Increased expression of renal aquaporin water channels in spontaneously hypertensive rats.

AIMS: The present study was aimed to determine whether there exists an altered regulation of aquaporin (AQP) water channels in hypertension. METHODS: Male spontaneously hypertensive rats (SHR) aged 10-12 weeks were used. Age-matched Wistar-Kyoto (WKY) rats served as control. The abundance of AQP1-4 proteins in the kidney was determined by Western blot analysis. The protein expression and activity of adenylyl cyclase were also determined. RESULTS: The medullary expression of AQP2 and AQP3 proteins was increased in SHR compared with that in WKY rats. The expression of AQP1 protein was also significantly increased in the inner medulla, while that of AQP4 was not. Immunohistochemistry of AQP2 revealed that principal cells of the collecting duct have strong immunoreactivity, the degree of which was augmented with prominent apical labeling in SHR. The plasma level of arginine vasopressin (AVP) was higher in SHR; the adenylyl cyclase activity stimulated by AVP was augmented, along with increased expression of type VI adenylyl cyclase. The urine was more concentrated with its volume decreased in SHR. CONCLUSION: The expression of AQP1-3 channels is increased in the kidney, in association with enhanced activity of the AVP/cAMP pathway, in SHR.

Indomethacin enhances shuttling of aquaporin-2 despite decreased abundance in rat kidney.

The effect of nonsteroidal antiinflammatory drugs on the regulation of aquaporin-2 (AQP2) water channels in the kidney was determined. Male Sprague-Dawley rats were injected with indomethacin (5 mg/kg twice a day intraperitoneally) for 2 d. The control group was injected with vehicle. The expression of AQP2 proteins was determined in the kidney by immunoblotting and immunohistochemistry. The expression of G(salpha) and type VI adenylyl cyclase was determined by immunoblotting. The activity of adenylyl cyclase complexes was determined by stimulated accumulation of cAMP. Immunoblotting revealed that indomethacin markedly decreased the expression of AQP2. Accordingly, however, the ratio of AQP2 expression in the membrane fraction versus that in the cytoplasmic fraction was increased. The urinary excretion of AQP2 proteins also increased. Immunohistochemistry demonstrated almost exclusive apical labeling of AQP2 with scanty cytoplasmic localization along the collecting duct. The expression of G(salpha) and adenylyl cyclase VI proteins was decreased. The generation of cAMP provoked by arginine vasopressin, sodium fluoride, or forskolin was blunted. These results suggest that indomethacin increases the shuttling of AQP2 while it decreases its abundance in the collecting duct.

Indomethacin decreases particulate guanylyl cyclase activity in rat kidney.

1. Effects of non-steroidal anti-inflammatory drugs on the local atrial natriuretic peptide (ANP) and nitric oxide (NO) systems in the kidney were investigated. 2. Male Sprague-Dawley rats were treated with indomethacin (5 mg/kg, every 12 h, i.p.) for 2 days. The expression of ANP and natriuretic peptide receptor-A (NPR-A) mRNA was determined in the kidney, as was that of endothelial NO synthase (NOS) proteins. Particulate and soluble guanylyl cyclase activities were determined separately. 3. Following treatment with indomethacin, urinary sodium excretion decreased significantly. Although the renal expression of ANP was not changed significantly, that of NPR-A decreased significantly. The expression of NOS increased significantly. Particulate guanylyl cyclase activity was decreased, whereas the activity of soluble guanylyl cyclase was increased. The catalytic activity of Na(+)/K(+)-ATPase was increased, with no significant changes in its expression. The expression of the type 3 Na/H exchanger and Na-K-2CL cotransporters increased significantly. 4. The indomethacin-induced decrease in urinary sodium excretion may be attributed, at least in part, to decreased activity of the local ANP/cGMP system. The increased activity of the NO/cGMP system may be a compensatory response to the diminished activity of the prostaglandin system.

