Basic fibroblast growth factor maintains calcium homeostasis and granulosacell viability by stimulating calcium efflux via a PKC delta-dependent pathway

Previous studies have demonstrated that basic fibroblast growth factor prev ents granulosa cell apoptosis. The following six observations provide insig ht into the mechanism by which basic fibroblast growth factor mediates its antiapoptotic action. First, loading granulosa cells with 1,2 bis(2-aminoph enoxy)-ethane-N,N,N ' ,N ' -tetraacetic acid, an intracellular calcium chel ator, prevented apoptosis when granulosa cells were deprived of basic fibro blast growth factor. Second, treatment with thapsigargin, an agent known to increase intracellular free calcium, induced granulosa cell apoptosis even in the presence of basic fibroblast growth factor. Third, an activator of PKC mimicked, whereas PKC inhibitors blocked, basic fibroblast growth facto r's antiapoptotic action. Fourth, continuous basic fibroblast growth factor exposure maintained relatively constant levels of intracellular free calci um, and a PKC inhibitor induced a sustained 2- to 3-fold increase in intrac ellular free calcium. Fifth, granulosa cells, as well as spontaneously immo rtalized granulosa cells, were shown to express PKC delta, -lambda, and -ze ta. Finally, the PKC delta -specific inhibitor, rottlerin, blocked basic fi broblast growth factor's antiapoptotic action in granulosa cells and sponta neously immortalized granulosa cells. These studies suggest that basic fibr oblast growth factor regulates intracellular free calcium through a PKC del ta -dependent mechanism and that a sustained increase in intracellular free calcium is sufficient to induce and is required for granulosa cell apoptos is.