An analysis of renal nitric oxide contribution to hyperfiltration in diabetic rats (original) (raw)
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Posttranslational regulation of NO synthase activity in the renal medulla of diabetic rats
AJP: Renal Physiology, 2004
Shear stress increases NO production by endothelial cells, inner medullary collecting duct cells, and thick ascending limb. We postulated that the osmotic diuresis accompanying type 1 diabetes is associated with increased NOS activity and/or expression in the renal medulla. Diabetes was induced by injection of streptozotocin, with insulin provided to maintain moderate hyperglycemia (HYP) or euglycemia (EUG) for 3 weeks. SHAM rats received vehicle treatments. A separate group of rats (PHZ) received phlorizin to produce a glucose-dependent osmotic diuresis. Renal medullary NOS1 and NOS2 activities did not differ between groups, whereas NOS3 activity was significantly increased in HYP. Neither NOS1 nor NOS3 protein levels differed significantly between groups. Reduced phosphorylation of NOS3 at Thr 495 and Ser 633 was evident in medullary homogenates from HYP rats, with no difference apparent at Ser 1177 . Immunohistochemical analysis indicated prominent expression of pThr 495 NOS3 in the thick ascending limb and collecting duct of SHAM and PHZ rats. HYP rats displayed staining in the collecting duct but minimal thick ascending limb staining. Immunostaining with anti-pSer 1177 NOS3 was evident only in the thick ascending limb, with no apparent differences between groups. In summary, glucosedependent osmotic diuresis alone did not alter NOS activity or expression in the renal medulla. Diabetic hyperglycemia increased medullary NOS3 activity without a concomitant increase in NOS3 protein levels; however, NOS3 phosphorylation was reduced at Thr 495 and Ser 633 . Thus, changes in the phosphorylation of NOS at known regulatory sites might represent the primary mechanism underlying increased renal medullary NOS activity in diabetic hyperglycemia.
Evidence for the Existence of Two Distinct Functions for the Inducible NO Synthase in the Rat Kidney
Journal of the American Society of Nephrology, 2002
ABSTRACT. The functional role of the NO synthase (NOS) isoforms in the normal or diseased kidney is uncertain. This study examined the renal expression of the endothelial (eNOS), neuronal (nNOS), and inducible (iNOS) isoforms by both immunohistochemistry and Western blot analyses in sham-operated rats (S) and in rats subjected to 5/6 nephrectomy (Nx). Primary antibodies from two different sources were used to detect iNOS. Additional S and Nx rats were chronically treated with aminoguanidine (AG), a selective iNOS inhibitor. All three isoforms were clearly expressed in S kidney. Their renal abundance, evaluated by Western blot analysis, fell in Nx rats. With the use of anti-iNOS antibodies from two distinct sources, the immunohistochemical analysis showed the presence of what appeared to be two distinct iNOS fractions: a “tubular” fraction, present in S and with decreased intensity in Nx; and an “interstitial” fraction, observed only in inflamed areas of Nx rats. AG treatment greatly...
Nitric Oxide Synthesis Is Reduced in Subjects With Type 2 Diabetes and Nephropathy
Diabetes, 2010
OBJECTIVE-Nitric oxide (NO) is a key metabolic and vascular regulator. Its production is stimulated by insulin. A reduced urinary excretion of NO products (NOx) is frequently found in type 2 diabetes, particularly in association with nephropathy. However, whether the decreased NOx excretion in type 2 diabetes is caused by a defective NOx production from arginine in response to hyperinsulinemia has never been studied.
