Intracellular pH Increase Driven by an Na+/H+Exchanger upon Activation of Surf Clam Oocytes

Abstract

Intracellular pH (pHi) measurements were performed in surf clam (Spisula solidissima) oocytes before and after artificial activation or fertilization [evidenced by germinal vesicle breakdown (GVBD)] by the dimethyloxazolidinedione (DMO) and 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF) methods. Results using both methods showed increases of pHiof 0.3 pH unit after activation by excess K+. Using BCECF, we found an increase of similar magnitude after fertilization or after the addition of serotonin. By contrast, GVBD did not occur when the pHiwas increased to similar or even higher levels by exposing the oocytes to ammonia. In sodium-free seawater, excess K+induced GVBD but the pHiof K+-activated oocytes decreased significantly below the resting level of unactivated oocytes. The pHiincreases in K+-activated oocytes were otherwise proportional to the external Na+concentration. The amiloride derivatives dimethylamiloride and hexamethylene amiloride (at 10–50 μM) efficiently inhibited the K+-induced increase of pHibut did not block GVBD. These two derivatives were able, however, to retard K+-induced GVBD, hexamethylene amiloride being the more efficient. This retardation of K+-induced GVBD could be abolished by the simultaneous addition of ammonia. Taken altogether, these results show that a pHiincrease, driven by a typical Na+/H+exchanger, follows activation of surf clam oocytes but that this pHiincrease is neither sufficient nor required for GVBD, though it does allow its progression at an optimal rate.