Effects Of L-Arginine And Furosemide On Blood Pressure And Renal Function In Volume-Expanded Rats

1. The aim of the present study was to investigate the effects of L-arginine (L-Arg) on blood pressure and water and electrolyte excretion in control and extracellular fluid volume-expanded rats (10% bodyweight with 0.9% NaCl) and to determine whether diuretic treatment with furosemide (FUR) can be optimized by the administration of L-Arg in this model. 2. Both groups of animals were anaesthetized, divided into groups and treated with either 7.5 mg/kg FUR, 250 mg/kg L-Arg, 1 mg/kg NG-nitro-L-arginine methyl ester (L-NAME), FUR + L-NAME or FUR + L-Arg. Mean arterial pressure (MAP), diuresis, natriuresis and kaliuresis were determined. 3. Extracellular fluid volume expansion induced no changes in MAP in control and volume-expanded rats (92+/-6 vs 100+/-8 mmHg, respectively). The hypotension induced by FUR in control and volume-expanded rats (69+/-7 and 76+/-5 mmHg, respectively) was significantly (P < 0.01) enhanced by the administration of L-Arg (54+/-3 and 64+/-3 mmHg, respectively). 4. Injection of L-NAME increased MAP and diminished diuresis, natriuresis and kaliuresis in both groups. 5. Furosemide-induced water and electrolyte excretion was blunted by the administration of L-NAME. 6. The combination of L-Arg + FUR increased diuresis induced by FUR alone (control rats: 29.33+/-1.68 vs 12.91+/- 0.41 microL/min per 100 g, respectively; volume-expanded rats: 248.5+/-25.4 vs 112,6+/-8.3 microL/min per 100 g, respectively; P < 0.01). 7. The administration of the combination of L-Arg + FUR promoted a decrease in the sodium/water excretion ratio compared with the administration of FUR alone (control rats: 0.230+/-0.018 vs 0.45+/-0.03, respectively, P < 0.001; volume-expanded rats: 0.091+/-0.010 vs 0.22+/-0.03, respectively, P < 0.01). 8. The potassium/water excretion rate induced by FUR alone and in the presence of L-Arg followed a pattern similar to that seen for natriuresis (control rats: 0.35+/-0.05 vs 0.20+/-0.05 microEq/min per 100 g, respectively; volume-expanded rats: 0.045+/-0.008 vs 0.014+/-0.003 microEq/min per 100 g, respectively; P < 0.01). 9. The decrease in the electrolyte/water excretion ratio observed with FUR + L-Arg in volume-expanded rats was greater than in control animals. 10. The results of the present study show that the administration of FUR with L-Arg contributes to enhanced hypotensive and diuretic effects of FUR, thus diminishing the relative electrolyte excretion in normal conditions and in extracellular fluid volume expansion.     

Microcystin-LR promote intestinal secretion of water and electrolytes in rats.

We showed previously that exposure to microcystin-LR causes renal toxic effects in isolated perfused rat kidney, and that inflammatory mediators from supernatants of macrophages stimulated by microcystin-LR are involved in this process. The aim of this research was to examine water and electrolytes secretion in vivo, induced by microcystin-LR and supernatant of macrophages stimulated for this toxin (SUP.MphiS + MCLR), using perfused rat ileal segment and ligated intestinal loop models. We found microcystin-LR at 1 microg/ml (0.09 +/- 0.003* vs. control 0.07 +/- 0.001 g of secretion/2 cm of loop; P < 0.05*) and the SUP.MphiS + MCLR after 18 h postinoculation (0.10 +/- 0.003 vs. control 0.03 +/- 0.002 g/cm) caused intestinal secretion. In addition, microcystin-LR caused significant sodium secretion (-2.18 +/- 0.72* vs. control 2.18 +/- 0.50 microEq g(-1) min(-1)), potassium (-0.26 +/- 0.04* vs. control 0.32 +/- 0.03 microEq g(-1) min(-1)), chloride (MCLR = -3.29 +/- 1.93* vs. control 0.88 +/- 1.25 microEq g(-1) min(-1)) and water (-0.012 +/- 0.004* vs. control 0.002 +/- 0.002 ml g(-1) min(-1)). We also demonstrated SUP.MphiS + MCLR to induce intestinal secretion of electrolytes (sodium, potassium, chloride) and water. These findings suggested that microcystin-LR and lamina propria macrophages-derived mediators are able to induce intestinal secretion in vivo, probably via inhibition of protein phosphatase.     

Contribution of the renin-angiotensin system to short-term blood pressure variability during blockade of nitric oxide synthesis in the rat.

