Pertussis toxin modification of PC12 cells lowers cytoskeletal F-actin and enhances norepinephrine secretion: involvement of protein kinase C and protein phosphatases.

We have investigated the relationship between norepinephrine secretion and cytoskeletal F-actin in rat phaeochromocytoma PC12 cells. Stimulation of PC12 cells with extracellular ATP or high K+ caused both the release of norepinephrine and a decrease in F-actin. The stimulation of secretion and the decrease in F-actin were dependent on extracellular Ca2+. The addition of Ca2+ to digitonin-permeabilized PC12 cells also stimulated norepinephrine release and decreased F-actin. Modification of PC12 cells with pertussis toxin caused a 35% decrease in F-actin, and it enhanced ATP-stimulated and K+ stimulated norepinephrine secretion from intact cells and Ca(2+)-dependent norepinephrine secretion from permeabilized cells. After down regulation of protein kinase C, pertussis toxin still enhanced secretion, but it had no effect on F-actin indicating that the effect of pertussis toxin on F-actin was dependent on protein kinase C activity. The addition of okadaic acid, an inhibitor of serine/threonine protein phosphatases, to PC12 cells caused a decrease F-actin, but it had no effect on ATP-stimulated or K(+)-stimulated norepinephrine secretion. After down regulation of protein kinase C, much higher concentrations of okadaic acid were need to decrease F-actin. The similarity between the effects of pertussis toxin and low concentrations of okadaic acid suggest that the effect of pertussis toxin on cytoskeletal F-actin in PC12 cells may result from an inhibition of protein phosphatase 2A.