Basolateral membrane Cl-, Na+-, and K+-coupled base transport mechanisms in rat MTALH

Mechanisms involved in basolateral HCO3- transport were examined in the in vitro microperfused rat medullary thick ascending limb of Henle (MTALH) by microfluorometric monitoring of cell pH. Removing peritubular Cl- induced a cellular alkalinization that was inhibited in the presence of peritubular 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and blunted in the absence of external CO2/HCO3-. The alkalinization elicited by removing peritubular Cl- persisted in the bilateral absence of Na+, together with a voltage clamp. When studied in Cl--free solutions, lowering peritubular pH induced a base efflux that was inhibited by peritubular DIDS or by the absence of external CO2/HCO3-. Removing peritubular Na+ elicited a cellular acidification that was accounted for by stimulation of a DIDS- and ethylisopropylamiloride (EIPA)-insensitive Na+/HCO3- cotransport and inhibition of a basolateral Na+/H+ exchange. Increasing bath K+ induced an intracellular alkalinization that was inhibited in the absence of external CO2/HCO3-. At 2 mM, peritubular Ba2+, which inhibits the K+-Cl- cotransport, did not induce any change in transepithelial voltage but elicited a cellular alkalinization and inhibited K+-induced cellular alkalinization, consistent with the presence of a basolateral, electroneutral Ba2+-sensitive K+-Cl- cotransport that may operate as a K+-HCO3- cotransport. This cotransport was inhibited in the peritubular presence of furosemide, [(dihydroindenyl)oxy] alkanoic acid, 5-nitro-2-(3-phenylpropylamino) benzoate, or DIDS. At least three distinct basolateral HCO3- transport mechanisms are functional under physiological conditions: electroneutral Cl-/HCO3- exchange, DIDS- and EIPA-insensitive Na+-HCO3- cotransport, and Ba2+-sensitive electroneutral K+-Cl-(HCO3-) cotransport.