1. The aim of this study was to investigate, by use of spectral analysis, (1) the blood pressure (BP) variability changes in the conscious rat during blockade of nitric oxide (NO) synthesis by the L-arginine analogue NG-nitro-L-arginine methyl ester (L-NAME); (2) the involvement of the renin-angiotensin system in these modifications, by use of the angiotensin II AT1-receptor antagonist losartan. 2. Blockade of NO synthesis was achieved by infusion for 1 h of a low-dose (10 micrograms kg-1 min-1, i.v., n = 10) and high-dose (100 micrograms kg-1 min-1, i.v., n = 10) of L-NAME. The same treatment was applied in two further groups (2 x n = 10) after a bolus dose of losartan (10 mg kg-1, i.v.). 3. Thirty minutes after the start of the infusion of low-dose L-NAME, systolic BP (SBP) increased (+10 +/- 3 mmHg, P < 0.01), with the effect being more pronounced 5 min after the end of L-NAME administration (+20 +/- 4 mmHg, P < 0.001). With high-dose L-NAME, SBP increased immediately (5 min: +8 +/- 2 mmHg, P < 0.05) and reached a maximum after 40 min (+53 +/- 4 mmHg, P < 0.001); a bradycardia was observed (60 min: -44 +/- 13 beats min-1, P < 0.01). 4. Low-dose L-NAME increased the low-frequency component (LF: 0.02-0.2 Hz) of SBP variability (50 min: 6.7 +/- 1.7 mmHg2 vs 3.4 +/- 0.5 mmHg2, P < 0.05), whereas the high dose of L-NAME not only increased the LF component (40 min: 11.7 +/- 2 mmHg2 vs 2.7 +/- 0.5 mmHg2, P < 0.001) but also decreased the mind frequency (MF: 0.2-0.6 Hz) component (60 min: 1.14 +/- 0.3 mmHg2 vs 1.7 +/- 0.1 mmHg2, P < 0.05) of SBP. 5. Losartan did not modify BP levels but had a tachycardic effect (+45 beats min-1). Moreover, losartan increased MF oscillations of SBP (4.26 +/- 0.49 mmHg2 vs 2.43 +/- 0.25 mmHg2, P < 0.001), prevented the BP rise provoked by the low-dose of L-NAME and delayed the BP rise provoked by the high-dose of L-NAME. Losartan also prevented the amplification of the LF oscillations of SBP induced by L-NAME; the decrease of the MF oscillations of SBP induced by L-NAME was reinforced after losartan. 6. We conclude that the renin-angiotensin system is involved in the increase in variability of SBP in the LF range which resulted from the withdrawal of the vasodilating influence of NO. We propose that NO may counterbalance LF oscillations provoked by the activity of the renin-angiotensin system.     

Troglitazone enhances glucose uptake induced by alpha-adrenoceptor stimulation via phosphatidylinositol 3-kinase in rat heart

1. Thiazolidinedione-derived agents have been reported to act as insulin sensitizers by augmenting insulin-dependent stimulation of phosphatidylinositol 3-kinase (PI3K) activity in a specific manner. It has been suggested that alpha-adrenoceptor stimulation mediates glucose uptake through PI3K in the heart. 2. To elucidate whether the thiazolidinedione-derived agent troglitazone (TRO) affects glucose uptake induced by alpha-adrenoceptor stimulation through PI3K, the rate of glucose uptake was quantified from the rate of accumulation of sugar phosphate (d[SP]/dt) using [(31)P] nuclear magnetic resonance spectroscopy after substitution of glucose with 2-deoxyglucose in rat perfused heart. Hearts were stimulated with 100 micromol/L phenylephrine plus 10 micromol/L propranolol (alpha-adrenoceptor stimulation), or 1 micromol/L isoproterenol plus 10 micromol/L phentolamine (beta-adrenoceptor stimulation). 3. The d[SP]/dt in the alpha- and beta-adrenoceptor-stimulated groups (0.45 +/- 0.06 and 0.42 +/- 0.04 micromol/min per g, respectively) was higher than that of the control group (0.27 +/- 0.02 micromol/min per g; P < 0.01). The addition of 2 microg/mL troglitazone to alpha-adrenoceptor stimulation augmented d[SP]/dt (0.72 +/- 0.08 micromol/min per g; P < 0.05 vs the alpha-adrenoceptor-stimulated group), which was effectively blocked by 3 micromol/L wortmannin (0.35 +/- 0.06 micromol/min per g; P < 0.01 vs troglitazone + alpha-adrenoceptor stimulation group). However, addition of troglitazone to beta-adrenoceptor stimulation did not alter d[SP]/dt (0.33 +/- 0.02 micromol/min per g; P = NS vs the beta-adrenoceptor-stimulated group). 4. These results indicate that troglitazone acutely enhances alpha-adrenoceptor stimulation on glucose uptake through a PI3K-dependent pathway, thus possibly improving glucose utilization in a catecholamine-released state.     

Modulation of haemostatic function and prevention of experimental thrombosis by red wine in rats: a role for increased nitric oxide production.

1. The effects of ethyl alcohol and wine (red and white) on haemostatic parameters and experimental thrombosis were studied in rats; NO was evaluated as a possible mediator of these effects. 2. We found that red wine (12% alcohol) supplementation (8.4 +/- 0.4 ml d-1 in drinking water, for 10 days) induced a marked prolongation of 'template' bleeding time (BT) (258 +/- 13 vs 132 +/- 13 s in controls; P < 0.001), a decrease in platelet adhesion to fibrillar collagen (11.6 +/- 1.0 vs 32.2 +/- 1.3%; P < 0.01) and a reduction in thrombus weight (1.45 +/- 0.33 vs 3.27 +/- 0.39 mg; P < 0.01). 3. Alcohol-free red wine showed an effect similar to red wine. In contrast, neither ethyl alcohol (12%) nor white wine (12% alcohol) affected these systems. 4. All these effects were also observed after red wine i.v. injection (1 ml kg-1 of 1:4 dilution) 15 min before the experiments. 5. The effects of red wine were prevented by the NO inhibitor, N omega nitro-L-arginine-methyl ester (L-NAME). L-arginine, not D-arginine, reversed the effect of L-NAME on red wine infusion. 6. Red wine injection induced a 3 fold increase in total radical-trapping antioxidant parameter values of rat plasma with respect to controls, while white wine and alcohol did not show any effect. 7. Our study provides evidence that red wine modulates primary haemostasis and prevents experimental thrombosis in rats, independently of its alcohol content, by a NO-mediated mechanism.     

