The cytosolic pH (pH(i)) of transitional cells from the ampulla of the
gerbil was measured micro-fluorometrically with the pH-sensitive dye
2',7'-bicarboxyethyl-5(6)-carboxyfluorescein (BCECF) to assess the pos
sible contribution of a Na+/H+ exchanger to the regulation of pH(i). A
ll experiments were conducted in virtually HCO3--free solutions. Under
control conditions, pH(i) was 7.19 and addition of 10(-5) M ethylisop
ropylamiloride (EIPA), a blocker of Na+/H+ exchange, caused a small bu
t significant acidification of pH(i). A transient exposure to 21.4 mM
NH4Cl caused a rapid cytosolic alkalinization followed by a brisk acid
ification and prompt recovery of pH(i) to its control value. The cytos
olic buffer capacity (B(i)) was determined in the absence of Na+ from
changes in pH(i) which were elicited by [NH4+] steps. B(i) was 4.7 mM/
pH at pH(i) 7.19 and varied with pH(i). The initial net proton flux J(
H), representative of Na+/H+ exchange activity, was calculated from th
e product of the initial rate of alkalinization after an NH4+-prepulse
and B(i) at the corresponding pH(i). J(H) was dependent on the extrac
ellular Na+ with an apparent K(m) of 64 mM, sensitive to the cytosolic
[H+] with an apparent K(m) of 2.7 10(-7) M (i.e. pH 6.6), and inhib
ited by EIPA with an IC50 of 5 10(-7) M. These data suggest that tra
nsitional cells contain in the basolateral membrane a Na+/H+ exchanger
of the amiloride-sensitive subtype.