Transcriptional and translational inhibitors block serotonergic phase advances of the SCN circadian pacemaker in vitro

The mammalian circadian pacemaker is located in the suprachiasmatic nucleus (SCN). Phase shifts of the pacemaker are modulated by various inputs. The input that we investigated is the serotonergic (5HT) input from the raphe nuclei. 5HT phase-advances the SCN pacemaker when applied during mid-subjective day. In vitro studies indicate that 5HT phase-advances the mammalian circadian pacemaker through a process that includes stimulation of 5-HT7 receptors, activation of protein kinase A, and opening of K+ channels. How these cytoplasmic and membrane events translate into a shift in the molecular core of the circadian oscillator is not known. To further understand this process, we investigated whether serotonergic phase advances require protein synthesis. Using two reversible translational inhibitors, anisomycin and cycloheximide, we show that inhibition of protein synthesis blocks SHTergic phase shifts. We further show that a transcriptional inhibitor, 5,6-dichloro-l-P-ribobenzimidazole (DRB), also blocks the SHTergic phase shifts in the SCN circadian pacemaker. These results are similar to those found previously with respect to 5HTergic modulation of the Aplysia ocular circadian clock, and suggest that 5HT may phase-shift the SCN pacemaker through increasing transcription and translation of specific proteins.