Diminished expression of sodium transporters in the ureteral obstructed kidney in rats.

BACKGROUND/AIMS: Whether the postobstructive natriuresis and diuresis is related with an altered regulation of sodium transporters in the kidney was examined. METHODS: Male Sprague-Dawley rats underwent either bilateral (BUO) or unilateral obstruction (UUO) of the proximal ureters for 24 h. The expression of Na,K-ATPase, type-3 sodium-hydrogen exchanger (NHE3), type-1 bumetanide-sensitive sodium cotransporter (BSC1), and thiazide-sensitive sodium cotransporter (TSC) proteins was determined in the obstructed kidney by Western blot analysis and immunohistochemistry. Catalytic activity of Na,K-ATPase was also determined. RESULTS: The expression of alpha1 and beta1 subunit proteins and the catalytic activity of Na,K-ATPase were significantly decreased in the obstructed kidney in BUO. The expressions of NHE3, BSC1 and TSC proteins were also significantly decreased. Immunohistochemistry confirmed the downregulation of these sodium transporters in the obstructed kidney. In UUO, the expression of sodium transporters was similarly decreased in the obstructed kidney. CONCLUSION: The postobstructive natriuresis and diuresis may in part be accounted for by a reduced abundance of sodium transporters in the kidney.

Long-term treatment with cyclosporine decreases aquaporins and urea transporters in the rat kidney.

The aim of this study was to evaluate the long-term effects of cyclosporine (CsA) treatment on urinary concentration ability. Rats were treated daily for 4 wk with vehicle (VH; olive oil, 1 ml/kg sc) or CsA (15 mg/kg sc). The influence of CsA on the kidney's ability to concentrate urine was evaluated using functional parameters and expression of aquaporins (AQP1-4) and of urea transporters (UT-A-1-3, and UT-B). Plasma vasopressin levels and the associated signal pathway were evaluated, and the effect of vasopressin infusion on urine concentration was observed in VH- and CsA-treated rats. Toxic effects of CsA on tubular cells in the medulla as well as the cortex were evaluated with aldose reductase (AR), Na-K-ATPase-alpha(1) expression, and by determining the number of terminal transferase-mediated dUTP nick end-labeling (TUNEL)-positive cells. Long-term CsA treatment increased urine volume and fractional excretion of sodium and decreased urine osmolality and free-water reabsorption compared with VH-treated rats. These functional changes were accompanied by decreases in the expression of AQP (1-4) and UT (UT-A2, -A3, and UT-B), although there was no change in AQP2 in the cortex and outer medulla and UT-A1 in the inner medulla (IM). Plasma vasopressin levels were not significantly different between two groups, but infusion of vasopressin restored CsA-induced impairment of urine concentration. cAMP levels and Gsalpha protein expression were significantly reduced in CsA-treated rat kidneys compared with VH-treated rat kidneys. CsA treatment decreased the expression of AR and Na-K-ATPase-alpha(1) and increased the number of TUNEL-positive renal tubular cells in both the cortex and medulla. Moreover, the number of TUNEL-positive cells correlated with AQP2 or UT-A3) expression within the IM. In conclusion, CsA treatment impairs urine-concentrating ability by decreasing AQP and UT expression. Apoptotic cell death within the IM at least partially accounts for the CsA-induced urinary concentration defect.

Upregulation of vascular renin-angiotensin and endothelin systems in rats inhibited of nitric oxide synthesis.