Effects of glycine on glomerular filtration rate and segmental tubular handling of sodium in conscious rats

1. Infusion of the amino acid glycine leads to an increase in effective renal plasma flow (ERPF) and glomerular filtration rate (GFR) by a mechanism that possibly involves stimulation of nitric oxide (NO). Because NO also increases proximal tubular fluid output (Vprox) by inhibition of proximal tubular Na+ reabsorption and modulation of the tubuloglomerular feedback system, we hypothesized that glycine would increase Vprox as measured by lithium clearance (CLi). 2. In the first series of experiments, the effect of glycine infusion (4 mg/min) was examined in conscious, unstressed, chronically catheterized rats. In an additional series of experiments, the effect of glycine was examined under similar conditions in rats pretreated with a NO synthase (NOS) inhibitor (NG-nitro-L-arginine methyl ester (L-NAME), 2.5 microg/min). 3. Glycine significantly increased ERPF (from 3268 to 4018 microL/min per 100 g bodyweight (BW)), GFR (from 874 to 1009 microL/min per 100 g BW), CLi (from 275 to 461 microL/min per 100 g BW) and Na+ clearance (CNa; from 2.9 to 14.0 microL/min per 100 g BW). Fractional excretion of lithium (FELi; from 32 to 46%) and CNa/CLi (from 0.99 to 2.99%) also rose, indicating inhibition of proximal and distal nephron Na+ reabsorption, respectively. In the rats pretreated with L-NAME, similar haemodynamic and tubular responses to glycine infusion were seen, suggesting that the effects were not mediated by NO. 4. We conclude, that glycine increases ERPF and GFR and it also inhibits proximal and distal nephron Na+ reabsorption leading to an increase in CLi and CNa. There was no indication that any of these effects were mediated by NO.     

Impact of the endothelial nitric oxide synthase gene G894T polymorphism on renal endothelial function in patients with type 2 diabetes

OBJECTIVE: Endothelial dysfunction and increased oxidative stress contribute to the progression of diabetic nephropathy. To analyze the functional significance of the G894T polymorphism of NOS3, the gene encoding endothelial nitric oxide synthase (NOS), we assessed basal nitric oxide activity and the amount of oxidative stress in the renal circulation of patients with type 2 diabetes. METHODS: Renal plasma flow (RPF) was assessed by steady-state input clearance technique with sodium para-aminohippurate in 84 patients with type 2 diabetes and 84 patients without diabetes. RPF was measured at baseline and after the infusion of the NOS inhibitor N-monomethyl-L-arginine (4.25 mg/kg); the substrate of NOS L-arginine (100 mg/kg); and coinfusion of vitamin C (3 g) with L-arginine (100 mg/kg). RESULTS: The decrease of RPF to N-monomethyl-L-arginine was similar between carriers of the T allele and homozygous carriers of the G allele in patients with diabetes (-56+/-40 vs. -68.1+/-74 ml/min/1.73 m, P=0.342) and patients without diabetes (-66.7+/-81 vs. -58.3+/-63 ml/min/1.73 m, P=0.606). In patients with diabetes, however, carriers of the T allele revealed a more pronounced increase of RPF to coinfusion of vitamin C with L-arginine than homozygous carriers of the G allele (61.8+/-75 vs. 22.3+/-73 ml/min/1.73 m, P=0.021), whereas in patients without diabetes the response of RPF to coinfusion of vitamin C with L-arginine was similar between both groups (46.2+/-80 vs. 70.7+/-86 ml/min/1.73 m, P=0.217). Gene-environment interaction between disease (diabetes) and genotype (genotype GG vs. genotype GT/TT) was observed for increase of RPF to coinfusion of vitamin C with L-arginine (P=0.020). CONCLUSION: G894T polymorphism of NOS3 has no impact on the basal nitric oxide activity of renal circulation. In contrast, the T allele is associated with increased oxidative stress in the renal circulation in patients with diabetes suggesting a specific role of the G894T polymorphism in the pathogenesis of diabetic nephropathy.     

