Ca2+ ions play an important role during rhythmic bursting of thalamocortical neurons within sleep. The function of Ca2+ during the tonic relay mode of these neurons during wakefulness is less clear. Here, we report that tonic activity in thalamocortical cells results in an increase in the intracellular Ca2+ concentration and subsequent release of Ca2+ from intracellular stores mediated via ryanodine receptors (RyRs). Blockade of Ca2+ release shifted the regular firing of single action potentials toward the generation of spike clusters. Regular spike firing and intracellular Ca2+ release thus appear to be functionally coupled in a positive feedback manner, thereby supporting the relay mode of thalamocortical cells during wakefulness. Regulatory influences may be coupled to this system via the cyclic ADP ribose pathway.