A dual mechanism for intracellular pH regulation by leech neurones

Neutral-carrier pH-sensitive micro-electrodes were used to investigate intracellular pH (pHi) in leech neurones. When used in snail neurones such electrodes gave very similar pHi values to those recorded simultaneously by recessed-tip glass micro-electrodes. Leech Retzius neurones superfused with a pH 7.4 HCO3--free physiological saline were found to have a pHi of 7.3, too high to be explained by a passive distribution of H+ or OH-. To investigate pHi regulation the pHi was decreased by one of three methods: by exposure to propionate, by adding and then removing NH4Cl or by exposure to CO2. Acidification by any method was followed by a recovery to normal pHi values within 15-20 min. In HCO3--free solutions, pHi recovery from acidification was blocked by removing external Na or by amiloride (2 mM). In solutions buffered with 2% CO2 and 11 mM-HCO3-, amiloride slowed but did not block pHi recovery. The anion exchange inhibitor SITS (4-acetamido-4'-isothiocyanato-stilbene-2,2'-disulphonic acid) also slowed pHi recovery in the presence of HCO3-. In CO2/HCO3- solution the removal of external Na either slowed or blocked pHi recovery, and blocked it completely in the presence of amiloride. We conclude that in HCO3--free solutions pHi regulation is by a Na-H exchange system; but in the presence of HCO3- there is an additional mechanism which is probably a Na-dependent Cl-HCO3 exchanger.