Acidification of extracellular environment inhibits CCK-8-induced Ca2+ entry into pancreatic acinar cells of rat evaluated by a Mn(2+)-quenching method.
The fluorescence of fura 2, a Ca(2+)-sensitive dye, is quenched in the presence of other divalent cations such as Mn2+. This characteristic of the dye provides a useful measure for investigating the role of Ca2+ in cellular functions. The present experiments were thus designed to examine the effects of extracellular pH (pHo) on cholecystokinin octapeptide (CCK-8)-induced Ca2+ entry into isolated perifused pancreatic acini of the rat using a Mn(2+)-quenching method. At a standard pHo (7.4), addition of Mn2+ (1 mM) during continuous stimulation with 100 pM CCK-8 quenched fura 2 fluorescence excited by both excitation wavelengths of 340 and 380 nm. Lowering pHo to 6.0 weakened the quenching, whereas elevating pHo to 8.0 during the stimulation accelerated the quenching compared with that at standard pHo. Accordingly, when acinar cells were stimulated continuously with 100 pM CCK-8 at pHo 6.0, the second sustained increase in cytosolic Ca2+ concentration ([Ca2+]i) was decreased compared with that at pHo 7.4, but the [Ca2+]i increase reappeared when the pHo was neutralized from 6.0 to 7.4 even after cessation of the stimulation. The second sustained increase of [Ca2+]i was achieved at pHo 8.0, but it declined rapidly when CCK-8 stimulation was terminated and pHo was simultaneously restored to 7.4. Amylase release induced by continuous stimulation with 100 pM CCK-8 showed a biphasic time course. A second sustained phase of amylase release was significantly reduced at pHo 6.0, but amylase release increased again when pHo was restored to 7.4. These results indicate that extracellular pH modifies the extent of secretagogue-induced Ca2+ entry into the pancreatic acinar cell, and the inhibitory effects of lowered pHo on the secretory response are largely due to a limited increase in [Ca2+]i.
Asada
N
N
Medical Sciences Laboratory, Hokkaido University of Education at Asahikawa, Japan.
Habara
Y
Y
Kanno
T
T
D000681:Amylases / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D005110:Extracellular Space / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008345:Manganese; D010179:Pancreas / Q000378:metabolism*; D051381:Rats; D012636:Secretory Rate / Q000187:drug effects; D012844:Sincalide / Q000494:pharmacology*
D008345:Manganese; D000681:Amylases; D012844:Sincalide; D002118:Calcium