CELLULAR DISTRIBUTION OF THE RENAL BUMETANIDE‐SENSITIVE Na–K–2Cl COTRANSPORTER BSC‐1 IN THE INNER STRIPE OF THE OUTER MEDULLA DURING THE DEVELOPMENT OF HYPERTENSION IN THE SPONTANEOUSLY HYPERTENSIVE RAT (original) (raw)
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Increased Expression of the Sodium Transporter BSC-1 in Spontaneously Hypertensive Rats
Journal of Pharmacology and Experimental Therapeutics, 2004
The purpose of this study was to compare the expression of BSC-1 (bumetanide-sensitive Na +-K +-2Clcotransporter) in kidneys of spontaneously hypertensive (SHR) versus Wistar-Kyoto (WKY) rats by immunoblotting and RT-PCR. To determine the specificity of any observed changes in BSC-1 expression, we also compared expression of the thiazide sensitive Na +-Clcotransporter (TSC), the type-3 Na +-H + exchanger (NHE-3), Na +-K +-ATPase-α 1 , the inwardly rectifying K + channel (ROMK-1), the type-1 Na +-HCO 3-cotransporter (NBC-1), aquaporin-1 and aquaporin-2. Analyses were performed on outer cortex, outer medulla and inner medulla. BSC-1 protein was detected in outer medulla and was markedly (6-fold) higher in SHR. TSC protein was detected in the cortex and was not over-expressed in SHR. Aquaporin-1 protein was detected in all three regions and was not over-expressed in SHR. Aquaporin-2 and ROMK-1 proteins were detected in all three regions, but were moderately elevated (2-fold) only in SHR inner medulla. Na +-K +-ATPase and NHE-3 proteins were detected in all three regions. Na +-K +-ATPase-α 1 was modestly (25%) increased in SHR outer and inner medulla, while NHE-3 was moderately (2-fold) increased in the SHR cortex and inner medulla. NBC-1 protein was detected only in the cortex, and was higher in SHR (2-fold). mRNA levels of BSC-1, aquaporin-2 and ROMK-1 were not elevated in SHR, indicating a post-translational mechanism of protein over-expression. High-dose furosemide increased fractional sodium excretion more in SHR than WKY (3-fold). We conclude that increased expression of BSC-1, and to a lesser extent, aquaporin-2, ROMK-1, NHE-3 and NBC-1, may contribute to the pathogenesis of hypertension in the SHR.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 1990
, and (C) Sabra salt-sensitive hypertensive (SBED and salt-resistant (SBN). Initially, Na+-dependent furosemide-or bumetanide-inhibited aSRb+ fluxes were characterised using Wistar rat microsomes. The latter were partially purified on a metrizamide cushion, and assay conditions were optimized for use with microsomes from the other rats. The major result is that in microsomes from adult Milan hypertensive (MILS) rats the rate of the Na÷/K+/CI--cotransporter mediated s6 Rb flux at sub-saturating concentrations of Rb, appears to be significantly greater than in the normotensive (MNS) controls. The effect reflects an increased apparent Rb affinity of the cotransporter in MHS microsomes. There is no difference in maximal rate or in the apparent Na ÷ activation affinity of the aSRb+ flux. In addition bumetanide appears to be a somewhat more effective inhibitor in MHS compared to MNS microsomes. The aSRb+ flux result is compatible with a previous finding that in red cells, Na+/K+-cotransporter mediated fluxes are increased in MHS compared to MNS. It supports the notion that the Na+/K+/CI--cotransporter in both red cells and kidney is a genetic marker for hypertension. It is of interest that apparently more than one Na ÷ transport system is affected in MIlS hypertensive kidneys (a) the Na÷/K+/C! -cotransporter in the thick ascending limb of Heule and (b) the Na+/H ÷ exchanger and/or a conductive Na÷-pathway in brush-border membranes from proximal tubule. It is conceivable that in the hypertensive animals a common regulatory pathway (e.g., phosphorylation) or protein (e.g., cytoskeleton) is affected along the length of the nephron. In Sabra SBH and SBN rat microsomes, no difference was found for the aSRb+ flux via the Na+/K+/CI-cotransporter (or via a K + channel).
