Involvement of Gαq/11 in m3-Muscarinic Receptor Stimulation of Phosphatidylinositol 4,5 Bisphosphate-Specific Phospholipase C in Rat Parotid Gland Membranes

Carbachol stimulation of phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis by rat parotid gland membranes is dependent on the presence of GTPγS and is a result of m3-muscarinic receptor regulation of G-protein coupled, PIP2-specific phospholipase C (PLC). The PLC activity (>80%) was solubilized with 1% Na-cholate but the solubilized enzyme was not stimulated by GTPγS and carbachol. Immunoblotting of rat parotid membranes with polyclonal antiserum, which recognizes α-subunits of the Gq/11 family, indicated the presence of two immunoreactive proteins of approximate molecular weights 41 and 42 kDa. Incubation of membranes with the common Gαq/11 antiserum attenuated the stimulation of PIP2 hydrolysis, induced by GTPγS alone and by carbachol, in the presence of GTPγS. The antiserum had no effect on PIP2 hydrolysis in unstimulated membranes or in the cholate extract, where it is uncoupled from the G-protein. Antiserum against Gαi, which is also coupled to the m3-muscarinic receptor in this tissue, had no effect on either basal or stimulated PIP2 hydrolysis. These results demonstrate that in rat parotid gland, activation of PIP2-specific PLC by m3-muscarinic receptor stimulation is mediated via α-subunits of the Gq/11 family of G-proteins.