Ryanodine-sensitive intracellular Ca²⁺ channels are involved in the output from the SCN circadian clock

The suprachiasmatic nuclei (SCN) contain the major circadian clock responsible for generation of circadian rhythms in mammals. The time measured by the molecular circadian clock must eventually be translated into a neuronal firing rate pattern to transmit a meaningful signal to other tissues and organs in the animal. Previous observations suggest that circadian modulation of ryanodine receptors (RyR) is a key element of the output pathway from the molecular circadian clock. To directly test this hypothesis, we studied the effects of RyR activation and inhibition on real time expression of PERIOD2::LUCIFERASE, intracellular calcium levels and spontaneous firing frequency in mouse SCN neurons. Furthermore, we determined whether the RyR-2 mRNA is expressed with a daily variation in SCN neurons. We provide evidence that pharmacological manipulation of RyR in mice SCN neurons alters the free [Ca²⁺ ]_i in the cytoplasm and the spontaneous firing without affecting the molecular clock mechanism. Our data also show a daily variation in RyR-2 mRNA from single mouse SCN neurons with highest levels during the day. Together, these results confirm the hypothesis that RyR-2 is a key element of the circadian clock output from SCN neurons.