AJP: Renal Physiology, 2008
We investigated which of the NaCl transporters are involved in the maintenance of salt sensitive hypertension. Milan hypertensive (MHS) rats were studied 3 months after birth. In MHS, as compared to normotensive strain (MNS), mRNA abundance, quantified by competitive PCR on isolated tubules, was unchanged both for Na + -H + isoform 3 (NHE3) and Na + -K + -2Cl -(NKCC2), but higher (119%, n=5, p<0.005) for Na + -Cl -(NCC) in distal convoluted tubules (DCT). These results were confirmed by Western blots which revealed: the renal tubule cells to reabsorb sodium (13). Therefore the importance of the Na + -K + -ATPase in the pathogenesis of hypertension is fully established (3).
The regulation of proximal tubular salt transport in hypertension: an update
Current Opinion in Nephrology and Hypertension, 2009
Purpose of review-Renal proximal tubular sodium reabsorption is regulated by sodium transporters, including the sodium glucose transporter, sodium amino acid transporter, sodium hydrogen exchanger isoform 3 and sodium phosphate cotransporter type 2 located at the luminal/ apical membrane, and sodium bicarbonate cotransporter and Na + /K + ATPase located at the basolateral membrane. This review summarizes recent studies on sodium transporters that play a major role in the increase in blood pressure in essential/polygenic hypertension. Recent findings-Sodium transporters and Na + /K + ATPase are segregated in membrane lipid and nonlipid raft microdomains that regulate their activities and trafficking via cytoskeletal proteins. The increase in renal proximal tubule ion transport in polygenic hypertension is primarily due to increased activity of NHE3 and Cl/HCO 3 exchanger at the luminal/apical membrane and a primary or secondary increase in Na + /K + ATPase activity.
Sodium Transport in Membrane Vesicles from Kidney of the Milan Hypertensive Rat Strain
Annals of the New York Academy of Sciences, 1986
Sodium uptake was evaluated in brush border membrane vesicles prepared from kidney cortex of rats of the Milan hypertensive strain (MHS)' during the prehypertensive (70 g animals) and hypertensive (200 g animals) stages. Control measurements were carried out with the corresponding Milan normotensive strain (MNS). The vesicles were prepared by differential centrifugation following precipitation of contaminating membranes with MgC12.2 Uptake of "Na was measured by a rapid filtration technique3 in the following intravesicular (in) and extravesicular (out) pH conditions:
American journal of physiology. Renal physiology, 2003
The effect of ANG II treatment of rats for 7 days was examined with respect to the abundance and subcellular localization of key thick ascending limb (TAL) Na+ transporters. Rats were on a fixed intake of Na+ and water and treated with 0, 12.5, 25, 50 (ANG II-50), 100 (ANG II-100), and 200 (ANG II-200) ng x min(-1) x kg(-1) ANG II (sc). Semiquantitative immunoblotting revealed that Na+/H+ exchanger 3 (NHE3) abundance in the inner stripe of the outer medulla (ISOM) of ANG II-treated rats was significantly increased: 179 +/- 28 (ANG II-50, n = 5), 166 +/- 23 (ANG II-100, n = 7), and 167 +/- 19% (ANG II-200, n = 4) of control levels (n = 6, P < 0.05), whereas lower doses of ANG II were ineffective. The abundance of the bumetanide-sensitive Na(+)-K(+)-2Cl(-) cotransporter (BSC-1) in the ISOM was also increased to 187 +/- 28 (ANG II-50), 162 +/- 23 (ANG II-100), and 166 +/- 19% (ANG II-200) of control levels (P < 0.05), but there were no changes in the abundance of Na(+)-K(+)-ATPas...
Increased Na pump activity in the kidney cortex of the Milan hypertensive rat strain
FEBS Letters, 1991
The (Na',K')-ATPase activity from the kidney cortex of the Milan hypertensive rat strain (MHS) and the corresponding normotensive control (MNS) was measured both in active solubilized enzyme preparations and in isolated basolateral membrane vesicles. Kineticanalysis of the purified enzyme showed that the V,,,,, value was significantly higher in MHS rats. The difference between MHS and MNS was not linked to a different number of sodium pumps, but was related to the molecular activity of the enzyme. Using basolateral membrane vesicles, an increased ATPdependent ouabain-sensitive sodium transport was also demonstrated in MHS rats. These results support the hypothesis that a higher tubular sodium reabsorption may be involved in the pathogenesis of hypertension in this rat strain.