Diabetes, Obesity and Metabolism, 1999
Objective: Nitric oxide (NO) has been proposed to play a signi®cant role in renal function. In addition, NO production has been found to increase in diabetes mellitus. The present study aimed to clarify the mechanism responsible for NO action in renal function in rats with short (10 days) or prolonged periods (8 weeks) of diabetic induction. Methods: Male Wistar rats were induced to develop diabetes mellitus by intraperitoneal injection of streptozotocin (STZ) (65 mg/kg b.w.), whereas the age-matched control rats were given normal saline. After diabetic induction for 10 days or 8 weeks, the experiment was begun. Three consecutive periods of 30 min each, were designed consisting of one control period, the ®rst and the second period of L-arginine or L-NAME or insulin infusion. Mean arterial pressure (MAP) was determined every 15 min. Arterial blood and urine samples were collected to determine the plasma glucose level (PG), glomerular ®ltration rate (GFR), effective renal plasma¯ow (ERPF), urine¯ow rate (V), urinary protein excretion (Upro), fractional excretion of glucose (FEG) and fractional excretion of sodium (FENa) in each period. Results: No signi®cant differences of MAP were apparent between control rats and rats with diabetic induction. L-arginine infusion had no effect whereas L-NAME markedly increased MAP in normal rats and rats after the short period of diabetes induction. Pressor response to L-NAME in rats exposed to the prolonged period of diabetes induction was lower than that of age-matched control rats. During L-NAME infusion, the PG level signi®cantly declined from 394.9 6 13.1± 338.0 6 14.1 mg/dl and from 399.9 6 7.9±354.3 6 18.8 mg/dl in rats after short and prolonged periods of diabetic induction, respectively. GFR signi®cantly increased whereas ERPF slightly increased in diabetic rats. The elevation of GFR could be reversed by L-NAME or insulin infusion but it increased again after simultaneous infusion of insulin and glucose. Increases in V, the Upro and FEG without changes of FENa, were apparent in diabetic rats. Either L-arginine or L-NAME infusion could not reverse elevations of V, Upro and FEG. The rise of both V and Upro was reversed along with the attenuation of high FEG during insulin infusion, and it rose again close to the diabetic level during simultaneous infusion of insulin and glucose. Elevation of GFR, V and Upro appeared along with a rise of the PG level by » 300±350 mg/ dl in diabetic rats. Conclusions: Both NO and hyperglycaemia are involved in modulating renal hyper®ltration in diabetic rats. The elevations of urine¯ow rate and urinary excretion of both protein and glucose would be expected to represent the reduction of renal tubular reabsorption rather than renal hyper®ltration in diabetic rats. NO does not participate in the change of renal tubular function in diabetic rats. There was a parallel change of urine¯ow rate and urinary excretion of protein in diabetic rats. The rise of the PG level itself would account for the increases of GFR, V, Upro and FEG in diabetic rats. Glomerular hyper®ltration, diuresis and proteinuria in diabetic rats are not exhibited until the PG level rises to » 300±350 mg/ dl.
Nitric oxide in the kidney: Physiological roles and regulation
Physiological roles of nitric oxide (NO) in the kidney include the regulation of renal and glomerular haemodynamics, mediation of pressure natriuresis, maintenance of medullary perfusion, blunting of tubuloglomerular feedback (TGF), inhibition of tubular sodium reabsorption and modulation of renal sympathetic nerve activity. The net effect of NO in the kidney is to promote natriuresis and diuresis and NO plays an important role in the renal adaptation to variations of dietary salt intake. Reduction of NO production in the renal medulla, either pharmacologically or genetically, has a great impact on the delivery of blood to the medulla and on the long-term regulation of sodium excretion and blood pressure. Conversely, infusion of L-arginine to increase nitric oxide, abrogates hypertension and enhances medullary blood flow. It is evident that medullary NO production serves as an important counterregulatory factor to buffer vasoconstrictor hormone-induced reduction of medullary blood f...
Nitric oxide in ischaemic acute renal failure of streptozotocin diabetic rats
Diabetologia, 1996
Changes in nitric oxide (NO) levels were determined in ischaemic acute renal failure in streptozotocin-induced diabetes mellitus rats. Two weeks after streptozotocin administration and immediately after right nephrectomy, the left renal artery was occluded for 60 rain. Similar procedures were carried out in non-diabetic rats. The nitrite (NOz) + nitrate (NO3) levels were measured in plasma and urine. The effects of chronic oral supplementation with Larginine and an NO synthase inhibitor (N-omega-nitro-L-arginine) were also studied in both diabetic and non-diabetic rats before and after renal artery clamping. The rats with diabetic acute renal failure had a much lower creatinine clearance (90 _+ 22 ~tl 9 min-~ 9 100 g body weight-1, p < 0.005), and higher fractional excretion of sodium (FENa)% (10.90+ 4.2, p < 0.001) and protein excretion (2078 + 69 ~tg/ ml creatinine clearance, p < 0.001) compared with the respective values in the non-diabetic groups (163 +_ 30; 1.46 + 86; 453.3 + 31). The plasma and urine NO 2 + NO 3 levels were significantly higher in the untreated diabetic rats compared with the untreated normal rats before ischaemia (p < 0.001). The ischaemic acute renal failure in non-diabetic rats increased the plasma and urinary NO 2 + NO 3 excretion after ischaemia. The urinary excretion of these metabolites decreased significantly and their plasma levels remained unchanged in the ischaemic diabetic rats. The rc-arginine administration resulted in a small but significantly higher creatinine clearance after clamping in the non-diabetic rats. The NO synthase inhibitor caused deterioration in renal function in all ischaemic and non-ischaemic groups. In summary, the greater vulnerability to ischaemia of the diabetic kidney seems to be associated with both impaired response to and impaired production of NO. [Dia
