Intracellular pH and its regulation in Pelvetia zygotes

We have investigated the role of intracellular pH (pHi) in early plant development using ion-selective microelectrodes to record from eggs and embryos of the brown alga Pelvetia. Temporal changes in pHi were investigated by recording from zygotes at all stages of the first cell cycle. pHi was 7.57 +/- 0.09 in recently fertilized eggs, but decreased by approximately 0.2 units a few hours postfertilization. Proton motive force (pmf) was also monitored and found to be less than -50 mV throughout the first cell cycle. Because of the low pmf values, we suggest that secondary active transport is probably not coupled to H+, and instead we propose that solute transport is driven by the Na+ electrochemical potential. Zygotes strictly regulated pHi over a wide range of extracellular pH (pHo); pHi varied by less than 0.2 units over pHo values from 6.2 to 9.2. Inhibitor studies were conducted to investigate the mechanism of regulation. Agents known to inhibit the H(+)-ATPase (antimycin A, KCN, carbonylcyanide m-chlorophenylhydrazone, N,N'-dicyclohexylcarbodiimide, and erythrosin B) caused marked cytoplasmic acidification. Addition of amiloride, an inhibitor of Na+/H+ antiport, also resulted in acidification, as did removal of NaCl from the medium. By contrast, increasing extracellular NaCl concentration caused transient alkalinization of the cytoplasm. Taken together, these results indicate that proton pumping by an H(+)-ATPase and Na+/H+ antiporter contribute to pH regulation.