Caffeine‐Sensitive Calcium Stores in Bovine Adrenal Chromaffin Cells

Abstract

Caffeine was used to study the intracellular Ca2+ pools of bovine chromaffin cells. Its effects on cytosolic Ca2+ concentration ([Ca2+]i) were examined using fura-2. Caffeine caused a transient increase in [Ca2+]i in the presence or absence of extracellular Ca2+. In the former case, the caffeine-induced [Ca2+]i increase was higher and stayed above the basal value for several minutes. In the latter case, the [Ca2+]i rise was lower and fell to the basal level within 1 min. These results suggest that caffeine increases [Ca2+]i by causing both Ca2+ influx and Ca2+ release from intracellular pools. In the absence of extracellular Ca2+, ionomycin but not caffeine caused a further increase in [Ca2+]i in cells that had been treated with caffeine. Apparently there are at least two intracellular Ca2+ pools, only one of which is sensitive to caffeine. The caffeine-induced [Ca2+]i rise became smaller when the cells were pretreated with the inositol trisphosphate-generating agonists, methacholine and bradykinin. In addition, methacholine was unable to initiate a [Ca2+]i transient after the cells had been treated with caffeine. The results indicate that the caffeine-sensitive Ca2+ pools overlap with the inositol trisphosphate-sensitive pool and that the size of the latter pool is smaller than that of the former. The caffeine-sensitive Ca2+ pools were refilled after high K+ treatment, which suggests that the caffeine-sensitive Ca2+ pools may be important in buffering the cytosolic Ca2+. The effect of caffeine on [Ca2+]i is not due to inhibition of phosphodiesterase. Our results support a Ca2+ entry model in which depletion of intracellular Ca2+ pools controls the rate of Ca2+ entry across the plasma membrane.

Oxidative glutamate toxicity involves mitochondrial dysfunction and perturbation of intracellular Ca2+ homeostasis

“…The concentration of intracellular Ca 2 + was measured with Fura-2/AM, a calcium fluorescent ester chelator and indicator (Di Virgilio et al, 1988). After glutamate incubation, cells were harvested by gentle scraping, washed and resuspended in a standard medium containing 140 mM NaCl, 5 mM KCl, 1 mM MgCl 2 , 5.6 mM glucose, 1.5 mM CaCl 2 and 20 mM Hepes-Na (final pH 7.4), which was used for loading and for the subsequent fluorescence measurements (Liu et al, 1991). Fura-2/AM (5 mM) was added to the cell suspension, which was subsequently incubated for 30 min.…”

Pituitary Adenylate Cyclase‐Activating Polypeptide Causes Ca2+ Release from Ryanodine/Caffeine Stores Through a Novel Pathway Independent of Both Inositol Trisphosphates and Cyclic AMP in Bovine Adrenal Medullary Cells

“…The results that the Ca2~re-lease induced by histamine or carbachol, which depends primarily on Ca2~release from 1P 3 stores, was potently inhibited by ryanodine/caffeine, as well as by cinnarizine, indicate that these two ligands activate a large amount ofCa 2tinduced Ca2~release from ryanodine/caffeine stores by causing primary Ca2~release from 1P 3 stores. In fact, it is reported that 1P3 stores have less capacity than caffeine stores in bovine adrenal chromaffin cells (Liu et al, 1991) and that 1P3 caused less Ca 2~release than caffeine in permeabilized chromaffin cells (Robinson and Burgoyne, 1991;Stauderman et al, 1991).…”

Catecholamine Secretion from Bovine Adrenal Chromaffin Cells: The Role of the Na+/Ca2+ Exchanger and the Intracellular Ca2+ Pool

“…The [Ca2~] 1 of a single cell was measured as previously described (Liu et al, 1991). Each chromaffin cell selected for fluorescence measurement was alternately excited at 340 and 380 nm with a dualexcitation fluorometer (CM system, SPEX).…”

“…It has also been shown that the nonmitochondrial Ca2* pooi accumulates Ca2b rought into the cells through the voltage-gated Ca2c hannels (Liu et a!., 1991). The ability of this pool to release Ca2~by various Ca2~-re1easingagents has been characterized in detail; however, the physiological role of this pool in bovine chromaffin cells remains unclear.…”