Protective effect of alpha-lipoic acid against ischaemic acute renal failure in rats

1. In the present study, we investigated whether treatment with alpha-lipoic acid (LA), a powerful and universal anti-oxidant, has renal protective effects in rats with ischaemic acute renal failure (ARF). 2. Ischaemic ARF was induced by occlusion of the left renal artery and vein for 45 min followed by reperfusion, 2 weeks after contralateral nephrectomy. Blood urea nitrogen (BUN), plasma concentrations of creatinine (Pcr) and urinary osmolality (Uosm) were measured for the assessment of renal dysfunction. Creatinine clearance (Ccr) and fractional excretion of Na+ (FENa) were used as indicators of glomerular and tubular function, respectively. 3. Renal function in ARF rats decreased markedly 24 h after reperfusion. Intraperitoneal injection of LA at a dose of 10 mg/kg before the occlusion tended to attenuate the deterioration of renal function. A higher dose of LA (100 mg/kg) significantly (P < 0.01) attenuated the ischaemia/reperfusion-induced increases in BUN (19.1 +/- 0.7 vs 7.2 +/- 0.7 mmol/L before and after treatment, respectively), Pcr (290 +/- 36 vs 78.1 +/- 4.2 micromol/L before and after treatment, respectively) and FENa (1.39 +/- 0.3 vs 0.33 +/- 0.09% before and after treatment, respectively). Treatment with 100 mg/kg LA significantly (P < 0.01) increased Ccr (0.70 +/- 0.13 vs 2.98 +/- 0.27 mL/min per kg before and after treatment, respectively) and Uosm (474 +/- 39 vs 1096 +/- 80 mOsmol/kg before and after treatment, respectively). 4. Histopathological examination of the kidney of ARF rats revealed severe lesions. Tubular necrosis (P < 0.01), proteinaceous casts in tubuli (P < 0.01) and medullary congestion (P < 0.05) were significantly suppressed by the higher dose of LA. 5. A marked increase in endothelin (ET)-1 content in the kidney after ischaemia/reperfusion was evident in ARF rats (0.43 +/- 0.02 ng/g tissue) compared with findings in sham- operated rats (0.20 +/- 0.01 ng/g tissue). Significant attenuation (P < 0.01) of this increase occurred in ARF rats treated with the higher dose of LA (0.24 +/- 0.03 ng/g tissue). 6. These results suggest that administration of LA to rats prior to development of ischaemic ARF prevents renal dysfunction and tissue injury, possibly through the suppression of overproduction of ET-1 in the postischaemic kidney.     

Central and peripheral haemodynamic effects of L-NAME infusion in healthy volunteers

AIMS: To evaluate the effects of the intravenous administration of the nitric oxide synthesis inhibitor N(g)nitro-L-arginine methyl ester (L-NAME) in healthy volunteers. METHODS: L-NAME (0.25, 0.5 and 0.75 mg/kg over 8 min) was infused in 13 healthy male volunteers. Finally, subjects were infused with either L- or D-arginine. RESULTS: L-NAME resulted in dose-dependent falls in heart rate 60 bpm (55-64 bpm) to 49 bpm (46-52 bpm) (P<0.01) and increased mean arterial pressure 77.0 mmHg (73.2-80.8 mmHg) to 90.0 mmHg (87.1-92.8 mmHg) (P<0.01). The cardiac output was significantly reduced after each L-NAME infusion, and systemic vascular resistance increased linearly over the dosage range. Cardiac stroke volume was significantly reduced only following 0.75 mg/kg/min L-NAME: from 100 ml (91.3-108.7 ml) to 83 ml (74.7-91.4 ml); P<0.01. Forearm blood flow was unchanged at any dosage. L-arginine but not D-arginine infusion reversed the haemodynamic effects of L-NAME. CONCLUSIONS: Contrasting with the profound dose-dependent effects of L-NAME had significant effects on central haemodynamics but no discernible effects on peripheral blood flow.     

Regional blood flow responses to acute ANG II infusion: effects of nitric oxide synthase inhibition.

We hypothesized that nitric oxide (NO) opposes regional vasoconstriction caused by acute angiotensin II (ANG II) infusion in conscious rats. Mean arterial pressure (MAP), blood flow, and vascular conductance (regional blood flow/ MAP; ml/min/100 g/mm Hg) were measured and/or calculated before and at 2 min of ANG II infusion (0.05 or 1 microg/kg/min, i.a.) in the absence and presence of NO synthase (NOS) inhibition [N(G)-nitro-L-arginine methyl ester (L-NAME), 0.25 or 1 mg/kg, i.a.]. ANG II reduced stomach and hindlimb conductance only after NOS inhibition. For example, whereas 0.05 microg/kg/min ANG II did not attenuate conductance in the stomach (i.e., 1.04+/-0.08 to 0.93+/-0.12 ml/min/100 g/mm Hg), this variable was reduced (i.e., 0.57+/-0.14 to 0.34-/+0.05 ml/min/100 g/mm Hg; p < 0.05) when ANG II was infused after 0.25 mg/kg L-NAME. In addition, whereas hindlimb conductance was similar before and after administering 1 microg/kg/min ANG II (i.e., 0.13+/-0.01 and 0.09+/-0.02, respectively), this variable was reduced (i.e., 0.07+/-0.01 and 0.02+/-0.00, respectively; p < 0.05) when ANG II was infused after 1 mg/kg L-NAME. These findings indicate that NO opposes ANG II-induced vasoconstriction in the stomach and hindlimb. In contrast, whereas both doses of ANG II decreased (p < 0.05) vascular conductance in the kidneys and small and large intestine regardless of whether NOS inhibition was present, absolute vascular conductance was lower (p < 0.05) after L-NAME. For example, 1 microg/kg ANG II reduced renal conductance from 3.34+/-0.31 to 1.22+/-0.14 (p < 0.05). After 1 mg/kg L-NAME, renal conductance decreased from 1.39+/-0.18 to 0.72+/-0.16 (p < 0.05) during ANG II administration. Therefore the constrictor effects of NOS inhibition and ANG II are additive in these circulations. Taken together, our results indicate that the ability of NO to oppose ANG II-induced constriction is not homogeneous among regional circulations.     

