ABSTRACT
In mouse luteinized-granulosa cells (MGLC), ATP induces an increase in intracellular Ca2+ concentration by stimulating phospholipase C (PLC) associated with purinergic receptors, leading to production of inositol 1,4,5-trisphosphate (IP3) and subsequent release of Ca2+ from intracellular stores. In this study, we examined the cross-talk between the ryanodine receptors (RyR) and IP3 receptors (IP3R) in response to ATP in MGLC. Specifically, the effect of RyR modulators on ATP response was examined. The results showed that ATP-induced intracellular calcium elevation was abolished by inhibitors of the RyR, such as dantrolene (25 microM) and ryanodine (80 microM). When the MGLC were stimulated with activators of RyR, 2 microM ryanodine and 10 mM caffeine, the ATP-elicited response was decreased. These actions were independent of IP3 production stimulated by ATP. Hence, ATP-induced intracellular Ca2+ mobilization involves the coordinated action of both types of calcium release channels (CRCs). Using fluorescent probes, it was shown that IP3R is uniformly distributed throughout the cell; in contrast, RyR is mainly found around the nuclei. It is concluded that the IP3R and the RyR are functionally associated, and both play a role in the pattern of Ca2+ increase observed during purinergic stimulation of MGLC. This coupling may provide a highly efficient amplification mechanism for ATP stimulation of Ca2+ mobilization.
Subject(s)
Adenosine Triphosphate/pharmacology , Calcium Channels/metabolism , Granulosa Cells/metabolism , Receptors, Cytoplasmic and Nuclear/metabolism , Ryanodine Receptor Calcium Release Channel/metabolism , Animals , Calcium Channels/drug effects , Dantrolene/pharmacology , Extracellular Space/metabolism , Female , Granulosa Cells/drug effects , Granulosa Cells/ultrastructure , Inositol 1,4,5-Trisphosphate/biosynthesis , Inositol 1,4,5-Trisphosphate Receptors , Luteinization/physiology , Mice , Receptors, Cytoplasmic and Nuclear/drug effects , Ryanodine Receptor Calcium Release Channel/drug effects , Uridine Triphosphate/pharmacologyABSTRACT
The aim of the present study was to characterize the role played by intracellular and extracellular calcium and sodium on the maitotoxin (MTX) response in Chinese hamster ovary (CHO) cells. The results presented here indicated that: (1) MTX activates calcium and sodium influx in a concentration-dependent manner; (2) extracellular calcium is required for the sodium influx; (3) removal of the extracellular sodium did not prevent the MTX-induced calcium influx; (4) elevation in the intracellular calcium concentration potentiates the MTX response; and (5) MTX, at the concentrations tested, did not compromise cell viability.
