Effect of a mu-opioid agonist (D-Ala2,N-MePhe4,Gly5-ol-enkephalin, DAGO), on glycine (Gly)-induced chloride current (IGly) was investigated in the periaqueductal gray (PAG) neurons acutely dissociated 1-2-week-old Wistar rats by the use of nystatin-perforated patch recording configuration under voltage-clamp condition. At a holding potential (VH) of -40 mV, DAGO caused a sustained potentiation of IGly at the low concentrations (10(-6)-10(-5) M) but reduced slightly the Gly response at the high concentration (10(-4) M). The reversal potential of IGly was equal to the Cl- equilibrium potential (ECl) and was not changed in the presence of 10(-6) M DAGO. The 10(-5) M Gly response was inhibited by the simultaneous treatment of forskolin and 3-isobutyl-1-methylxanthine (IBMX). H-89, a protein kinase A (PKA) inhibitor, increased the 10(-5) M Gly response but had little effect on the 10(-4) M Gly response. DAGO increased 10(-5) M Gly response in the presence of forskolin and IBMX but, not more than in the absence of forskolin and IBMX. The 10(-5) M Gly response augumented by DAGO was not affected by adding H-89. The present results suggest that the glycine-induced chloride current is cAMP dependent and is inhibited by PKA, and that the potentiation of the glycine response by DAGO is also cAMP dependent and is due to the inhibition of PKA as that of H-89. We conclude that the potentiation of glycine response by DAGO is mediated by an inhibition of cAMP-dependent PKA in the PAG neurons.