This continuing investigation is intended to elucidate the mechanism and significance of the enhanced turnover of certain acidic phospholipid classes resulting from the activation of Alpha1-adrenergic and muscarinic cholinergic receptors on the cell surface. This phospholipid effect, triggered by ligand binding to specific receptors, is presumed to be a vital link in the transmission and amplification of an extracellular signal which translates into characteristic intracellular events. We plan to obtain information on the mechanism of neurotransmitter action in intact dissociated rat pinealocytes, rat smooth muscle cells, and cultured mouse neuroblastoma cells by selectively and specifically inhibiting calcium entry or reactions leading to hydrolysis and resynthesis of phosphoinositides, to arachidonic acid release and metabolism or to cyclic GMP formation and breakdown. After each incubation, control, stimulated or characteristically modified by the addition of a targeted inhibitor at 0 time or up to 5 minutes after the start, we shall determine ligand binding, membrane fluidity, phospholipid turnover, Ca2+ fluxes and intracellular levels, cyclic GMP levels and formation, and arachidonic acid release, in order to derive clues about the relationship among the several parameters. Because events before the point of inhibition are expected to continue to occur while those after are blocked, we shall be able to obtain clear information about the sequence and physiological significance of the several phenomena.
