Impairment of Na/K-ATPase Signaling in Renal Proximal Tubule Contributes to Dahl Salt-sensitive Hypertension

Abstract

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/KATPase. 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.The Dahl salt-resistant (R) and -sensitive (S) strains were developed by selective breeding of the outbred SpragueDawley rat strain for resistance or susceptibility to the hypertensive effects of high dietary sodium (1). Whereas the blood pressure (BP) 2 response to salt-loading in Dahl R and S rats involves many regulatory factors (2), it has been proposed that renal proximal tubule (RPT) Na ϩ handling may be a critical determinant of the different BP responses in these strains (3, 4). Cardiotonic steroids (CTS) such as ouabain and marinobufagenin (MBG) appear to be involved in the regulation of BP and renal Na ϩ handling in vivo (5-7) as well as the ion handling of both primary cultures and proximal tubular cell lines in vitro (7-12). Based on this background, we performed the following studies to examine whether the proximal tubules of Dahl R and S rats had different responses to CTS. MATERIALS AND METHODSAnimals-Male Sprague-Dawley rats (290 -310 g body weight) were purchased from Charles River. Male, agematched Dahl R (SS/JrHsd) and S (SR/JrHsd) rats were bred and maintained in-house and used at the age of 12-14 weeks. Rats were maintained with 12 h dark/light cycle and had ad libitum access to the food (Teklad Laboratory animal diets from Harlan Laboratories) and water. All animal experimentation was conducted in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals using protocols approved by the University of Toledo Institutional Animal...

Identification of a Mutant α1 Na/K-ATPase That Pumps but Is Defective in Signal Transduction

“…Although their pathophysiological significance has been a subject of debate for many years (29), several gene replacement studies from Lingrel and co-workers (30,31) have unequivocally demonstrated an important role of endogenous CTS in regulation of renal Na ϩ excretion and blood pressure. In addition, we and others showed that CTS not only induced hypertension in rats but also caused significant cardiovascular remodeling independent of their effect on blood pressure (5,20,31,32). Moreover, CTS may play an important role in embryonic development (19).…”

“…This pumping function is essential for eukaryotic cells to maintain ionic homeostasis as well as to provide transmembrane Na ϩ gradients for the Na ϩ -dependent transport of nutrients. Recent studies from different laboratories have revealed that the ␣1 Na/K-ATPase also has important signal transduction functions and could act as a functional receptor for cardiotonic steroids to activate protein kinase cascades (2)(3)(4), which play an important role in renal salt handling and remodeling of the heart and the kidney under pathological conditions (5). Using in vitro binding assays, we have identified two pairs of domain interactions that seem to be essential for the formation of this functional receptor.…”

Alteration of Tight Junction Protein Expression in Dahl Salt-Sensitive Rat Kidney

“…A mutation in the α1 Na/K-ATPase gene, leading to higher sodium reabsorption and SS hypertension, was identified in Dahl SS rats [20, 21], but its existence was later denied by other researchers [22]. The contributory role of the proximal tubule Na + /H + exchanger type 3 was also unclear in Dahl SS rat kidneys [23, 24]. Although Hoagland et al reported that the thick ascending limb Na + -K + -2Cl - cotransporter was upregulated in Dahl SS rat kidneys, the control animals used were not Dahl SR rats, but rather salt-resistant brown Norway rats [25].…”

Renal mechanisms of salt-sensitive hypertension: contribution of two steroid receptor-associated pathways

“…The last fraction is reabsorbed via epithelial Na ϩ channels (ENaC) in the CNT/CD. Na ϩ /H ϩ exchanger (NHE)3 in proximal tubules (62,68) and NKCC2 in the thick ascending limb (3,10,66) are associated with salt-sensitive hypertension in animal models. Salt-induced NHE3 suppression is more impaired in Dahl salt-sensitive rats than in Dahl salt-resistant rats (68).…”

“…Na ϩ /H ϩ exchanger (NHE)3 in proximal tubules (62,68) and NKCC2 in the thick ascending limb (3,10,66) are associated with salt-sensitive hypertension in animal models. Salt-induced NHE3 suppression is more impaired in Dahl salt-sensitive rats than in Dahl salt-resistant rats (68). NKCC2 activity in thick ascending limb is also upregulated in salt-sensitive rats (3,10,66).…”