Vitamin C prevents radiation-induced endothelium-dependent vasomotor dysfunction and de-endothelialization by inhibiting oxidative damage in the rat

1. The present study was undertaken to determine whether endothelial function or morphology was altered in aortic rings of rats after irradiation, to investigate the mechanism of radiation effects on the endothelium and to examine the effect of vitamin C treatment against radiation-induced damage of the endothelium. 2. Female Sprague-Dawley rats were randomized into four groups (control, radiation, radiation + vitamin C, radiation + vitamin C + NG-nitro-L-arginine methyl ester (L-NAME); n = 10 for each group and n = 7 for the control group) and were irradiated with 10 Gy of 137Cs as a radiation source. Segments of the thoracic aorta were obtained and isometric tension, levels of 8-hydroxydeoxyguanosine (OH-dG) and immunohistochemical staining were measured. 3. Irradiation significantly impaired the acetylcholine-induced vasodilation of aortic segments, an effect that could be prevented by pretreatment with vitamin C (500 mg/kg per day). This beneficial effect of vitamin C was abolished by the addition of L-NAME (100 microg/kg per day), an inhibitor of nitric oxide (NO) synthesis. Irradiation significantly increased the level of OH-dG in the aorta (1.02 +/- 0.27 vs 2.61 +/- 0.78 OH-dG/105 deoxyguanosine (dG) for control and irradiated tissues, respectively; P < 0.01), an increase that was prevented by vitamin C treatment (1.59 +/- 0.23 OH-dG/105 dG; P < 0.01). Irradiation caused significant de-endothelialization (von Willebrand factor (vWF) staining was 93 +/- 7 vs 100% in irradiated and control tissues, respectively; P < 0.05) and this was prevented by vitamin C treatment (vWF staining 98 +/- 3%; P < 0.05). 4. Radiation caused endothelial damage and impaired NO production through oxidative injury, resulting in a selective impairment of endothelial-dependent vasodilation that could be prevented by vitamin C, partly through anti-oxidant mechanisms.     

Cerebral blood flow and cerebrovascular reactivity after inhibition of nitric oxide synthesis in conscious goats.

1. The role of nitric oxide in the cerebral circulation under basal conditions and after vasodilator stimulation was studied in instrumented, conscious goats, by examining the action of inhibiting endogenous nitric oxide production with NG-nitro-L-arginine methyl ester (L-NAME). 2. In 6 unanaesthetized goats, blood flow to one brain hemisphere (electromagnetically measured), systemic arterial blood pressure and heart rate were continuously recorded. L-NAME (35 mg kg-1 by i.v. bolus) decreased resting cerebral blood flow by 43 +/- 3%, increased mean arterial pressure by 21 +/- 2%, and decreased heart rate by 41 +/- 2%; cerebrovascular resistance increased by 114 +/- 13% (P < 0.01); the immediate addition of i.v. infusion of L-NAME (0.15-0.20 mg kg-1 during 60-80 min) did not significantly modify these effects. Cerebral blood flow recovered at 72 h, arterial pressure and cerebrovascular resistance at 48 h, and heart rate at 6 days after L-NAME treatment. 3. A second treatment with L-NAME scheduled as above reproduced the immediate haemodynamic effects of the first treatment, which (except bradycardia) reversed with L-arginine (200-300 mg kg-1 by i.v. bolus). 4. Acetylcholine (0.01-0.3 micrograms), sodium nitroprusside (3-100 micrograms) and diazoxide (0.3-9 mg), injected into the cerebral circulation of 5 conscious goats, produced dose-dependent increases in cerebral blood flow, and decreases in cerebrovascular resistance; sodium nitroprusside (30 and 100 micrograms) also caused hypotension and tachycardia.(ABSTRACT TRUNCATED AT 250 WORDS)     

cGMP介导以注入L—精氨酸诱导的大鼠血管加压素释放效应

目的：探讨ｃＧＭＰ是否介导Ｌ－精氨酸（一氧化氮合酶物）引起的血管加压素（ＡＶＰ）释放增多效应。方法：用放射免疫法测定大鼠血浆中ＡＶＰ水平。结果：侧脑室分别注射Ｌ－精氨酸和８－溴－ｃＧＭＰ（一种可透过膜的ｃＧＭＰ衍生物）能刺激血浆ＡＶＰ水平增加［分别从（３．２±０．５）升至（５．８±１．４）ｎｇ·Ｌ＾－１，从（２．６±０．３）升至６６．６±０．４）ｎｇ·Ｌ＾－１，Ｐ〈０．０１］，同时注射Ｌ－精氨酸和     

Inhibition of nitric oxide synthase by 1-(2-trifluoromethylphenyl) imidazole (TRIM) in vitro: antinociceptive and cardiovascular effects.

