Increased renal glomerular endothelin-1 release in gentamicin-induced nephrotoxicity.

Gentamicin-induced acute renal failure is characterized by a decrease in renal plasma flow and creatinine clearance. Endothelins (ET) are potent renal vasoconstrictors. The aim of this work is to assess the role of ET-1 in gentamicin-induced renal failure. Renal glomerular release of ET-1 was measured in rats with gentamicin-induced nephrotoxicity (100 mg/kg/day, s.c. for 2, 4 or 6 days). Glomeruli were isolated and incubated for 24 h in RPMI-1640. Glomerular supernatant and plasma concentration of ET-1 were measured by RIA. Renal failure was assessed by insulin, para-aminohippuric and creatinine clearance and histological studies. Gentamicin induced a dose number-dependent increase in plasma creatinine and a decrease in creatinine clearance. This was accompanied by a marked decrease in inulin and para-aminohippuric acid clearance, as well as by a marked tubular necrosis, without alterations in glomerular structures. Plasma ET-1 concentration and glomerular ET-1 release were also increased in gentamicin-treated rats. When 10-5 M gentamicin was added to control glomeruli, ET-1 production was not modified (36.4 +/- 2.2 vs. 35.2 +/- 1.7 pg/ml/24 h). All these results suggest that elevated ET-1 plasma levels and increased glomerular release of ET-1 could mediate, at least in part, the decrease in glomerular filtration rate observed in gentamicin-induced ARF.

Gentamicin treatment increases mesangial cell nitric oxide production.

Gentamicin-induced decreases in glomerular filtration rate have been associated with a marked decline in the glomerular capillary ultrafiltration coefficient which could be mediated by mesangial cell contraction. We have assessed a possible role of endogenous nitric oxide (NO) as a modulator of the proliferative and contractile effects of gentamicin on mesangial cells. NO synthesis and release, measured as nitrite production, were increased in the presence of gentamicin; this increase was blocked by coincubation with L-NAME. Mesangial cells treated with gentamicin, but not cells under control conditions, expressed mac-iNOS mRNA and presented positive immunoreactivity for mac-iNOS. Gentamicin induced a reduction of the planar surface area of cultured rat mesangial cells; cell treatment with gentamicin plus L-arginine significantly decreased the contractile effect of gentamicin. Gentamicin increased both [3H]thymidine incorporation into DNA and viable cell number; when L-arginine was added together with gentamicin, this abolished the effect of gentamicin on mesangial cell proliferation. The present studies demonstrate that gentamicin induces the expression of mac-iNOS and produces contraction and proliferation in mesangial cells. These actions seem to be modulated by mesangial NO synthesis and release.

Effect of nitric oxide synthesis modification on renal function in gentamicin-induced nephrotoxicity.

We evaluated the effect of acute or chronic nitric oxide (NO) synthesis activation or inhibition in rats with gentamicin-induced acute renal failure. Rats received gentamicin 100 mg/kg per day for 6 days, or isotonic saline. Some animals of each group also received N(G)-monomethyl-l-arginine (l-NAME, 4 mg/kg per day) or l-arginine (1%) in the drinking water for 6 days (chronic NO synthesis modification). In another experimental set, animals were treated with gentamicin or saline for 6 days and glomerular filtration rate (GFR) and renal plasma flow (RPF) were measured before and after the infusion of l-NAME (50 mg/h per kg) or l-arginine (60 mg/h per kg) (acute NO synthesis modification). Acute l-NAME administration induced a decrease in GFR and RPF both in control and gentamicin treated animals. Chronic l-NAME treatment induced an impairment in GFR only in gentamicin-treated animals. Acute l-arginine administration did not modify renal function in any experimental group whereas chronic l-arginine administration improved renal function only in gentamicin-treated animals. Urinary excretion of N-acetyl-ß-d-glucosaminidase and alkaline phosphatase was increased by chronic treatment with l-NAME in both groups, whereas l-arginine had no effect. In conclusion, NO synthesis inhibition aggravates gentamicin-induced renal damage. However, chronic NO synthesis stimulation partially prevents against gentamicin nephrotoxicity, thus suggesting that increased renal NO synthesis during gentamicin-induced nephrotoxicity plays a protector role on renal function.

Effects of the beta 3-adrenoceptor agonist BRL 37344 on lipomobilization and plasma glucose levels in conscious fasted rabbits.

