Intracellular pH alterations induced by tacrine in a rat liver biliary epithelial cell line

Abstract

1. The effects of tacrine (THA) on intracellular pH (pH(i)) were examined in a rat liver biliary epithelial cell line (RLEC) in HEPES-buffered medium. pH(i) was recorded using the pH-sensitive fluoroprobe, carboxy-SNARF-1 (carboxy-seminaphtorhodafluor). 2. In the steady state, short-term exposures to THA resulted in alkalinization and re-acidification at 0.1 and 0.25 mM. Following a 24 h-treatment, no significant difference in pH(i) could be detected at 0.1 and 0.25 mM THA, whereas at 0.05 mM, pH(i) was slightly more acid (7.17 ± 0.02, n = 16 versus 7.21 ± 0.02, n = 24 [control]). 3. In control and short-term treated cells, intracellular intrinsic buffering power (β(i)) increased roughly linearly as pH(i) decreased. This dependence was not seen following long-term treatment. In all cases, β(i) was increased by THA (by 1.6 to 3.5 fold). 4. Following an acid load (induced by 20 mM NH4Cl removal), pH(i) recovery in RLEC relied upon Na+/H+ exchange. A short-term treatment (0.25 mM THA) did not affect total acid extrusion. In contrast, a 24 h-treatment with 0.05 mM THA reduced it (by ≃ 36% at a pH(i) of 6.73) while at 0.25 mM, a large increase was detected (by ≃ 109% at a pH(i) of 6.75). 5. In Na+-free medium, THA (0.25 mM) still induced an alkalinization in the steady state. Following an acid load, THA stimulated a Na+-independent acid efflux in a dose-dependent manner, inhibitable by α-cyano-4-hydroxy cinnamate (CHC, 4 mM) but not by quercetin (0.125 mM). 6. In conclusion, this work demonstrates that THA affects pH(i) in RLEC, through a decrease in Na+/H+ exchange and an increase in β(i). Stimulation of a CHC-inhibitable, Na+-independent acid efflux is also detected.