INTRACELLULAR PH INCREASE DRIVEN BY AN NA+ H+ EXCHANGER UPON ACTIVATION OF SURF CLAM OOCYTES/

Intracellular pH (pH(i)) measurements were performed in surf clam (Spi sula solidissima) oocytes before and after artificial activation or fe rtilization [evidenced by germinal vesicle breakdown (GVBD)] by the di methyloxazolidinedione (DMO) and -bis-(2-carboxyethyl)-5-(and-6)-carbo xyfluorescein (BCECF) methods. Results using both methods showed incre ases of pH(i) of 0.3 pH unit after activation by excess K+. Using BCEC F, we found an increase of similar magnitude after fertilization or af ter the addition of serotonin. By contrast, GVBD did not occur when th e pH(i) was increased to similar or even higher levels by exposing the oocytes to ammonia. In sodium-free seawater, excess K+ induced GVBD b ut the pH(i) of K+-activated oocytes decreased significantly below the resting level of unactivated oocytes. The pH(i) increases in K+-activ ated oocytes were otherwise proportional to the external Na+ concentra tion. The amiloride derivatives dimethylamiloride and hexamethylene am iloride (at 10-50 mu M) efficiently inhibited the K+-induced increase of pH(i) but did not block GVBD. These two derivatives were able, howe ver, 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 pH(i) increase, driven by a typical Na+/H+ exchanger, foll ows activation of surf clam oocytes but that this pH(i) increase is ne ither sufficient nor required for GVBD, though it does allow its progr ession at an optimal rate. (C) 1997 Academic Press.