Reversible effects of acute hypertension on proximal tubule sodium transporters
American Journal of Physiology-Cell Physiology, 1998
Acute hypertension provokes a rapid decrease in proximal tubule sodium reabsorption with a decrease in basolateral membrane sodium-potassium-ATPase activity and an increase in the density of membranes containing apical membrane sodium/hydrogen exchangers (NHE3) [Y. Zhang, A. K. Mircheff, C. B. Hensley, C. E. Magyar, D. G. Warnock, R. Chambrey, K.-P. Yip, D. J. Marsh, N.-H. Holstein-Rathlou, and A. A. McDonough. Am. J. Physiol.270 ( Renal Fluid Electrolyte Physiol.39): F1004–F1014, 1996]. To determine the reversibility and specificity of these responses, rats were subjected to 1) elevation of blood pressure (BP) of 50 mmHg for 5 min, 2) restoration of normotension after the first protocol, or 3) sham operation. Systolic hypertension increased urine output and endogenous lithium clearance three- to fivefold within 5 min, but these returned to basal levels only 15 min after BP was restored. Renal cortex lysate was fractionated on sorbitol gradients. Basolateral membrane sodium-potassiu...
Journal of Biological Chemistry, 2011
We have observed that, in renal proximal tubular cells, cardiotonic steroids such as ouabain in vitro signal through Na/K-ATPase, which results in inhibition of transepithelial 22 Na ؉ transport by redistributing Na/K-ATPase and NHE3. In the present study, we investigate the role of Na/K-ATPase signaling in renal sodium excretion and blood pressure regulation in vivo. In Sprague-Dawley rats, high salt diet activated c-Src and induced redistribution of Na/K-ATPase and NHE3 in renal proximal tubules. In Dahl salt sensitive (S) and resistant (R) rats given high dietary salt, we found different effects on blood pressure but, more interestingly, different effects on renal salt handling. These differences could be explained by different signaling through the proximal tubular Na/K-ATPase. Specifically, in Dahl R rats, high salt diet significantly stimulated phosphorylation of c-Src and ERK1/2, reduced Na/K-ATPase activity and NHE3 activity, and caused redistribution of Na/K-ATPase and NHE3. In contrast, these adaptations were either much less effective or not seen in the Dahl S rats. We also studied the primary culture of renal proximal tubule isolated from Dahl S and R rats fed a low salt diet. In this system, ouabain induced Na/K-ATPase/c-Src signaling and redistribution of Na/K-ATPase and NHE3 in the Dahl R rats, but not in the Dahl S rats. Our data suggested that impairment of Na/K-ATPase signaling and consequent regulation of Na/K-ATPase and NHE3 in renal proximal tubule may contribute to salt-induced hypertension in the Dahl S rat. . 2 The abbreviations used are: BP, blood pressure; CTS, cardiotonic steroids; EE, early endosome; ERK1/2, extracellular signal-regulated kinases 1 and 2; MBG, marinobufagenin; NHE3, sodium/hydrogen exchanger isoform 3; RPT, renal proximal tubule; HS, high salt diet; LS, low salt diet.
American Journal of Physiology-Renal Physiology, 1998
A bumetanide-sensitive Na-K-2Cl cotransporter, BSC-1, is believed to mediate the apical component of transcellular NaCl absorption in the thick ascending limb (TAL) of Henle’s loop. To study its ultrastructural localization in kidney, we used an affinity-purified, peptide-derived polyclonal antibody against rat BSC-1. Immunoblots from rat kidney cortex and outer medulla revealed a solitary 161-kDa band in membrane fractions. Immunocytochemistry of 1-μm cryosections demonstrated strong BSC-1 labeling of the apical and subapical regions of medullary and cortical TAL cells. Notably, macula densa cells also exhibited distinct labeling. Distal convoluted tubules and other renal tubule segments were unlabeled. Immunoelectron microscopy demonstrated that BSC-1 labeling was associated with the apical plasma membrane and with subapical intracellular vesicles in medullary and cortical TAL and in macula densa cells. Smooth-surfaced TAL cells, in particular, had extensive BSC-1 labeling of intr...