1. The ability of a range of substituted imidazole compounds to inhibit mouse cerebellar neuronal nitric oxide synthase (nNOS), bovine aortic endothelial NOS (eNOS) and inducible NOS (iNOS) from lungs of endotoxin-pretreated rats was investigated. In each case the substrate (L-arginine) concentration employed was 120 nM. 2. 1-(2-Trifluoromethylphenyl) imidazole (TRIM) was a relatively potent inhibitor of nNOS and iNOS (IC50S of 28.2 microM and 27.0 microM respectively) but was a relatively weak inhibitor of eNOS (IC50, 1057.5 microM). The parent compound, imidazole, was a weak inhibitor of all three NOS isoforms (IC50S: nNOS, 290.6 microM; eNOS, 101.3 microM; iNOS, 616.0 microM). Substitution of imidazole with a phenyl group to yield I-phenylimidazole (PI) resulted in an isoform non-selective increase in inhibitory potency (IC50S: nNOS, 72.1 microM; eNOS, 86.9 microM; iNOS, 53.9 microM). Further substitution of the attached phenyl group resulted in an increase in nNOS and a decrease in eNOS inhibitory potency as in TRIM, 1-chlorophenylimidazole (CPI; IC50S: nNOS, 43.4 microM; eNOS, 392.3 microM; iNOS, 786.5 microM) and 1-(2,3,5,6-tetrafluorophenyl) imidazole (TETRA-FPI; IC50S; nNOS, 56.3 microM; eNOS, 559.6 microM; iNOS, 202.4 microM). 3. The ability of TRIM to inhibit mouse cerebellar nNOS activity in vitro was influenced by the concentration of L-arginine (0.12-10.0 microM) in the incubation medium. When mouse cerebellar nNOS was used as enzyme source a double reciprocal (Lineweaver-Burk) plot in the presence/absence of TRIM (50 microM) revealed a competitive inhibitory profile. The K(m) for L-arginine and the Ki for TRIM calculated from these data were 2.4 microM and 21.7 microM, respectively. The ability of TRIM to inhibit mouse cerebellar nNOS activity in vitro was unaffected by varying the time of exposure of the enzyme to TRIM from 0-60 min at 0 degree C. 4. TRIM exhibits potent antinociceptive activity in the mouse as evidenced by inhibition of acetic acid induced abdominal constrictions. The ED50 for TRIM following i.p. administration was 20 mg kg-1 (94.5 mumol kg-1). The antinociceptive effect of TRIM was reversed by pretreatment of animals with L-arginine (50 mg kg-1, i.p.) and was not accompanied by sedation, motor ataxia or behavioural changes (rearing, crossing, circling, dipping) as assessed by use of a box maze procedure. 5. L-NG nitro arginine methyl ester (L-NAME, 20 mg kg-1, i.v.) but not TRIM (0.5-20 mg kg-1, i.v.) increased mean arterial blood pressure (MAP) in the urethane-anaesthetized rat. 6. L-NAME (100 microM) potentiated the contractile response of the rabbit isolated aorta to phenylephrine (ED50; 0.084 +/- 0.01 microM in the presence and 0.25 +/- 0.05 microM in the absence of L-NAME; maximum response, 7.7 +/- 0.4 g in the presence and 5.6 +/- 0.5 g in the absence of L-NAME, n = 6, (P < 0.05) whilst TRIM (1-100 microM) was without effect. L-NAME (100 microM) but not TRIM (1-100 microM) also reduced carbachol-induced relaxation of the phenylephrine-precontracted rabbit aorta preparation. 7. L-NAME (50 microM) potentiated the vasoconstrictor effect of bolus-injected noradrenaline (10-1000 nmol) and reduced the vasodilator effect of carbachol (10 microM) added to the Krebs reservoir in the rat perfused mesentery preparation. L-NAME (50 microM) also reduced nitric oxide (NO) release (measured by chemiluminescence of nitrite in the Krebs perfusate) in response to noradrenaline (100 nmol; 53.8 +/- 4.0 pmol ml-1 in the presence and 84.8 +/- 8.0 pmol ml-1 in the absence of L-NAME, n = 15, P < 0.05) and carbachol (10 microM; 63.9 +/- 5.0 pmol ml-1 in the presence and 154.0 +/- 9.0 pmol ml-1 in the absence of L-NAME, n = 15, P < 0.05). TRIM (50 microM) did not affect either the vasoconstrictor response to noradrenaline or the vasodilator response to carbachol or the accompanying release of NO from the perfused rat mesentery.     

