Regulation of intracellular pH and water movement by the Cl/HCO3 exchanger in RCCD1 cell line

RIVAROLA, VALERIA; FORD, PAULA; CHARA, OSVALDO; PARISI, MARIO; CAPURRO, CLAUDIA

Buenos Aires - Argentina

Congreso; XIV International Biophysics Congress; 2002

The aim of this work was to characterize the regulation of intracelular pH (pHi) and water movement by the Cl-/HCO3- exchanger (AE) in RCCD1 cell line. In order to study this we used the pH sensitive dye, BCECF. Apical and basolateral Cl- replacement led to a gradual and reversible stable rise in pHi (D pHi = 0,2 ± 0,05, n=4, p<0,05). This alkalinization was inhibited by DIDS 100 m M. HCO3- replacement caused a transitory alkalinization (D pHi = 0,27 ± 0,03, n=7, p<0,001) whose acidification kinetic was partially inhibited by DIDS (43%) and completely blocked by acetazolamide 100 m M. Basolateral Cl- replacement induced an alkalinization similar to the observed with apical and basolateral replacement of Cl- (D pHi-basolat= 0,22 ± 0,03, n=5, p<0,001). This was not the case with apical Cl- replacement that produce no change in pHi. These results suggests expression of a basolateral AE in RCCD1 cell line. We compared water fluxes studies with pHi measures to see if the AE was related to water movement. Replacement of Cl- and HCO3- induced a diminution of basal water secretion (54%) while producing an alkalinization (D pHi= 0.20 ± 0.07, n=6, p<0.01). These results suggests that water secretion is Cl- and HCO3- dependent. Finally RT-PCR studies showed the expression of AE2 isoform in RCCD1 cell line.D pHi = 0,2 ± 0,05, n=4, p<0,05). This alkalinization was inhibited by DIDS 100 m M. HCO3- replacement caused a transitory alkalinization (D pHi = 0,27 ± 0,03, n=7, p<0,001) whose acidification kinetic was partially inhibited by DIDS (43%) and completely blocked by acetazolamide 100 m M. Basolateral Cl- replacement induced an alkalinization similar to the observed with apical and basolateral replacement of Cl- (D pHi-basolat= 0,22 ± 0,03, n=5, p<0,001). This was not the case with apical Cl- replacement that produce no change in pHi. These results suggests expression of a basolateral AE in RCCD1 cell line. We compared water fluxes studies with pHi measures to see if the AE was related to water movement. Replacement of Cl- and HCO3- induced a diminution of basal water secretion (54%) while producing an alkalinization (D pHi= 0.20 ± 0.07, n=6, p<0.01). These results suggests that water secretion is Cl- and HCO3- dependent. Finally RT-PCR studies showed the expression of AE2 isoform in RCCD1 cell line.