The effects of interfering with GTP-binding proteins on the activation mechanism of calcium release-activated calcium current.

In electrically non-excitable cells, Ca2+ entry is mediated predominantly by the store-operated Ca2+ influx pathway, which is activated by emptying the intracellular Ca2+ stores. Just how the Ca2+ content of the stores is communicated to the activity of store-operated Ca2+ channels in the plasma membrane is unclear. It has been suggested that, in some cell types, the link is accomplished by either a small or a heterotrimeric GTP-binding protein, which is inhibited by guanosine 5'-O-(3-thiotriphosphate) (GTP[gamma-S]) and, in some cases, pertussis toxin. Using the whole-cell patch-clamp technique to directly measure the store-operated Ca2+ current ICRAC (Ca2+-release-activated Ca2+ current) in RBL cells, we report that manipulations designed to interfere with GTP-binding protein activity (dialysis with GTP[gamma-S], exposure to pertussis toxin) routinely fail to affect the activation of ICRAC. However, these agents alter the activity of a K+ current in the same cells, demonstrating biological activity. Furthermore, activation of ICRAC does not seem to require the presence of a pre-existing diffusible messenger in the cytoplasm to any appreciable extent because the current reaches the same amplitude irrespective of the whole-cell dialysis time. We conclude that neither a mobile pre-existing molecule nor a GTP-dependent step is necessary for the activation of ICRAC in RBL-1 cells.