Intravascular granulocyte aggregates caused by the selectin-binding carbohydrate fucoidin in pig kidneys

1. Renal postischaemic reperfusion injury constitutes a significant problem after kidney transplantation. The polysaccharide fucoidin improves postischaemic function in lamb hearts, presumably by blocking selectin-mediated leucocyte adhesion. 2. In the present study, eight pairs of ischaemic pig kidneys were reperfused in an ex vivo model with autologous blood with or without fucoidin (100 mg/L). 3. Fucoidin resulted in a significant decrease in renal blood flow (45 +/- 5 vs 178 +/- 22 mL/min per 100 g; P < 0.001) and increased vascular resistance (3.80 +/- 0.07 vs 0.60 +/- 0.12 mmHg/mL per min per 100 g; P < 0.001). 4. Histological examination revealed granulocyte emboli in afferent glomerular arteries in five of six fucoidin-treated kidneys and in one of six controls (Fisher's exact test; P < 0.001). 5. In vitro experiments with human granulocytes showed that large granulocyte aggregates were induced by fucoidin at concentrations similar to those used in reperfused kidneys, whereas slightly lower doses of fucoidin prevented l-selectin-dependent homotypic granulocyte adhesion. 6. The formation of embolizing granulocyte aggregates defines a narrow therapeutic range for fucoidin and calls into question its experimental use as an inhibitor of selectin-mediated leucocyte adhesion.     

Differential effects of L-arginine on the inhibition by NG-nitro-L-arginine methyl ester of basal and agonist-stimulated EDRF activity.

1. An isolated, buffer-perfused rabbit ear preparation was used to investigate the influence of NG-nitro-L-arginine methyl ester (L-NAME) on endothelium-dependent vasodiltation and modulation of vasoconstrictor responses and vascular conductance. 2. Acetylcholine (0.55 pmol-1.6 nmol) caused dose-related vasodilatation of preparations constricted by the combination of 5-hydroxytryptamine and histamine (both 1 microM), with an ED50 = 31.1 +/- 7.8 pmol and a maximum dilatation of 69.9 +/- 4.3%. In the presence of 10 microM L-NAME the dose-response for vasodilator effects was shifted significantly (P less than 0.001) to the right (ED50 = 3.07 +/- 1.18 nmol) and there was a significant (P less than 0.01) depression of the maximum response (Rmax = 44.3 +/- 4.0%). The higher concentration of 100 microM L-NAME completely abolished vasodilatation to acetylcholine. L-Arginine (10 mM) did not reverse the inhibitory actions of L-NAME at either concentration. 3. L-NAME 100 microM, augmented vascular tone induced by 1 microM 5-hydroxytryptamine and 1 microM histamine, thus altering the characteristics of both pressure/flow and conductance/flow relationships such that conductance was reduced at all flow rates. The augmentation of constrictor tone was reversed in a concentration-dependent manner by L-arginine (10 microM-10 mM) and the effect of L-NAME on the conductance/flow relationships was similarly reversed by 10 mM L-arginine. The augmentation of tone was endothelium-dependent as it did not occur following functional destruction of the endothelium by perfusion of the vascular bed with the detergent CHAPS (0.3%) for 150s.(ABSTRACT TRUNCATED AT 250 WORDS)     

The influence of chronic inhibition of nitric oxide synthesis on contractile and relaxant properties of rat carotid and mesenteric arteries

Balloon denudation of the rat carotid artery leads to an immediate decrease in beta-adrenoceptor-medi-ated vasodilator response. However, this arterial function becomes significantly enhanced during subsequent formation of neointima with the endothelial cell lining still being absent. It was therefore hypothesized that chronic suppression of endothelium-dependent nitric oxide (NO) synthesis may eventually upregulate the beta-adrenoceptor system on vascular smooth muscle. To investigate this hypothesis, male Wistar rats were treated chronically with the L-arginine analogue NG-nitro-L-arginine methyl ester (L-NAME) to inhibit the synthesis of NO (i.e. 15 mg/kg per day or 0.06 mmol/kg per day for 6 weeks p.o.). Prior to the experiments the mean arterial blood pressure (MAP) was significantly elevated in the L-NAME-treated rats (i.e. 128.4+/-3.4 mmHg vs. 100.0+/-2.9 mmHg, L-NAME vs. control, n=4, P<0.05). The functional properties of the isolated vessel preparations were established by isometric force measurement in a myograph setup, in the absence of L-NAME. The maximal contractile responses to high potassium chloride solution (100 mM), to the alpha1-adrenoceptor agonist phenylephrine, and to the thromboxane A2-agonist U46619, were not influenced by chronic L-NAME-treatment in the carotid and mesenteric artery preparations. The vasodilator responses to the cholinergic agonist methacholine were significantly impaired in the carotid arteries of L-NAME-treated animals: 30.9+/-7.9% vs. 64.6+/-2.0%, P<0.05, L-NAME vs. control, n=10. However, these responses appeared not to be modulated in the mesenteric artery preparations after chronic L-NAME treatment. Separate experiments showed that these responses could be blocked both in the rat carotid and mesenteric artery with L-NAME (10 mM) in vitro. Addition of the NO synthase substrate L-arginine could partially but significantly reverse this blockade. Chronic inhibition of NO synthesis caused significant deterioration of beta-adrenoceptor-mediated vasodilator responses. For carotid arteries: Emax=36.1+/-9.4% vs. 65.9+/-6.0%, P<0.05, L-NAME vs. control, n=5; and pD2=6.7+/-0.2 and 7.4+/-0.1, respectively, P<0.05, n=5. For mesenteric arteries: pD2=7.7+/-0.0 and 8.0+/-0.0, respectively, P<0.05, n=5. From these data, it may be concluded that chronic L-NAME treatment results in a stable impairment of the endothelium-dependent NO system in the rat carotid but not mesenteric arteries. The stated hypothesis fails as the beta-adrenoceptor-induced vasorelaxation of carotid and mesenteric arteries became significantly impaired, rather than enhanced. Taken together, the beta-adrenoceptor function in the rat carotid artery is apparently more dependent on endothelial NO synthesis than that in the rat mesenteric artery.     