The existence of a functional beta 3-adrenoceptor in conscious 24 h fasted rabbits was investigated by studying the effect of BRL 37344 (a beta 3-adrenoceptor agonist) in comparison with CGP 12177 (a beta 1- and beta 2-adrenoceptor antagonist, which also acts as a partial beta 3 agonist) and isoproterenol (a nonselective beta 1-, beta 2-, and beta 3-adrenoceptor agonist) on plasma nonesterified fatty acids (NEFA) and its possible relation with glucose metabolism. All drugs were intravenously infused at the same dose, 0.3 microgram.kg-1.min-1 (30 min), BRL 37344, CGP 12177, and isoproterenol significantly elevated plasma NEFA levels without increasing plasma glucose levels. Isoproterenol elevated plasma insulin and lactate levels, but BRL 37344 and CGP 12177 did not. Bupranolol (0.1 mg/kg, subcutaneously) blocked the BRL 37344 mediated effect on plasma NEFA levels. In the presence of CGP 20712A (1 mg/kg, subcutaneously), ICI 118551 (30 micrograms/kg, subcutaneously), or the mixture of both selective beta-antagonists, BRL 37344 also evoked an increase in plasma NEFA levels, but the response was less pronounced when compared with BRL 37344 alone. These data suggest that in conscious rabbits deprived of food for 24 h (i) BRL 37344 induces lipomobilization through beta 3-adrenoceptor stimuli, but the effect is less than by beta 1- and beta 2-adrenoceptor stimuli, and (ii) an excess of plasma NEFA evoked by BRL 37344 may not necessarily contribute to plasma glucose increases.

Increased severity of gentamicin nephrotoxicity in aging rats is mediated by a reduced glomerular nitric oxide production.

The present experiments have been performed to assess whether the increased severity of gentamicin-induced nephrotoxicity in old animals could be mediated by a decreased production of nitric oxide. Aging rats (12 months) treated with gentamicin showed higher plasma creatinine and a higher reduction in creatinine clearance. After gentamicin treatment, glomerular nitrite production was higher in young than in old animals, whereas no differences in cortical gentamicin concentration were observed between young and aging animals. The increased severity of gentamicin-induced acute renal failure in old animals could be based on a decreased glomerular NO production after gentamicin treatment.

Glomerular angiotensin II receptors in gentamicin-induced renal failure in the rat.

We evaluated the properties of glomerular angiotensin II receptors in renal glomeruli isolated from control rats and from rats with gentamicin-induced renal failure. There were no differences in the affinity of angiotensin II for its receptor between glomeruli from control and those from rats treated with gentamicin. Angiotensin II receptor density was lower in glomeruli from rats with renal failure than in those from control rats (985 +/- 71 in gentamicin treated rats vs. 1602 +/- 213 fmol/mg prot in controls). No significant differences were observed in renin activity in the supernatant from glomeruli isolated from control rats (3.74 +/- 0.29 ng angiotensin l/mL h) and those isolated from rats with gentamicin-induced renal failure (2.99 +/- 0.29 ng angiotensin l/mL h, p > 0.1). These findings do not support the contention of a role of angiotensin II in the development and maintenance of gentamicin-induced ARF.

Effect of felodipine on systemic hemodynamics of spontaneous mild-hypertensive aged rats.

The aim of the present study was to examine the effects of felodipine, a dihydropyridinic calcium ions channels blocker, on mean arterial blood pressure (MAP), cardiac output (CO), peripheral resistances (TPR) and blood flow distribution in spontaneously mildly hypertensive female Wistar 30-34 months old rats. Under pentobarbital anesthesia, CO and regional organ blood flow were measured by the radioactive microspheres method, before and 30 min after administration of felodipine 0.5 mumol/kg b. w. by gastric gavage. They were compared to the corresponding values in normotensive rats of the same strain and age. Fifteen (from twenty five) rats were hypertensive with a MAP averaging 139 +/- 2 mm Hg. CO and TPR were slightly higher in these hypertensive rats. Cerebral blood flow (CBF) was lower, though the difference did not reach significant values. MAP significantly decreased after felodipine with no significant changes in CO and TPR in hypertensive as well as in normotensive animals. Renal blood flow (RBF) was similar before and after felodipine which significantly decreased renal vascular resistance in both groups. Felodipine administration did not induce significant changes in CBF but a significant increase in portal venous inflow (PVI) in hypertensive rats only. In conclusion, in old female rats with mild spontaneously hypertension, acute felodipine oral administration reduced arterial blood pressure without diminishing CBF.

Nitric oxide production by mononuclear leukocytes in alcoholic cirrhosis.

