N-ACETYLASPARTYLGLUTAMATE INHIBITS FORSKOLIN-STIMULATED CYCLIC-AMP LEVELS VIA A METABOTROPIC GLUTAMATE-RECEPTOR IN CULTURED CEREBELLAR GRANULE CELLS

The neuronal dipeptide N-acetylaspartylglutamate (NAAG) fulfills sever al of the criteria for classification as a neurotransmitter including localization in synaptic vesicles, calcium-dependent release after neu ronal depolarization, and low potency activation of N-methyl-D-asparta te receptors. In the present study, the influence of NAAG on metabotro pic receptor activation in cerebellar granule cells was examined in ce ll culture. Stimulation of granule cell adenylate cyclase with forskol in increased cyclic AMP (cAMP) several hundredfold above basal levels within 10 min in a concentration-dependent manner. Although glutamate, NAAG, and the metabotropic receptor agonist trans-1-amino-1,3-cyclope ntanedicarboxylic acid did not alter the low basal cAMP levels, the ap plication of 300 muM glutamate or NAAG or trans-1-amino-1,3-cyclopenta nedicarboxylic acid reduced forskolin-stimulated cAMP in granule cells by 30-50% in the absence or presence of inhibitors of ionotropic acid ic amino acid receptors, as well as 2-amino-4-phosphonobutyrate. No ad ditivity in the inhibition of cAMP was found when 300 muM NAAG and tra ns-1-amino-1,3-cyclopentanedicarboxylic acid were coapplied. The beta- analogue of NAAG failed to reduce cAMP levels. Similar effects of NAAG and glutamate were obtained under conditions of inhibition of phospho diesterase activity and were prevented by pretreatment of the cells wi th pertussis toxin. These data are consistent with the activation by N AAG of a metabotropic acidic amino acid receptor coupled to an inhibit ory G protein. In contrast, the metabotropic acidic amino acid recepto r coupled to phosphoinositol turnover in these cells was not activated by NAAG. Granule cells in culture expressed very low levels of extrac ellular peptidase activity against NAAG, converting to glutamate < 0.1 % of the 10 muM through 1 mM NAAG applied to these cells during 15-min in vitro assays.