A caffeine-sensitive Ca2+ store modulates K+-evoked secretion in chromaffin cells.
Catecholamine release from bovine adrenal medulla chromaffin cells superfused with a Krebs-N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid solution was monitored on-line with an electrochemical detector. Caffeine (10 mM) progressively depressed the magnitude of secretory responses to depolarizing pulses of 70 mM K+ and 2 mM Ca2+ (70 K+/2 Ca2+) in cells superfused with a Krebs-N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid solution containing 0 mM Ca2+ + 0.5 mM EGTA; blockade reached 80% at the third 70 K+/2 Ca2+ challenge given in the presence of caffeine. A similar effect was obtained when, instead of continuous superfusion, prepulses of caffeine were applied (10 mM for 60 s). The blocking effects of caffeine on K+-induced secretion depended on the time of exposure to the drug: the longer the exposure time the greater the blockade. The recovery of the K+ secretory responses previously impaired by caffeine was always gradual and followed a staircase mode. This contrasts with the effects of caffeine on various parameters measuring Ca2+ entry through Ca2+ channels, which did not parallel its effects on K+-evoked secretion. The secretion data, however, are compatible with the disappearance and recovery of an intracellular Ca2+ concentration signal triggered by K+ in single chromaffin cells loaded with fura 2 and treated with 10 mM caffeine. Thus, contrary to previous views, the depression of secretion by caffeine does not seem to be associated with inhibition of extracellular Ca2+ entry through Ca2+ channels. These functional data are, rather, compatible with the view that the degree of filling of a caffeine-sensitive intracellular Ca2+ store might regulate the extent of exocytosis. When emptied, such a store might act as a sink for the external Ca2+ entering through Ca2+ channels during cell depolarization, thus decreasing the intracellular Ca2+ concentration available for exocytosis.
Lara
B
B
Departamento de Farmacología y Terapéutica, Facultad de Medicina, Universidad Autónoma de Madrid, Spain.
López
M G
MG
Villarroya
M
M
Gandía
L
L
Cleeman
L
L
Morad
M
M
García
A G
AG
D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D015220:Calcium Channels / Q000378:metabolism / Q000502:physiology; D002395:Catecholamines / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D007425:Intracellular Membranes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000494:pharmacology*; D013997:Time Factors
D015220:Calcium Channels; D002395:Catecholamines; D002110:Caffeine; D011188:Potassium; D002118:Calcium