Fluoride but not phorbol esters stimulate rat urinary bladder prostanoid synthesis: Investigations into the roles of G proteins and protein kinase C

Abstract

The role of G proteins and protein kinase C in mediating muscarine receptor-linked prostanoid synthesis by the rat urinary bladder was investigated using the G protein activator, sodium fluoride (NaF); the protein kinase C activators, phorbol myristate (PMA) and phorbol dibutyrate (PDBU); the protein kinase C inhibitor, H7, and the parasympathomimetic, carbachol. NaF stimulated in vitro rat urinary bladder prostacyclin (PGI₂) synthesis (EC₅₀ = 6 mmol. ⁻¹), an action inhibited by the presence of EDTA (10 mmol. 1⁻¹). Carbachol potentiated the stimulatory action of NaF. NaF (10 mmol.l⁻¹)-stimulated PGI₂ synthesis was inhibited by the calcium channel blockers verapamil, nifedipine and the protein kinase C inhibitor, H7, in concentrationdependent manners. Carbachol-stimulated PGI₂ synthesis was also inhibited by H7. PDBU and PMA were without effect on de novo, NaF-or carbachol-stimulated urinary bladder PGI₂ synthesis. Other prostanoids (PGE₂ and PGF_2α) were stimulated to the same degree as PGI₂ by NaF, and inhibited equally by H7 and calcium channel blockers. Dibutyryl adenosine 3′:5′-cyclic monophosphate was without effect on de novo or NaF-stimulated prostanoid synthesis. Since fluoride activates G proteins, these data indicate that: (1) muscarine receptorprostanoid synthesis coupling is mediated by G proteins in the rat urinary bladder; (2) fluoride action is mediated by protein kinase C and not adenyl cyclase, probably through activation of phospholipase C and therefore the generation of the protein kinase C activator diacyl glycerol; (3) activated protein kinase C may initiate Ca²⁺⁺ mobilisation linked to prostanoid synthesis; and (4) the lack of effect of the phorbol esters on urinary bladder PGI₂ synthesis, in contrast to that on other smooth muscle, indicates that in different smooth muscle tissues there are varying forms of protein kinase C.