We have investigated the influence of the H+ and Na+ gradients across the cell membrane on the regulation of the intracellular pH (pHi) and of the intracellular Na activity (aNai) in sheep heart Purkinje fibres, using Na+- and pH-sensitive microelectrodes. 2. In oxygenated, nominally bicarbonate-free solutions (buffered with HEPES) the steady-state pHi changed linearly with the extracellular pH (pHo) by 0.23 pH units/pHo unit change over the pHo range of 5.4-8.4. The H+ equilibrium potential changed by about 47 mV/pHo unit change. 3. Both the steady-state pHi and the pHi recovery from acidification induced by lowering the pHo to 6.4 were affected only to a small extent by reducing the extracellular Na concentration [Na]o, to one half or to one tenth normal. 4. The steady-state aNai decreased by 5 to 20% when the pHo was reduced to 6.4 and increased by 3 to 8% when the pHo was raised to 8.4. These changes in aNai were still present when the Na-K pump had been inhibited by the cardioactive steroid strophanthidin (10(-5) M). 5. Exposure to K-free solutions caused an increase in aNai. Following addition of 6 mM-K (to re-activate the Na-K pump) the rate of decrease of aNai was not affected by pHo changes from 6.4 to 8.4. 6. Inhibition of the Na+-K+ pump by strophanthidin (10(-5) M) caused aNai to rise rapidly within 2-3 min. The pHi remained unchanged for the first 1-30 min after the pump inhibition, but then decreased by several tenths of a pH unit. 7. Amiloride (10(-3) M) caused a small decrease in a Nai and an intracellular acidification of up to 0.2 pH units. 8. Under conditions, where aNai was high due to inhibition of the Na-K pump by strophanthidin, lowering the [Na]o to one tenth normal produced a very large intracellular acidification, while aNai decreased. Amiloride increased this intracellular acidification even more, while the decrease in aNai remained unaffected. 9. Application of NH4Cl (20 mM) produced a decrease of aNai and a rapid intracellular alkalinization, followed by a slower acidification. Upon removal of NH4Cl the pHi dropped by several tenths of a pH unit but rapidly recovered. During this pH recovery there was a small transient increase in aNai above the control level before returning to normal. 10. The pHi recovery after the removal of NH4Cl was slowed by lowering the [Na]o to one tenth normal, and it was greatly inhibited in the presence of amiloride. The transient overshoot of aNai after NH4Cl removal was suppressed by amiloride. 11. We conclude that under some conditions there appears to be an exchange of intracellular H+ for extracellular Na+ across the cell membrane and that this exchange can help regulate the intracellular pH.