Nitric oxide is now established as a biological mediator of clinical relevance. The present study investigated the production of nitric oxide by lympho-mononuclear leukocytes from alcoholic patients with either hepatitis or cirrhosis. The study included 42 patients, 12 without any liver disease and 30 alcoholic patients, 13 of whom had histologically confirmed cirrhosis and 17 alcoholic hepatitis. Cells were obtained from peripheral blood by density gradient and incubated in sterile conditions in RPMI 1640 for 6 h at 37 degrees C. Culture supernatants were assayed for nitrite concentration using the Griess reaction. Cells from cirrhotic but not from hepatopathic patients showed significantly higher nitrite production than controls (cirrhotic, 0.36 +/- 0.07; hepatopathic, 0.13 +/- 0.02; control: 0.25 +/- 0.05 nmol/10(6) cells/6 h). In cirrhotic patients L-Nitro-arginine methylester inhibited nitrite production (0.18 +/- 0.05). These data suggest that alcoholic cirrhotic but nonhepatopathic patients show an increased nitric oxide production by blood lymphomononuclear cells. This production could be involved in the systemic vasodilation in cirrhotic patients.

Effect of NG-nitro-L-arginine methyl ester on nephrotoxicity induced by gentamicin in rats.

The present experiments were designed to assess the effect of inhibiting NO synthesis on the renal failure induced in rats by treatment with high doses of gentamicin. Eighteen Wistar rats were given gentamicin 100 mg/kg body weight/day for 5 days, whereas another 18 rats were used as control. Half of the gentamicin-treated rats and half of the controls also received the specific inhibitor of NO synthesis, NG-nitro-L-arginine methyl ester (L-NAME), 0.05 mg/ml in the drinking water for 5 days. Arterial pressure and renal function were measured on the 5th day of the study. In the animals treated with L-NAME, arterial pressure was higher than in untreated rats, thus suggesting that the treatment was effective in inhibiting NO synthesis. Rats that received L-NAME and gentamicin, showed higher plasma creatinine levels and higher score of renal damage, as well as lower Na+ and K+ excretion and creatinine clearance than rats that received gentamicin alone. These data showing that NO inhibition aggravates gentamicin-induced renal failure, suggest that endogenously released NO plays a protective role in gentamicin nephrotoxicity.

Increased glomerular nitric oxide synthesis in gentamicin-induced renal failure.

Renal glomeruli isolated from normal rats and from rats with gentamicin-induced renal failure were incubated with substances that modify nitric oxide (NO) synthesis and the resulting changes in glomerular cyclic GMP (cGMP) levels were measured by radioimmunoassay. Glomeruli from normal and gentamicin-treated rats contained 0.17 +/- 0.04 and 2.02 +/- 0.63 pmol cGMP/mg protein respectively. In normal glomeruli, acetylcholine and bradykinin significantly increased cGMP levels whereas NG-nitro-L-arginine-methyl ester, an inhibitor of NO synthesis, completely blocked this increase. In glomeruli from gentamicin-treated animals, acetylcholine and bradykinin also stimulated cGMP accumulation, while NG-nitro-L-arginine-methyl ester decreased the cGMP content to levels significantly below the basal concentrations. These data suggest an increased glomerular production of NO in rats with gentamicin-induced renal failure.

Effect of gentamicin treatment on glutamine and lactate metabolism by the renal cortex of the rat.

The present study was performed to measure the uptake of main renal cortical fuel substrates (glutamine and lactate) and the release of the main renal cortical products (ammonia and glucose) by cortical slices from gentamicin-treated rats. Experiments were done in 2 groups of female Wistar rats (250 g): In gentamicin group (n = 13), rats were injected s.c. with gentamicin-sulphate 100 mg/Kg body wt/day for 5 days. Control rats (n = 13) received isotonic saline. After anesthesia and blood sampling, renal cortical slices were obtained and incubated with L-glutamine and/or lactate at 1 or 5 mM concentration, containing L-glutamate and/or pyruvate at 0.1 or 0.5 mM. Creatinine clearance was reduced to a 50% in gentamicin-treated rats. In addition these animals showed a sharp increase in urinary excretion of N-acetyl-beta-D-glucosaminidase and alkaline phosphatase. Light microscopy examination revealed extensive cell necrosis and tubular obstruction of the proximal tubules in kidneys of rats injected with gentamicin. The renal cortical gentamicin concentration of rats injected with gentamicin was 310 +/- 43 mu/g, whereas it was undetectable in control rats. Cortical slices from gentamicin-treated rats, compared to control ones, showed a reduced production of ammonia and glucose, without differences in glutamine or lactate extraction. These alterations can be explained by both the increased rate of anabolic reactions to recover cell damage associated to renal failure, as well as by a direct effect of gentamicin on the rate of carboxylation reactions.