Upregulation of renal endothelin receptors in rats with cyclosporine A-induced nephrotoxicity

Measurement of endothelin receptors by binding assay was performed in rats treated with cyclosporine A (CYA). Cyclosporine A administration at 50 mg/kg i.p. for 4 days resulted in renal function impairment as indicated by a significant increase in serum creatinine concentration (from 0.46 +/- 0.02 to 0.61 +/- 0.03 mg/dl. P less than 0.01) and a significant decrease in 24 h creatinine clearance (from 0.65 +/- 0.04 to 0.41 +/- 0.02 ml/min per 100 g, P less than 0.01). Renal endothelin (ET) receptor density was significantly higher in CYA-treated rats (312 +/- 34 vs. 196 +/- 26 fmol/mg protein, P less than 0.01). These data support the possible involvement for endothelin in the increased renal vascular resistance associated with CYA-induced nephrotoxicity.     

Effects of the angiotensin-converting enzyme inhibitor alacepril on exercise capacity and neurohormonal factors in patients with mild-to-moderate heart failure

1. Alacepril is a long-acting, sulphydryl-containing angiotensin-converting enzyme inhibitor. Data are limited regarding the effects of alacepril on exercise tolerance in patients with chronic heart failure (CHF). The aim of the present study was to determine the effects of chronic alacepril treatment on exercise capacity and neurohormones in patients with CHF. 2. The effects of 12 weeks treatment with alacepril on clinical, echocardiographic and cardiopulmonary exercise variables were studied in 18 CHF patients (mean age: 63 +/- 2 years; New York Heart Association (NYHA) class I n = 6, class II n = 10, class III n = 2) in a cross-over fashion. Resting levels of plasma noradrenaline, renin-angiotensin system activity and natriuretic peptides were evaluated. 3. Treatment with alacepril significantly improved NYHA functional class and decreased cardiothoracic ratio (60.1 +/- 2.0 vs 58.1 +/- 1.9% for baseline vs alacepril, respectively; P < 0.01). Cardiac dimensions by echocardiogram were decreased after alacepril therapy. Peak Vo2 (17.7 +/- 1.2 vs 19.5 +/- 1.3 mL/min per kg; P < 0.01) and anaerobic threshold (11.7 +/- 0.6 vs 13.2 +/- 0.9 mL/min per kg; P < 0.01) increased with alacepril treatment. Plasma noradrenaline and plasma angiotensin II levels were not altered, but plasma aldosterone (77.7 +/- 13.5 vs 51.7 +/- 9.7 pg/mL; P < 0.01), atrial natriuretic peptide (ANP; 86.5 +/- 20.3 vs 43.6 +/- 7.6 pg/mL; P < 0.05) and brain natriuretic peptide (BNP; 222.7 +/- 59.3 vs 117.7 +/- 34.3 pg/mL; P < 0.05) levels decreased after alacepril treatment. 4. These results suggest that treatment with alacepril improves functional status and exercise capacity in patients with mild-to-moderate CHF. Neurohormones were favourably influenced by alacepril therapy, with significant decreases in plasma aldosterone, ANP and BNP levels.     

The role of nitric oxide in iron-induced rat renal injury

Iron overload and enhanced hydroxyl radical (*OH) formation have been implicated as the causative factors of oxidative stress in different organs. Both pro-oxidant and anti-oxidant properties have been reported for nitric oxide (NO) in iron-mediated tissue injury. To determine the contribution of NO to iron-induced renal injury, eight groups of rats (eight in each group) were studied as follows: control (normal saline), L-Arg (L-arginine as a substrate of NO synthase, 400 mg/kg), L-NAME (an inhibitor of NO synthase, 8 mg/kg), Fe (iron dextran, 600 mg/kg), DFO (deferroxamine as a chelator of iron, 150 mg/kg), Fe+L-Arg, Fe+L-NAME, DFO+L-Arg. Twenty-four hours after the injections, blood samples were taken and kidneys removed for biochemical analysis. Plasma creatinine and urea were used to stimulate renal function. Renal tissue and plasma vitamin E levels, the most important endogenous fat soluble antioxidant, were measured by HPLC and UV detection. In this study, renal function was markedly reduced in the Fe group compared to controls (creatinine, 1.02+/-0.05 mg/dL versus 0.78+/-0.04 P <0.05; urea, 49.59+/-1.69 mg/dL versus 40.75+/-0.86, P <0.01). Vitamin E levels were significantly lower in the Fe group compared to controls (plasma P <0.01; renal tissue P <0.05). Administration of L-Arg to Fe-treated groups prevented these reductions. L-NAME increased iron-induced toxicity significantly, demonstrated by further reduction in the vitamin E levels and renal function compared to the Fe group alone. We concluded that NO plays an important role in protecting the kidney from iron-induced nephrotoxicity. NO synthase blockade enhances iron-mediated renal toxicity in this model.