Properties of large conductance Ca(2+)-activated K+ channels were studied in the soma of motoneurones visually identified in thin slices of neonatal rat spinal cord. The channels had a conductance of 82 +/- 5 pS in external Ringer solution (5.6 mM Ko+/(/)155 mM Ki+) and 231 +/- 4 pS in external high-Ko solution (155 mM Ko+/(/)155 mM Ki+). The channels were activated by depolarization and by an increase in internal Ca2+ concentration. Potentials of half-maximum channel activation (E50) were -13, -34, -64 and -85 mV in the presence of 10(-6), 10(-5), 10(-4) and 10(-3) M internal Ca2+, respectively. Using an internal solution containing 10(-4) M Ca2+, averaged KCa currents showed fast activation within 2-3 msec after a voltage step to +50 mV. Averaged KCa currents did not inactivate during 400 msec voltage pulses. External TEA reduced the apparent single-channel amplitude with a 50% blocking concentration (IC50) of 0.17 +/- 0.02 mM. KCa channels were completely suppressed by externally applied 100 mM charybdotoxin. It is concluded that KCa channels activated by Ca2+ entry during the action potential play an important role in the excitability of motoneurones.