity in glomeruli, proximal tubule and medullar collecting duct and in endothelium and vascular smooth muscle of renal arteries. DA induced an increase in NOS activity in renal medulla and cortex in both groups, but no changes in the aorta were observed. CONCLUSIONS: Our results suggest that renal DA would be associated with the renal response induced by NO during extracellular volume expansion. NO-system activation would be one of the mechanisms involved in renal DA activity during saline load, but NO appears not to be involved in DA vascular effects.     

Effect of combination of nitric oxide synthase and cyclooxygenase inhibitors on carrageenan-induced pleurisy in rats

In the present study, we examined the effects of L-nitroarginine methylester (L-NAME), a non-selective nitric oxide synthase (NOS) inhibitor, indomethacin (IND), a non-selective COX inhibitor and a combination of these agents (L-NAME+IND) on carrageenan-induced pleurisy in rats. Exudate volume, albumin leakage, leukocyte influx, exudate and plasma nitrite/nitrate (NO(x)) levels and exudate PGE(2) levels increased markedly 6 h after an intrapleural injection of 2% carrageenan. First, the effects of L-NAME and IND alone were investigated. L-NAME non-significantly reduced exudate volume by 26% at 10 mg/kg (i.p.), and significantly by 45% at 30 mg/kg. IND dose-dependently decreased the exudate volume at 0.3-10 mg/kg (p.o.) and the effect reached the maximal level at 1 mg/kg (33%). Second, the effects of L-NAME (10 mg/kg, i.p.), IND (1 mg/kg, p.o.) and L-NAME+IND were examined. L-NAME and IND alone at the dose employed significantly reduced the exudate volume and albumin levels by 21-26%. L-NAME but not IND tended to reduce the increased exudate and plasma NO(x) by 18% and 19%, respectively. IND but not L-NAME significantly reduced leukocyte numbers and PGE(2) levels in the exudates by 25% and 77%, respectively. L-NAME+IND significantly reduced exudate volume, albumin leakage, leukocyte number, PGE(2) and NO(x) by 43%, 41%, 31%, 80% and 37%, respectively. The inhibitory effects of L-NAME+IND on exudate volume, albumin leakage and NO(x) levels were greater than those of L-NAME and IND alone. In conclusion, a non-selective NOS inhibitor and COX inhibitor showed anti-inflammatory effects at the early phase of carrageenan-induced pleurisy, and a combination of both inhibitors had a greater effect than each alone probably via the potentiation of NOS inhibition. The simultaneous inhibition of NOS and COX could be a useful approach in therapy for acute inflammation.     

VEGF(121)- and bFGF-induced increase in collateral blood flow requires normal nitric oxide production

The angiogenic proteins basic fibroblast growth factor (bFGF; FGF-2) and vascular endothelial growth factor 121 (VEGF(121)) are each able to enhance the collateral-dependent blood flow after bilateral femoral artery ligation in rats. To study the effect of nitric oxide (NO) synthase (NOS) inhibition on bFGF- or VEGF(121)-induced blood flow expansion, the femoral arteries of male Sprague-Dawley rats were ligated bilaterally, and the animals were given tap water [non-N(G)-nitro-L-arginine methyl ester (L-NAME) group; n = 36] or water that contained L-NAME (L-NAME group; 2 mg/ml, n = 36). Animals from each group were further divided into three subgroups: vehicle (n = 12), bFGF (5 microg x kg(-1) x day(-1), n = 12), or VEGF(121) (10 microg x kg(-1) x day(-1), n = 12). Growth factors were delivered via intra-arterial infusion with osmotic pumps over days 1-14. On day 16, after a 2-day delay to permit clearance of bFGF and VEGF from the circulation, maximal collateral blood flow was determined by (85)Sr- and (141)Ce-labeled microspheres during treadmill running. L-NAME (approximately 137 mg x kg(-1) x day(-1)) for 18 days increased systemic blood pressure (approximately 26%, P<0.001). In the absence of L-NAME, collateral-dependent blood flows to the calf muscles were greater in the VEGF(121)- and bFGF-treated subgroups (85 +/- 4.5 and 80 +/- 2.9 ml x min(-1) x 100 g(-1), respectively) than in the vehicle subgroup (49 +/- 3.0 ml x min(-1) x 100 g(-1), P<0.001). In the presence of NOS inhibition by L-NAME, blood flows to the calf muscles were essentially equivalent among the three subgroups (54 +/- 3.0, 56 +/- 5.1, and 47 +/- 2.0 ml x min(-1) x 100 g(-1) in the bFGF-, VEGF(121)-, and vehicle-treated subgroups, respectively) and were not different from the blood flow in the non-L-NAME vehicle subgroup. Our results therefore indicate that normal NO production is essential for the enhanced vascular remodeling induced by exogenous bFGF or VEGF(121) in this rat model of experimental peripheral arterial insufficiency. These results imply that a blunted endothelial NO production could temper vascular remodeling in response to these angiogenic growth factors.     

Endothelial dysfunction and arterial pressure regulation during early diabetes in mice: roles for nitric oxide and endothelium-derived hyperpolarizing factor

We determined whether nitric oxide (NO) counters the development of hypertension at the onset of diabetes in mice, whether this is dependent on endothelial NO synthase (eNOS), and whether non-NO endothelium-dependent vasodilator mechanisms are altered in diabetes in mice. Male mice were instrumented for chronic measurement of mean arterial pressure (MAP). In wild-type mice, MAP was greater after 5 wk of N(omega)-nitro-L-arginine methyl ester (L-NAME; 100 mg x kg(-1) x day(-1) in drinking water; 97 +/- 3 mmHg) than after vehicle treatment (88 +/- 3 mmHg). MAP was also elevated in eNOS null mice (113 +/- 4 mmHg). Seven days after streptozotocin treatment (200 mg/kg iv) MAP was further increased in L-NAME-treated mice (108 +/- 5 mmHg) but not in vehicle-treated mice (88 +/- 3 mmHg) nor eNOS null mice (104 +/- 3 mmHg). In wild-type mice, maximal vasorelaxation of mesenteric arteries to acetylcholine was not altered by chronic L-NAME or induction of diabetes but was reduced by 42 +/- 6% in L-NAME-treated diabetic mice. Furthermore, the relative roles of NO and endothelium-derived hyperpolarizing factor (EDHF) in acetylcholine-induced vasorelaxation were altered; the EDHF component was enhanced by L-NAME and blunted by diabetes. These data suggest that NO protects against the development of hypertension during early-stage diabetes in mice, even in the absence of eNOS. Furthermore, in mesenteric arteries, diabetes is associated with reduced EDHF function, with an apparent compensatory increase in NO function. Thus, prior inhibition of NOS results in endothelial dysfunction in early diabetes, since the diabetes-induced reduction in EDHF function cannot be compensated by increases in NO production.