Exocytosis coupled to mobilization of intracellular calcium by muscarine and caffeine in rat chromaffin cells.
J Neurochem
; 67(1): 155-62, 1996 Jul.
Article
em En
| MEDLINE
| ID: mdl-8666986
ABSTRACT
We used cultured rat chromaffin cells to test the hypothesis that Ca2+ entry but not release from internal stores is utilized for exocytosis. Two protocols were used to identify internal versus external Ca2+ sources (a) Ca2+ surrounding single cells was transiently displaced by applying agonist with or without Ca2+ from an ejection pipette. (b) Intracellular stores of Ca2+ were depleted by soaking cells in Ca2+ -free plus 1 mM EGTA solution before transient exposure to agonist plus Ca2+. Exocytosis from individual cells was measured by microelectrochemical detection, and the intracellular Ca2+ concentration ([Ca2+]i) was measured by indo-1 fluorescence. KCl (35 mM) and nicotine (10 microM) caused an immediate increase in [Ca2+]i and secretion in cells with or without internal Ca2+ stores, but only when applied with Ca2+ in the ejection pipette. Caffeine (10 mM) and muscarine (30 microM) evoked exocytosis whether or not Ca2+ was included in the pipette, but neither produced responses in cells depleted of internal Ca2+ stores. Pretreatment with ryanodine (0.1 microM) inhibited caffeine- but not muscarine-stimulated responses. Elevated [Ca2+]i and exocytosis exhibited long latency to onset after stimulation by caffeine (2.9 +/- 0.38 s) or muscarine (2.2 +/- 0.25 s). However, the duration of caffeine-evoked exocytosis (7.1 +/- 0.8 s) was significantly shorter than that evoked by muscarine (33.1 +/- 3.5 s). The duration of caffeine-evoked exocytosis was not affected by changing the application period between 0.5 and 30 s. An approximately 20-s refractory period was found between repeated caffeine-evoked exocytosis bursts even though [Ca2+]i continued to be elevated. However, muscarine or nicotine could evoke exocytosis during the caffeine refractory period. We conclude that muscarine and caffeine mobilize different internal Ca2+ stores and that both are coupled to exocytosis in rat chromaffin cells. The nicotinic component of acetylcholine action depends primarily on influx of external Ca2+. These results and conclusions are consistent with our original observations in the perfused adrenal gland.
Buscar no Google
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Cafeína
/
Cálcio
/
Sistema Cromafim
/
Exocitose
/
Muscarina
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
J Neurochem
Ano de publicação:
1996
Tipo de documento:
Article