Mechanism underlying flow stimulation of sodium absorption in the mammalian collecting duct

Abstract

. Mechanism underlying flow stimulation of sodium absorption in the mammalian collecting duct. Am J Physiol Renal Physiol 291: F663-F669, 2006. First published April 25, 2006 doi:10.1152/ajprenal.00514.2005.-Vectorial Na ϩ absorption across the aldosterone-sensitive distal nephron plays a key role in the regulation of extracellular fluid volume and blood pressure. Within this nephron segment, Na ϩ diffuses from the urinary fluid into principal cells through an apical, amiloride-sensitive, epithelial Na ϩ channel (ENaC), which is considered to be the rate-limiting step for Na ϩ absorption. We have reported that increases in tubular flow rate in microperfused rabbit cortical collecting ducts (CCDs) lead to increases in net Na ϩ absorption and that increases in laminar shear stress activate ENaC expressed in oocytes by increasing channel open probability. We therefore examined whether flow stimulates net Na ϩ absorption (JNa) in CCDs by increasing channel open probability or by increasing the number of channels at the apical membrane. Both baseline and flow-stimulated J Na in CCDs were mediated by ENaC, as J Na was inhibited by benzamil. Flow-dependent increases in JNa were observed following treatment of tubules with reagents that altered membrane trafficking by disrupting microtubules (colchicine) or Golgi (brefeldin A). Furthermore, reducing luminal Ca 2ϩ concentration ([Ca 2ϩ ]) or chelating intracellular [Ca 2ϩ ] with BAPTA did not prevent the flow-dependent increase in J Na. Extracellular trypsin has been shown to activate ENaC by increasing channel open probability, and we observed that trypsin significantly enhanced J Na when tubules were perfused at a slow flow rate. However, trypsin did not further enhance J Na in CCDs perfused at fast flow rates. Similarly, the shear-induced increase in benzamil-sensitive J Na in oocytes expressing protease resistance ENaC mutants was similar to that of controls. Our results suggest the rise in J Na accompanying increases in luminal flow rates reflects an increase in channel open probability. epithelial sodium channel; in vitro microperfusion; protein trafficking; mechanoregulation; laminar shear; principal cell THE DISTAL CONVOLUTED TUBULE (DCT), connecting tubule (CNT), and collecting duct (CD) contribute to the final regulation of renal Na ϩ reabsorption (13-16, 21, 24, 26, 28, 29, 31, 32), a process that plays a key role in modifying extracellular fluid volume and blood pressure. Within the rabbit cortical collecting duct (CCD), a segment that has been utilized extensively for functional analysis by in vitro microperfusion, Na ϩ absorption is considered to be electrogenic and mediated by Na ϩ diffusion from the urinary fluid into the cell through the apical amiloride-sensitive epithelial Na ϩ channel (ENaC).We (32, 33) and others (13,27,38) previously reported that increases in tubular fluid flow rate stimulate net Na ϩ absorption in the mammalian CCD. We speculated that hydrodynamic forces associated with increases in urinary flow rate either directly activate...

Pendrin gene ablation alters ENaC subcellular distribution and open probability

“…The ␣-and ␥-subunits are fully active following two cleavage events, which reduce the apparent mobility of the ␣-subunit from 85 to 65 kDa (5) and the ␥-subunit from 93 to 70 kDa (5). These ␣-and ␥-ENaC cleavage events stimulate ENaC channel activity through increased channel open probability (5,6,23). Since pendrin gene ablation in aldosterone-treated mice reduces the abundance of the 70-kDa more than the 85-kDa ␥-ENaC fragment (15), aldosterone-induced ␥-ENaC cleavage may be blunted in the mutant mice, which should reduce ENaC channel open probability.…”

Role of NKCC in BK channel-mediated net K+secretion in the CCD

“…K ϩ secretion within these segments requires a favorable electrochemical gradient (a high cell K ϩ concentration and an apical membrane voltage that is depolarized with regard to the Nernst potential for K ϩ ), generated by apical Na ϩ entry through the epithelial Na ϩ channel (ENaC) and its electrogenic Na ϩ -K ϩ -ATPase-mediated basolateral extrusion in exchange for the uptake of K ϩ , as well as an apical membrane permeability to K ϩ . In animals fed a normal-K ϩ diet, K ϩ secretion in the cortical collecting duct (CCD) is completely dependent on Na ϩ absorption (14,43,68).…”

“…This response reflects, at least in part, an increase in delivery to and reabsorption of Na ϩ by the distal nephron (27,43,64), which in turn increases the driving force for passive K ϩ efflux across the apical membrane. Two types of apical K ϩ channels have been functionally identified in the distal nephron: the inwardly rectifying ATPsensitive secretory K ϩ (SK) channel encoded by ROMK (15,16,75), and a high-conductance Ca 2ϩ -and/or stretch-activated BK channel (32,46,63), considered to be comprised of a pore-forming ␣-subunit, a member of the slo family of K ϩ channels originally cloned from Drosophila, and a regulatory ␤-subunit (1,10,30).…”

Trypsin can activate the epithelial sodium channel (ENaC) in microdissected mouse distal nephron

“…Recently, some functional evidence is emerging that ENaC activation by extracellular proteases can indeed occur in native renal tissue (17). In microperfusion studies, it was observed that trypsin treatment enhanced net sodium absorption in isolated rabbit cortical collecting ducts (CCD), at least at slow tubular fluid perfusion rates (36). Moreover, it has been reported that exposure to luminal TK increases the intracellular sodium concentration ([Na ϩ ] i ) in principal cells of isolated and microperfused mouse CCD.…”