A caffeine- and ryanodine-sensitive intracellular Ca2+ store can act as a Ca2+ source and a Ca2+ sink in PC12 cells

Abstract

We have investigated the modulation of stimulus-induced changes in intracellular Ca2+ concentration ([Ca2+]i) by a caffeine-and ryanodine-sensitive Ca2+ store in PC12 cells. In populations of fura-2-loaded cells, caffeine cause a concentration-dependent increase in [Ca2+]i that was saturable, reversible and inhibited in a use-dependent fashion by ryanodine. Maximal Ca2+ release occurred with 40 mM caffeine, with an EC50 of 13 mM caffeine and a Hill coefficient (h) of 2.7, indicating that the release mechanism was co-operative. Pretreatment of intact cell populations with increasing concentrations of caffeine in nominally Ca(2+)-free medium inhibited the subsequent Ca2+ response to a maximal concentration of ATP, in a dose-dependent manner. In permeabilized cells, a maximal concentration (40 microM) of InsP3 still released Ca2+ in the presence of a supramaximal concentration (50 mM) of caffeine, whereas caffeine was unable to release Ca2+ after the InsP3-sensitive store had been completely emptied. These data suggest that PC12 cells contain a uniquely InsP3-sensitive Ca2+ store, and a store that is sensitive to both InsP3 and caffeine. Depletion of the caffeine-sensitive Ca2+ store by caffeine and ryanodine pretreatment in intact cells attenuated the Ca2+ response to ATP, but not to 55 mM K+, suggesting that the caffeine-sensitive Ca2+ store acts as a Ca2+ source after ATP stimulation, but not after depolarization with 55 mM K+. Pretreatment of intact cells with ATP and ryanodine resulted in a use-dependent block of both caffeine- and ATP-mediated Ca2+ release, confirming that ATP stimulation of PC12 cells brings about activation of ryanodine receptors. The rate of recovery, but not the magnitude or rate of onset, of the depolarization-induced [Ca2+]i transient was modulated by the state of filling of the caffeine-sensitive Ca2+ store such that recovery was prolonged if the store was either full, or empty and unable to refill. We conclude that the caffeine- and ryanodine-sensitive Ca2+ store can act as a Ca2+ source and a Ca2+ sink in PC12 cells, and that its role may in part be governed by the nature of the stimulating agent.

Caffeine-Induced Ca 2+ Transients and Exocytosis in Paramecium Cells. A Correlated Ca 2+ Imaging and Quenched-Flow/Freeze-Fracture Analysis

“…(v) In the presence of Ca o 2+ , this is superimposed by a Ca 2+ influx, possibly via store-operated Ca 2+ channels as described in other systems (Fasolato, Innocenti & Pozzan, 1994). Superposition of Ca 2+ mobilization from stores and Ca 2+ influx also occurs in some neuroendocrine cells during caffeine stimulation (Barry & Cheek, 1994), just as we observe.…”

Mechanisms underlying angiotensin II-induced calcium oscillations

“…7D). These results indicate that, together, IP 3 and CICR events in the microdomains control the appearance of Ca 2ϩ oscillations in the entire cell.…”

“…Many permutations of configurations, IP 3 R and RyR isoform variations, spatial distributions, and PM-ER/SR-cytosolic interactions, are conceptually possible and, moreover, likely to exist in living cells (3,5,14,17,18,27,45,46,50). As such, we recognize that the predictions of this model represent a limited exploration, guided largely by measurements obtained from DVR pericytes.…”

“…As a baseline, we assumed that store-operated cation channel activity is regulated by the average of Ca 2ϩ concentrations in the IP 3 and RyR storage pools. If SOC activity were controlled by [Ca] IP3R-SR alone, which is lower than the average SR Ca 2ϩ concentration during oscillations (Fig.…”

Expression and Function of Ryanodine Receptors in Nonexcitable Cells

“…However, there was an apparent change in isoform expression pattern from RyR1 and RyR2 in undifferentiated cells to only RyR1 in differentiated cells. The expression of RyR1 in undifferentiated PC12 cells may explain the observation that theophylline is equipotent with caffeine at stimulating Ca 2ϩ release from this clone (34), since RyR1 appears equally sensitive to these two methylxanthines (37).…”