Pacemaker activity of the Aplysia bursting pacemaker neuron R-15 was analyzed. It was shown that the free intracellular Ca2+ concentration, as measured by arsenazo III, increases during the depolarizing phase of the pacemaker cycle and declines throughout the hyperpolarizing phase that follows. This increase in Ca2+ results from the activation of voltage-dependent Ca2+ channels that open during the depolarizing phase of the cycle. The extracellular K+ concentration also increases during the depolarizing phase of the cycle and is correlated with an outward K+ current that opposes the inward current carried by Ca2+ ions. The increase in internal Ca2+ is sufficient to activate a K+ conductance that depends on the magnitude of the change in internal Ca2+ and on membrane potential, which is responsible for the hyperpolarizing phase of the cycle. It is proposed that the membrane oscillation depends on three separate but linked systems, which include a voltage-dependent Ca2+ channel, the internal Ca2+ concentration, and a Ca2+-activated K+ channel.