The present study was aimed at investigating whether the regulation of vascular renin-angiotensin and endothelin (ET) systems is altered by a chronic blockade of nitric oxide (NO) synthesis. Male Sprague-Dawley rats were supplemented with N(G)-nitro-L-arginine methyl ester (L-NAME, 100mgl(-1)) in drinking water for 4 weeks to inhibit the endogenous synthesis of NO. The mRNA expressions of renin, angiotensin converting enzyme (ACE), type-1 angiotensin II receptor (AT1R), ET-1, type-A ET receptor (ET(A)), and neutral endopeptidase (NEP) were determined in the thoracic aorta by reverse transcription-polymerase chain reaction. The treatment with L-NAME significantly increased the blood pressure, while it decreased the tissue levels of nitrite/nitrate. The mRNA expression of renin, ACE, and AT1R was increased in the aorta. The protein expression of AT1R assessed by Western blot analysis was also increased. The expression of ET-1 and ET(A) mRNA was increased, whereas that of NEP mRNA decreased. The increased expression of renin-angiotensin and ET system genes and the decreased expression of NEP may in part be causally related with the development of hypertension induced by a chronic blockade of NO synthesis.

Caffeine decreases the expression of Na+/K+-ATPase and the type 3 Na+/H+ exchanger in rat kidney.

1. The aim of the present study was to explore the mechanisms underlying the renal effects of caffeine. 2. Male Sprague-Dawley rats were treated with caffeine, consisting of a single oral bolus (0.2%, 20 mL/kg) followed by supplementation in drinking water (0.2%) for 1 day. Rats treated the same but given water without caffeine served as controls. 3. The expression of alpha1- and beta1-subunits of Na+/K+-ATPase, the type 3 Na+/H+ exchanger (NHE3) and aquaporin-1 was determined in the kidney by western blot analysis. 4. To explore possible involvement of local humoral mediators, the tissue expression of nitric oxide synthase (NOS) proteins was determined by western blot analysis and the expression of atrial natriuretic peptide (ANP) mRNA was determined by semiquantitative reverse transcription-polymerase chain reaction. 5. Following treatment with caffeine, the expression of alpha1- and beta1-subunits of Na+/K+-ATPase, as well as that of NHE3, was decreased. Accordingly, the catalytic activity of Na+/K+-ATPase was decreased. In contrast, the expression of aquaporin-1 was not altered significantly. 6. The expression of the endothelial isoform of NOS was increased, along with tissue nitrite/nitrate levels. The expression of ANP mRNA was increased. 7. It is suggested that caffeine decreases Na+/K+-ATPase and NHE3 activities and increases nitric oxide and ANP activities in the kidney.

Altered expression of vascular natriuretic peptide receptors in experimental hypertensive rats.

1. The aim of the present study was to determine whether the regulation of vascular natriuretic peptide receptors (NPR) is related to the local renin-angiotensin system (RAS). 2. Male Sprague-Dawley rats were made two-kidney, one-clip (2K1C) and deoxycorticosterone acetate (DOCA)-salt hypertensive to activate and inhibit the RAS, respectively. Another model of hypertension was induced by treatment with an inhibitor of nitric oxide synthesis, namely NG-nitro-L-arginine methyl ester (L-NAME). 3. The mRNA expression of NPR-A, NPR-C, angiotensin- converting enzyme (ACE) and angiotensin AT1 receptors was determined in the thoracic aorta by semiquantitative reverse transcription-polymerase chain reaction. The particulate guanylyl cyclase activity stimulated by atrial natriuretic peptide (ANP) was also determined in the membrane fraction of the thoracic aorta. 4. The plasma concentrations of ANP were increased significantly in the three models of hypertension. Plasma renin activity was increased in 2K1C hypertension, decreased in DOCA-salt hypertension and not significantly altered in L-NAME hypertension. 5. The mRNA expression of NPR-A and NPR-C was decreased, whereas that of ACE and AT1 receptors was increased in 2K1C and L-NAME hypertension. The mRNA expression of NPR-A and NPR-C was increased, whereas that of ACE and AT1 receptors was decreased in DOCA-salt hypertension. 6. The particulate guanylyl cyclase activity was decreased in 2K1C and L-NAME hypertension and increased in DOCA-salt hypertension. 7. The vascular expression of NPR may be reciprocally regulated by local RAS activity.