Dataset Information

Functional coupling between the caffeine/ryanodine-sensitive Ca2+ store and mitochondria in rat aortic smooth muscle cells.

ABSTRACT: We investigated the role of mitochondria in the agonist-induced and/or caffeine-induced Ca2+ transients in rat aortic smooth muscle cells. We explored the possibility that proliferation modulates the coupling between mitochondria and endoplasmic reticulum. Ca2+ transients induced by either ATP or caffeine were measured in presence or absence of drugs interfering with mitochondrial activity in freshly dissociated cells (day 1) and in subconfluent primary culture (day 12). We found that the mitochondrial inhibitors, rotenone or carbonyl cyanide m-chlorophenylhydrazone, as well as the permeability transition pore inhibitor, cyclosporin A, had no effect on the ATP-induced Ca2+ transient at either day 1 or day 12, but prevented caffeine-induced cytosolic Ca2+ increase at day 12 but not at day 1. Close connections between ryanodine receptors and mitochondria were observed at both day 1 and 12. Thapsigargin (TG) prevented ATP- and caffeine-induced Ca2+ transients at day 1. At day 12, where only 50% of the cells were sensitive to caffeine, TG did not prevent the caffeine-induced Ca2+ transient, and prevented ATP-induced Ca2+ transient in only half of the cells. Together, these data demonstrate that rat aortic smooth muscle cells at day 1 have an ATP- and caffeine-sensitive pool, which is functionally independent but physically closely linked to mitochondria and totally inhibited by TG. At day 12, we propose the existence of two cell populations: half contains IP3 receptors and TG-sensitive Ca2+ pumps only; the other half contains, in addition to the IP3-sensitive pool independent from mitochondria, a caffeine-sensitive pool. This latter pool is linked to mitochondria through the permeability transition pore and is refilled by both TG-sensitive and insensitive mechanisms.

SUBMITTER: Vallot O

PROVIDER: S-EPMC1221962 | biostudies-other | 2001 Jul

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description: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.

Dataset Information

Functional coupling between the caffeine/ryanodine-sensitive Ca2+ store and mitochondria in rat aortic smooth muscle cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets