Signal transduction and Ca2+ uptake activated by endothelins in rat brain endothelial cells.

The activation of signal transduction pathways by endothelin-1 or endothelin-3 were investigated in rat cerebromicrovascular endothelial cells. Endothelin-1 induced a rapid increase in inositol triphosphate (IP3) formation in these cells, whereas endothelin-3 was only moderately effective at high concentrations. Both endothelins also increased uptake of 45Ca2+ in these cells. Endothelin-1-induced IP3 formation or 45Ca2+ uptake were inhibited by endothelin ETA receptor antagonist BQ-123. Ryanodine, an inhibitor of intracellular Ca2+ mobilization, selectively endothelin-1-induced 45Ca2+ uptake, whereas nickel or suramin inhibited endothelin-3-induced 45Ca2+ uptake. The results indicate that endothelin-1 elevates 45Ca2+ uptake in rat brain endothelial cells by mechanisms coupled to the mobilization of intracellular Ca2+ stores. Both endothelin-1- and endothelin-3-induced 45Ca2+ uptake were inhibited by receptor operated Ca2+ channel blocker SK&F 96365, whereas they were insensitive to dihydropyridine derivatives nifedipine and nitrendipine. The release of arachidonic acid from rat brain endothelial cells observed in response to endothelin-1 was inhibited by ryanodine or SK&F 96365, implicating participation of both intra- and extra- cellular components of Ca2+ signaling in activating endothelial secretion of vasoactive substances.

The long-term down-regulation of dihydropyridine receptors by Bay K 8644 in PC12 cells.

Treatment of PC12 cells with Bay K 8644 for 12 hr or more leads to an almost 80% decrease in the subsequent ability of Bay K 8644 to stimulate the uptake of radioactive calcium into the cells. This effect is a property of the S(-)isomer of Bay K 8644; pre-treatment with the R(+)isomer, now known to be a calcium channel blocker, has the opposite effect. This treatment is specific in that it does not interfere with the stimulation of calcium uptake by potassium, ATP, or nerve growth factor. Such treatment is accompanied by a 90% decrease in the ability of Bay K 8644 to stimulate the release of norepinephrine. The characteristics of the binding of [3H]isradipine to control and to treated cells indicates that the decrease in the effect of dihydropyridines is accompanied by a marked decrease in the number of dihydropyridine binding sites with no apparent change in the affinity of the remaining sites. The continued ability of depolarizing levels of potassium to stimulate calcium uptake and the induction of the protooncogene c-fos in Bay K 8644-treated cells indicates that the L-type calcium channels are still intact, but are simply unresponsive to dihydropyridine agonists.