Membrane Transporters and Cytoplasmatic pH Regulation on Bovine Sertoli Cells

Sertoli cells are responsible for regulating a wide range of processes that lead to the differentiation of male germ cells into spermatozoa. Cytoplasmic pH (pH (i) ) has been shown to be an important parameter in cell physiology, regulating namely cell metabolism and differentiation. However, membrane transport mechanisms involved in pH (i) regulation mechanisms of Sertoli cells have not yet been elucidated. In this work, pH (i) was determined using the pH-sensitive fluorescent probe 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF). Addition of weak acids resulted in rapid acidification of the intracellular milieu. Sertoli cells then recovered pH (i) by a mechanism that was shown to be sensitive to external Na+. pH (i) recovery was also greatly reduced in the presence of 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and amiloride. These results point toward the action of an Na+-driven HCO (3) (-) /Cl- exchanger and/or an Na+/HCO (3) (-) cotransporter and the action of the Na+/H+ exchanger on pH (i) regulation in the experimental conditions used. pH (i) recovery was only slightly affected by ouabain, suggesting that the inhibition of Na+/K+-ATPase affects recovery indirectly, possibly via the shift on the Na+ gradient. On the other hand, recovery from the acid load was independent of the presence of concanamycin A, a specific inhibitor of the V-type ATPases, suggesting that these pumps do not have a relevant action on pH (i) regulation in bovine Sertoli cells.