PHORBOL ESTERS ATTENUATE GLUTAMATE-STIMULATED INOSITOL PHOSPHOLIPID HYDROLYSIS IN NEURONAL CULTURES

The phorbol diesters 12-O-tetradecanoylphorbol-13-acetate (TPA) and phorbol-12,13-dibutyrate, but not 4-α-phorbol-didecanoate, inhibited the stimulation of inositol phospholipid hydrolysis by excitatory amino acids and carbamylcholine in primary cultures of cerebellar neurons. This inhibition was mimicked by the synthetic diacylglycerol 1,2-dioleoyl-rac-glycerol (DOG) and was selective for a specific glutamate-phosphoinositide receptor subtype (GP2 receptor) activated by glutamate and quisqualate. TPA was nearly inactive in inhibiting the stimulation of inositol phospholipid hydrolysis by N-methyl-D-aspartate, a selective agonist of the GP1 receptor. Phorbol diesters and DOG attenuated the stimulation of inositol phospholipid hydrolysis by glutamate and quisqualate also in cerebellar slices from 9-15-day-old rats; however, using this preparation, their action was weak and required high concentrations (>1 μM). The inhibition of signal transduction by phorbol diesters was not consequent to a reduced binding of glutamate to its membrane recognition sites. In fact, TPA induced only a small increase in the K(D) but not change in the B(max) of [3H]glutamate binding in cerebellar membranes. Phorbol diesters may act to inhibit specific GTP-binding proteins or particular forms of phosphoinositidase C associated with GP2 or muscarinic cholinergic receptors.