Na(+)‐HCO3‐ symport in the sheep cardiac Purkinje fibre.

Abstract

1. Intracellular pH (pHi) was recorded in isolated sheep cardiac Purkinje fibres using liquid sensor ion‐selective microelectrodes in conjunction with conventional (3 M‐KCl) microelectrodes (to record membrane potential). 2. In HEPES‐buffered solution (pH0 7.4), pHi recovery from an intracellular acid load (20 mM‐NH4Cl removal) was blocked by 1 mM‐amiloride, consistent with the inhibition of Na(+)‐H+ exchange. Replacement of the HEPES buffer with CO2‐HCO3‐ caused a transient acidosis followed by an amiloride‐resistant recovery of pHi to more alkaline levels (n = 43). This implies the presence of a HCO3(‐)‐dependent pHi regulatory mechanism. 3. Comparison of the membrane potential with the equilibrium potential for HCO3‐ ions (EHCO3) estimated during amiloride‐resistant pHi recovery, showed that for polarized fibres (membrane potential Em approximately ‐80 mV), there was a net outward electrochemical driving force for HCO3‐ ions. Hence the amiloride‐resistant pHi recovery cannot be explained in terms of passive HCO3‐ influx through membrane channels. 4. Removal of external Na+ (Na0+ replaced by N‐methyl‐D‐glucamine) inhibited HCO3(‐)‐dependent pHi recovery, whereas removal of external Cl‐ (leading to depletion of internal Cl‐; Cl0‐ replaced by glucuronate) or short‐term removal of extracellular K+ had no inhibitory effect. We suggest that a Na(+)‐HCO3‐ co‐influx causes the recovery. Replacement of external Na+ with Li+ greatly reduced HCO3(‐)‐dependent pHi recovery indicating that Li0+ cannot readily substitute for Na0+ on the co‐transport. 5. The stilbene drug DIDS (4,4‐diisothiocyano‐stilbene‐disulphonic acid, 500 microM) slowed HCO3(‐)‐dependent pHi recovery. 6. Depolarization of the membrane potential in high K0+ (44.5 mM) solution or with 5 mM‐BaCl2 had no effect upon the rate of HCO3(‐)‐sensitive pHi recovery. This observation, when coupled with the fact that activation of HCO3(‐)‐dependent pHi recovery was associated with no consistent change of membrane potential, suggests that the Na(+)‐HCO3‐ co‐influx is electroneutral and voltage insensitive. 7. HCO3(‐)‐dependent pHi recovery was unaffected by the Na(+)‐K(+)‐2Cl‐ co‐transport inhibitor, bumetanide (150 microM). 8. The contribution of Na(+)‐H+ exchange and Na(+)‐HCO3‐ co‐transport to net acid efflux was assessed. At a pHi of 6.6, we estimate that the co‐transport should account for 20% of total acid equivalent efflux.(ABSTRACT TRUNCATED AT 400 WORDS)