Ca2+-Induced Ca2+Release in Chromaffin Cells Seen from inside the ER with Targeted Aequorin

• Published in: The Journal of cell biology Vol. 144; no. 2; pp. 241 - 254
• Main Authors: Alonso, Maria Teresa, Barrero, Maria José, Michelena, Pedro, Carnicero, Estela, Cuchillo, Inmaculada, García, Antonio G., García-Sancho, Javier, Montero, Mayte, Alvarez, Javier
• Format: Journal Article
• Language: English
• Published: United States Rockefeller University Press 25.01.1999; The Rockefeller University Press
• Subjects: Aequorin; Anatomy & physiology; Animals; Caffeine - pharmacology; Calcium; Calcium - metabolism; Calcium Channel Blockers - pharmacology; Cattle; Cells; Cellular biology; Chromaffin cells; Chromaffin Cells - drug effects; Chromaffin Cells - metabolism; Depolarization; Endoplasmic Reticulum - metabolism; Genes; Histamine - pharmacology; Histamines; Inositols; Microscopy, Confocal; Neurons; PC12 cells; Regular; Ryanodine - pharmacology; Secretion; Thapsigargin - pharmacology; Ungulates
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.144.2.241

The presence and physiological role of Ca2+-induced Ca2+release (CICR) in nonmuscle excitable cells has been investigated only indirectly through measurements of cytosolic [Ca2+]([Ca2+]c). Using targeted aequorin, we have directly monitored [Ca2+] changes inside the ER ([Ca2+]ER) in bovine adrenal chromaffin cells. Ca2+entry induced by cell depolarization triggered a transient Ca2+release from the ER that was highly dependent on [Ca2+]ERand sensitized by low concentrations of caffeine. Caffeine-induced Ca2+release was quantal in nature due to modulation by [Ca2+]ER. Whereas caffeine released essentially all the Ca2+from the ER, inositol 1,4,5-trisphosphate (InsP3)-producing agonists released only 60-80%. Both InsP3and caffeine emptied completely the ER in digitonin-permeabilized cells whereas cyclic ADP-ribose had no effect. Ryanodine induced permanent emptying of the Ca2+stores in a use-dependent manner after activation by caffeine. Fast confocal [Ca2+]cmeasurements showed that the wave of [Ca2+]cinduced by 100-ms depolarizing pulses in voltage-clamped cells was delayed and reduced in intensity in ryanodine-treated cells. Our results indicate that the ER of chromaffin cells behaves mostly as a single homogeneous thapsigargin-sensitive Ca2+pool that can release Ca2+both via InsP3receptors or CICR.