Intracellular mechanisms underlying the functional suppression of ionotropic glutamate receptors by activation of metabotropic glutamate receptors were investigated in cultured chick Purkinje neurons. The intracellularly recorded depolarization induced by L-AMPA (an ionotropic glutamate receptor agonist, (S)-alpha-amino-4-hydroxy-5-methyl-4-isoxazolepropionic acid) and the L-AMPA-induced inward current recorded by whole-cell voltage clamping were used. L-AMPA responses were suppressed by trans-ACPD (a selective agonist of metabotropic glutamate receptor, (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid) for various durations, with the longest duration about 60 min. This trans-ACPD effect was antagonized by l-(+)-2-amino-3-phosphonopropionic acid (a metabotropic glutamate receptor antagonist) and N(G) -monomethyl-L-arginine (a nitric oxide synthase inhibitor). Sodium nitroprusside, 3-morpholinosydnonimine (nitric oxide donors), and potassium ferricyanide mimicked trans-ACPD, and effects of trans -ACPD, sodium nitroprusside, and 3-morpholinosydnonimine were blocked by hemoglobin (a nitric oxide scavenger) but not by methemoglobin, while the effect of potassium ferricyanide was not affected by either hemoglobin or methemoglobin. 8-Bromo-cGMP also suppressed L-AMPA responses. KT5823 (a protein kinase G inhibitor) antagonized effects of trans-ACPD, 8-bromo-cGMP, and sodium nitroprusside. Phorbol 12,13-diacetate (a protein kinase C activator) also suppressed L-AMPA responses, and phorbol 12,13-diacetate plus trans-ACPD or phorhol 12,13-diacetate plus sodium nitroprusside showed an additive effect. Calphostin C and polymyxin B (protein kinase C inhibitors) antagonized the effect of trans-ACPD. These results suggest that activation of metabotropic glutamate receptors leads to the functional suppression of L-AMPA-sensitive ionotropic glutamate receptors in chick Purkinje neurons, and trans-ACPD-induced suppression of L-AMPA responses can be mimicked by activation of protein kinase G and/or protein kinase C. The involvement of nitric oxide in the trans-ACPD effect is discussed.