Role of Phospholipid in the Transmitter-Induced Synthesis of Cyclic GMP in Nervous Tissue

The mechanism by which acetylcholine and other transmitters stimulate the synthesis of cyclic GMP was studied at the cellular and molecular levels using neuroblastoma N1E 115 cells and rat brain. (i) Phosphatidic acid added to the medium stimulated the influx of calcium and elevated intracellular cyclic GMP content in the neuroblastoma cells. Treatment of the cells with phospholipase C increased the 32 Pi incorporation into phosphatidic acid and, in parallel, stimulated calcium influx and cyclic GMP synthesis in the cells. (ii) Guanylate cyclase was readily solubilized from neuroblastoma cells by homogenization with hypotonic buffer. The supernatant also contained an endogenous activator of small molecule for guanylate cyclase. The activator was also detected in the supernatant of rat brain and was purified to a high purity. Calcium was required for the activation of guanylate cyclase by the endogenous activator. (iii) Monoclonal antibody to the soluble guanylate cyclase from rat brain cross-reacted with the soluble enzyme from various tissues of rat, but not with the soluble enzyme of mouse brain and rabbit brain indicating a speciecs difference of soluble guanylate cyclase. Immunochemical titration study revealed that guanylate cyclase in the synaptosomal soluble fraction of rat brain was in a highly activated state.