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Circadian rhythms in mammals are orchestrated by a central clock in the suprachiasmatic nucleus (SCN). The clock emerges from cell-autonomous processes. Clock proteins cause the rhythmic expression of clock genes as well as output genes. Structural features and posttranslational modifications of clock proteins determine their intermolecular associations, subcellular localization, transcriptional activity, and stability. Some clock proteins positively and negatively regulate their own rhythmic expression via feedback on transcription. Patterns of clock gene expression within SCN cells and tissue are complex. The understanding of mammalian clock mechanisms critical to timekeeping is as yet incomplete.

The recent identification of specific genes responsible for the generation of endogenous circadian rhythmicity in the suprachiasmatic nucleus presents a new level of investigation into endogenous rhythmicity and mechanisms of synchronization of this circadian clock with the environmental light?dark cycle. This article describes techniques that employ antisense and decoy oligodeoxynucleotides (ODN) to determine the roles of specific molecular substrates both in endogenous rhythmicity and in regulating the effects of light on the mammalian circadian clock. Application of antisense ODN technology has revealed a role for timeless (Tim) in the core clock mechanism and established that induction of period1 (Per1) is required for light responsiveness. Likewise, a decoy ODN designed to sequester activated CREB protein definitively demonstrated a requirement for CRE-mediated transcription in light signaling. Experiments designed with these molecular tools offer new insights on the interaction of cellular processes and signaling with the molecular clockworks.

Publisher Summary The chapter explains that the clock properties predicted from in vivo studies are endogenous to the suprachiasmatic nucleus (SCN); they persist when the SCN is isolated in a brain slice and maintained under constant conditions in vitro. The SCN continues to generate a stable, near-24-hour rhythm of neuronal activity for up to three cycles in vitro, and this rhythm can be reset by a range of stimuli. At specific phases, or time domains, within the 24-hour cycle, the circadian clock in vitro is still receptive to only a subset of the various stimuli that can reset it. Therefore, the circadian clock regulates its own sensitivity to afferent signals. The chapter demonstrates that this gating is downstream from the second messengers in signaling pathways. It follows that the states of specific molecular gates regulated by the clock change over the 24-hour cycle and site(s) critical to gating signaling pathways that access the clock lie within the cells of the SCN.