Apical membrane permeability and kinetic properties of the sodium pump in rabbit urinary bladder.

Abstract

SUMMARY1. Previous studies have shown that aldosterone stimulates the rate of Na+ transport across the rabbit urinary bladder epithelium by increasing the apical membrane permeability to Na+. Paradoxically, ion-sensitive and conventional micro-electrode measurements demonstrated that intracellular Na+ activity aNa+ was essentially unchanged by aldosterone, i.e. aka+ was constant regardless of the rate of Na+ transport. The present study was designed to resolve this apparent contradiction.2. The effects of elevated, endogenous aldosterone levels produced by low-Na+ diet (Lewis & Diamond, 1976) on urinary bladder Na+ transport were investigated in vitro using Ussing-type chambers and intracellular conventional and ion-sensitive microelectrodes. Apical membrane selectivity and the kinetics of the Na+ pump were assessed as a function of hormone stimulation.3. The aldosterone-stimulated increase in Na+ transport was accounted for by increases in both the relative selective permeability of the apical membrane to Na+ and an increase in its absolute Na+ permeability.4. The kinetics of the Na+ pump were evaluated electrically by loading the cells with Na+ (monitored with Na+-sensitive micro-electrodes) or alternatively by manipulating serosal solution K+ concentration and measuring changes in the basolateral membrane electromotive forces and resistance. From these measurements the current generated by the pump was calculated as a function of intracellular Na+ or extracellular K+. The kinetics of the pump were not altered by aldosterone. A model of highly co-operative binding estimated Km for Na+ as 14-2 mm and 2-3 mm for K+. Hill coefficients for these ions were 2-8 and 1-8, respectively, consistent with a pump stoichiometry of 3 Na+ to 2 K+. The kinetic properties of the Na-K pump indicate that physiological levels of aka+ are poised at the foot of a step kinetic curve which energetically favours Na+ extrusion.

Electrogenic sodium absorption in rabbit cecum in vitro.

“…Although there is considerable evidence for in vitro pharmacologic modification of Na channels, there is also some precedent for in vivo alterations of this transport system (27,28). Along these lines, Leng-Peschlow and Marty (29) have observed that, in a surgically created cecal pouch, pretreatment of the pouch with cecal contents over several days resulted in net absorption of Na and water.…”

Defining protein expression in the urothelium: a problem of more than transitional interest

“…Perhaps the best example of functional characterization of a newly discovered protein in the urothelium was the amiloridesensitive sodium channel, which is present in the luminal membrane (41,(43)(44). Employing Ussing chambers and conventional as well as ion-sensitive microelectrodes to measure transepithelial resistance, conductance, short-circuit currents, and intracellular Na ϩ activity, the authors dissected the functional and regulatory signature of an ion channel which was ultimately cloned and shown to be the heteromeric epithelial sodium channel (ENaC) (8).…”

Regulation of Na channels of the rat cortical collecting tubule by aldosterone.

“…A primary action of the hormone is the increase in Na permeability of the apical membrane of these cells, leading to increased Na reabsorption. This has been demonstrated both with electrical measurements on intact epithelia (Nagel and Crabb6, 1980;Palmer, Li, Lindemann, and Edelman, 1982;Lewis and Wills, 1983;Sansom and O'Neil, 1985) and with Na fluxes into isolated membrane vesicles (Asher and Garty, 1988). There is, however, relatively little known about the action of aldosterone at the single channel level.…”