Changes in intracellular pH and pH regulating mechanisms in somitic cells of the early chick embryo: a study using fluorescent pH-sensitive dye

Abstract

1. Measurements of intracellular pH (pHi) have been made using the fluorescent dye dicyano-hydroquinone (DCH) in somites isolated from 2-day-old chick embryos. Measurements of pHi, from freshly dissected somites, gave values of 7.18 +/- 0.02 (mean +/- S.D., n = 12) while in somites kept in media containing 10% fetal calf serum for 2-5 h pHi was 7.36 +/- 0.02 (n = 11). 2. In freshly dissected control tissue recovery from an acid load (NH4Cl pre-pulse), in the nominal absence of HCO3-, was 0.007 +/- 0.003 pH units/min (n = 11). In the presence of 5 mM-HCO3- the rate of recovery was increased to 0.132 +/- 0.003 pH units/min (n = 11). This HCO3(-)-dependent recovery was inhibited by pre-treatment with DIDS (5 X 10(-4) M) and stopped when Na+ was replaced in the bathing medium with K+ or N-methylglucamine. Amiloride (10(-3) M) had no effect. 3. Replacement of Cl- with gluconate had little effect on pHi in control somites suggesting that absence of Cl(-)-HCO3- exchange. 4. These observations are consistent with the presence of a coupled Na+-HCO3- entry and the absence of Na+-H+ exchange. 5. In serum-treated cells recovery from an acid load was 0.101 +/- 0.01 pH units/min (n = 11) in the nominal absence of HCO3-. Increasing the HCO3- concentration (5-10 mM) or pre-treatment with DIDS had no effect on the rate of acid extrusion. Recovery is Na+ dependent and inhibitable with amiloride, indicating the presence of Na+-H+ exchange. 6. These results suggest that during somitic development the mechanisms regulating pHi and recovery from acid loading are modified from a Na+-HCO3- influx to a Na+-H+ exchanger. This transition allows the cells to increase pHi by up to 0.2 pH units and may have a role in somitic cell development.