Abstract
1. Intracellular pH (pHi) of surface fibres of the mouse soleus muscle was measured in vitro by recessed-tip pH-sensitive micro-electrodes. pHi was displaced in an acid direction by removal of external (NH4)2SO4 after a short exposure, and the mechanism of recovery from this acidification was investigated.
2. Removal of external K caused a very slow acidification (probably due to the decreasing Na gradient) but had no effect on the rate of pHi recovery following acidification. This indicates that K+—H+ exchange is not involved in the pHi regulating system.
3. Short applications of 10-4 M ouabain had no obvious effect on pHi and did not alter the rate of pHi recovery following acidification. This suggests that there is no direct connexion between the regulation of pHi and the Na pump.
4. Reduction of external Ca from 10 to 1 mM caused a transient fall in pHi, but the rate of pHi recovery following acidification was unaffected. This suggests that Ca2+—H+ exchange is not involved in the pHi regulating system.
5. An 11% reduction in external Na caused a significant slowing of pHi recovery following acidification. 90% or complete removal of external Na almost stopped pHi recovery. This suggests that Na+—H+ exchange is involved in pHi regulation.
6. Amiloride (10-4 M) reversibly reduced the rate of pHi recovery to much the same extent as removal of external Na. Its effect was not additive to that of removal of external Na.
7. Internal Na ion concentration ([Na+]i), measured using Na+-sensitive micro-electrodes, fell on application of (NH4)2SO4 and increased on its removal. The increase transiently raised [Na+]i above the level recorded before (NH4)2SO4 application. This overshoot of [Na+]i was almost completely inhibited by amiloride. This is consistent with the involvement of Na+—H+ exchange in the pHi regulating system.
8. Removal of external CO2 or application of SITS (10-4 M) caused some slowing of the rate of pHi recovery following acidification by removal of (NH4)2SO4. The effect of SITS was additive to that of Na-free Ringer or amiloride. These results suggest that Cl-—HCO3- exchange is also involved in the pHi regulating system and that it is a separate mechanism. Under the conditions used, Cl-—HCO3- exchange formed about 20% of the pHi regulating system.
9. Decreasing the temperature from 37 to 28 °C not only caused an increase in pHi, but also considerably slowed the rate of pHi recovery following acidification. We have calculated a Q10 for Na+—H+ exchange of 1·4 and for Cl-—HCO3- exchange, 6·9.
10. We conclude that the pHi regulating system is comprised of two separate ionic exchange mechanisms. The major mechanism is Na+—H+ exchange, which is probably driven by the transmembrane Na gradient. The other mechanism is Cl-—HCO3- exchange, which probably requires metabolic energy.
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Selected References
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