Renal and systemic nitric oxide synthesis in rats with renal mass reduction
Kidney International, 1997
Renal and systemic nitric oxide synthesis in rats with renal mass reduction. In rats undergoing renal mass reduction (RMR) oral supplementation with the nitric oxide (NO) precursor L-arginine increases glomerular filtration rate and ameliorates signs of glomerular injury, suggesting that chronic renal failure in the rats is a condition of low NO formation in the kidney. On the contrary, data are available that in the systemic circulation of uremics, both rats and human beings, NO is formed in excessive amounts and may contribute to platelet dysfunction and bleeding tendency, well-known complications of uremia. The present study was designed to clarify the pathophysiology of renal and systemic NO synthesis in uremia. We showed that renal er vivo NO generation, measured as the conversion of [3H] L-arginine to [3H} L-citrulline, was lower than normal in RMR rats, seven days after surgery, and progressively worsened with time in close correlation with signs of renal injury. Consistent with these results, urinary excretion of the stable NO metabolites, NO2 /N03, significantly decreased in rats with RMR. To go deeper into the cellular origin and biochemical nature of this abnormality we used two histochemical approaches that could locate either NO synthase (NOS) catalytic activity (NADPH-diaphorase) or NOS isoenzyme expression (immunoperoxidase). NADPH-diaphorase documented a progressive loss of renal NOS activity in RMR rats that co-localized with a strong progressive decrease of inducible NOS isoenzyme (iNOS) immunostaining. At variance with iNOS, endothelial cell NOS (ecNOS) staining was rather comparable in RMR and control kidneys. At variance to the kidney, in the systemic circulation of RMR rats the synthesis of NO increased as reflected by higher than normal plasma NO2 7NO3 concentrations. High systemic NO likely derives from vessels as documented by the increased NOS activity and higher expression of both iNOS and ecNOS in the aorta of RMR rats. Up-regulation of systemic NO synthesis might be an early defense mechanism against hypertension of uremia. On the other hand, more NO available to circulating cells may sustain the bleeding tendency, a well-known complication of uremia.
Renal vasoconstriction during inhibition of NO synthase: Effects of dietary salt
Kidney International, 1994
Renal vasoconstriction during inhibition of NO synthase: Effects of dietary salt. Since dietary salt loading enhances nitric oxide (NO) generation in the kidney, we investigated the hypothesis that changes in salt intake have specific effects on vascular resistance in the kidney mediated by the L-arginine-NO pathway. We contrasted changes in renal and hindquarter vascular resistances (RVR and HQVR) in anesthetized rats during intravenous infusions of graded doses of the NO synthase inhibitor N°-nitro-L-arginine methyl ester (L-NAME). Groups (N = 8 to 10) of rats were maintained on a high salt (HS) or low salt (LS) diet for two weeks. Compared to those on LS, rats on HS had a greater increase in mean arterial pressure (AMAP; +32 4 vs. +22 3%; P 0.05) and RVR (+ 160 17 vs. +83 10%; P < 0.005) and a greater fall in renal blood flow (zRBF;-47 3 vs.-32 4%; P < 0.01); changes in HQVR were similar in the two groups. The enhanced RVR response to L-NAME in HS rats could not be ascribed to the higher renal perfusion pressure (RPP) since it persisted in rats whose RPP was controlled by adjustment of a suprarenal aortic clamp. Changes in RVR with an NO donor (SIN-i) were similar in HS and LS rats. L-NAME reduced plasma renin activity in both HS and LS rats. After inhibition of ACE with captopril, or of angiotensin II type I (AT1) receptor with losartan, the increase in RVR with L-NAME remained greater in HS than LS rats. In conclusion, an increase in dietary salt potentiates the renal vascular response to L-NAME. This effect is specific for the kidney and cannot be ascribed to changes in NO responsiveness or RPP or to effects of Ang II generation or action on AT1 receptors. Generation of nitric oxide (NO) from L-arginine by nitric oxide synthase (NOS) leads to vasorelaxation of renal and systemic blood vessels [1-4]. Micropuncture and microperfusion studies in vivo have disclosed a functional role for NO as a signalling molecule between the macula densa (MD) and the afferent arteriole [5]. This MD-derived NO vasodilates the afferent arteriole and counteracts vasoconstrictor stimuli generated by the tubuloglomerular feedback (TGF) response [5]. Dietary salt loading enhances the renal production of NO, as indexed by the excretion of NO2 and NO3 and cyclic guanosine monophosphate (cGMP) [6]. Since TGF is a specific renal response, the first aim of these studies was to examine the hypothesis that there is a specific renal vasodilator pathway mediated by L-arginine-derived NO that is activated by dietary salt loading. Some investigators have shown that increases in renal vascular