Passive K+-Cl- fluxes in low-K+ sheep erythrocytes: modulation by A23187 and bivalent cations

Abstract

A fraction of the ouabain-resistant (OR) K+ flux of low-K+ (LK) sheep erythrocytes is Cl- dependent (K+-Cl- transport) and is activated reversibly by cell swelling or irreversibly by treatment with N-ethylmaleimide (NEM). The effect of the ionophore A23187 plus bivalent cations (Me2+) or ethyleneglycol-bis(beta-aminoethylether)-N,N'-tetraacetic acid (EGTA) was studied on K+-Cl- transport in control or NEM-treated LK cells. The following observations were made. 1) A23187 (6 microM), at a hematocrit of 10% (vol/vol) and in the presence of 1 mM EGTA, activated severalfold OR K+-Cl- transport in shrunken or swollen cells but failed to stimulate further K+-Cl- flux in NEM-treated cells. 2) In the absence of EGTA, but at very low external Ca2+ concentrations [( Ca2+]o = 10(-7) M), A23187 stimulated OR K+-Cl- flux in controls less than with EGTA and inhibited it slightly in NEM-treated cells. 3) When [Ca2+]o was raised to 10(-3) M, an almost complete inhibition of OR K+-Cl- fluxes occurred in shrunken, swollen, or NEM-treated cells. 4) Other Me2+ inhibited OR K+-Cl- flux in the presence of A23187 in the following order of decreasing potency: Mn2+ much greater than Ca2+ greater than Mg2+ greater than Sr2+ much much greater than Ba2+. 5) Stimulation of OR K+-Cl- flux by A23187 +/- EGTA and inhibition by A23187 + Ca2+ were reversible and did not alter significantly cellular ATP. 6) The stimulatory effect of A23187 plus EGTA, perhaps by Me2+ removal, on K+-Cl- flux and its inhibition by Ca2+ were reversibly abolished in metabolically depleted cells.(ABSTRACT TRUNCATED AT 250 WORDS)