In vitro studies of the role of cyclic-AMP in the mammalian circadian clock in the suprachiasmatic nuclei

The behavior and physiology of most organisms are organized into 24 hr rhythms that are produced by self-sustaining clocks located within the organisms. In mammals the primary circadian clock is located in the suprachiasmatic nuclei (SCN) of the brain. My research investigated the biochemical mechanisms underlying this clock, focusing in particular on the potential role of cAMP.All components of circadian oscillators share two properties. First, temporary perturbations in them induce permanent phase-shifts of the 24 hr clock. Second, the components exhibit 24 hr rhythms in the absence of external timing information. We attempted to determine whether cAMP showed these two characteristics in the SCN in vitro, where the clock could be maintained under constant conditions.We first determined that the SCN circadian clock can produce multiple daily cycles of SCN neuronal activity in vitro. Under our conditions the clock produces a stable 24 hr rhythm in electrical activity for three days.We then showed that 1-hr treatments that either increase cAMP levels or stimulate cAMP-dependent mechanisms permanently reset the SCN clock. These treatments are only effective during the daytime, inducing maximal advances of 4-5 hr when applied 3-8 hr into subjective daytime. These effects are specific for cAMP, since analogs of cGMP induce phase shifts only when applied during the subjective night, when cAMP analogs are ineffective.Finally, we showed that the level of cAMP fluctuates within the SCN in vitro, with high levels during late day and late night. These changes in cAMP concentration are accompanied by corresponding changes in the activity of phosphodiesterase, the enzyme mediating cAMP degradation.Together, these results are strong evidence for cAMP and phosphodiesterase being components of the mammalian circadian oscillator. As such, they represent an important advance in our understanding of this oscillator, and should lead to the identification of other elements of the SCN clock mechanism.U of I OnlyETDs are only available to UIUC Users without author permissio