The mechanism of proadrenomedullin N-terminal 20 peptide (PAMP)-induced inhibition of catecholamine release from adrenergic nerve was investigated in nerve growth factor-treated PC12 cells that have differentiated characteristics somewhat similar to noradrenergic neurons. The effect of PAMP on the excitability of these cells was investigated with the use of perforated whole-cell clamp. PAMP hyperpolarized the membrane by increasing a K+ conductance in a dose-dependent manner. The current-voltage relationship (I-V) relationship of the PAMP-induced K+ conductance exhibited inward-going rectification. The activation was abolished by microinjecting GDPbetaS into the cells or pretreating the cells with pertussis toxin. These results indicate that a pertussis toxin-sensitive G protein is involved in the signal transduction. The PAMP-induced activation of the K+ conductance was attenuated by microinjecting antibody against the carboxyl terminus of Galphai3, but it was not influenced by microinjecting antibody against the common carboxyl termini of Galphai1 and Galphai2, which indicated that the G protein coupling the PAMP receptor to the inwardly rectifying K+ current is Galphai3. The PAMP-induced hyperpolarization may inhibit the catecholamine release from the neurons by attenuating the action potential frequency.