Ex Vivo Entrainment of the Isolated Mammalian Biological Clock Reveals Intrinsic Plasticity in Clock Gene Rhythm Dynamics

Abstract The suprachiasmatic nucleus (SCN) in the brain, the central circadian clock, governs timekeeping under different lighting conditions throughout seasons. It remains elusive how circadian alignment with various light cycles is achieved by the SCN at the level of core clock gene rhythms. We instituted ex vivo long-term periodic optogenetic stimulation and bioluminescence recording on a timescale of weeks to study circadian entrainment of core clock gene PER2 rhythms in the isolated SCN. We show that a single early-night optogenetic stimulation elongates the PER2 falling phase, while a late-night stimulation contracts the rising phase, to induce delay and advance phase resetting. Phase resetting also resulted in subsequent persistent changes in clock period – with phase advances leading to period shortening, and phase delays resulting in lengthening. We demonstrate stable entrainment of the isolated clock to 22h and 25h stimulation cycles, which is achieved via repeated rising phase shortening and falling phase lengthening, respectively. When presented with optogenetic skeleton photoperiods (2 pulses/24h cycle) that simulate short-day or long-day photoperiods, isolated SCN entrained preferentially to the short-day photoperiod by phase-jumping, similar to circadian behavioral plasticity classically described in intact mice and files. Skeleton photoperiod entrainment is achieved via increasing the relative duration of the falling phase to the rising phase. Clock gene waveform changes are specific to entrainment as they do not persist following release into constant darkness. Our results reveal how core clock gene rhythms in the SCN encode different lighting conditions, and that key aspects of circadian behavioral plasticity reside within the SCN itself.