Endogenous K+/H+ exchange activity in the Sf9 insect cell line.
The fluorescent pH indicator 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF) was used to investigate changes in the intracellular pH (pHi) of individual Sf9 cells in response to changes in the composition of the external medium. Under standard conditions, the resting pHi was 0.2-0.3 unit lower than the extracellular pH (6.5). The extracellular concentration of K+ had a major influence on pHi. Removal of K+ from the medium resulted in a rapid but reversible acidification of the cells. The buffer capacity of the cells was a U-shaped function of pHi with a minimum at about pHi = 6.2. In the presence of K+, a change in the pH of the medium was followed by an equivalent change in pHi. In its absence, however, changes in the external pH had little effect on the pH of the cells. Following removal of K+ from the medium, the cells realkalinized at an initial rate which increased with increasing concentration of added K+. This cation was about 30 times more effective in promoting realkalinization of the cells than Li+ Na+, Rb+, and Cs+. The apparent Km for K(+)-dependent H+ efflux was about 12 mM and was slightly modulated by extracellular pH. These results strongly suggest that, in Sf9 cells, a K+/H+ antiporter plays a key role in the movement of protons across the cell membrane.