Ghrelin effects on the circadian system of mice.

The orexigenic peptide ghrelin stimulates both food intake and growth hormone release and is synthesized in the stomach and in hypothalamic areas involved in feeding control. The suprachiasmatic nuclei of the hypothalamus (SCN) control most circadian rhythms, although there is evidence that some oscillators, such as food-entrainable oscillators, can drive activity rhythms even after SCN ablation. Ghrelin levels exhibit a circadian rhythm and closely follow feeding schedules, making this peptide a putative candidate for food-related entraining signals. We examined the response of the SCN to ghrelin treatments in vitro, by means of electrophysiological and bioluminescence recordings, and in vivo, by assessing effects on the phase of locomotor activity rhythms. Ghrelin applied at circadian time 6 in vitro to cultured SCN slices induced an approximately 3 h phase advance. In addition, ghrelin phase advanced the rhythm of PER2::LUC (Period2::Luciferase) expression in cultured SCN explants from mPer2(Luc) transgenic mice. In vivo, intraperitoneal administration of ghrelin or a synthetic analog, growth hormone-releasing protein-6 (GHRP-6), to ad libitum fed animals failed to alter circadian phase. When injected after 30 h of food deprivation, GHRP-6 induced a phase advance compared with saline-injected animals. These results indicate that ghrelin may play a role in the circadian system by exerting a direct action on the SCN and that the system as a whole may become sensitive to ghrelin and other feeding-related neuropeptides under conditions of food restriction.

Neuropeptide Y differentially suppresses per1 and per2 mRNA induced by light in the suprachiasmatic nuclei of the golden hamster.

Neuropeptide Y (NPY), present in an input pathway to the suprachiasmatic nuclei (SCN), can block the effects of light on circadian rhythms. The authors have studied this interaction using an in vitro brain slice technique. Effects of NPY on light-induced period1 and period2 mRNA in the SCN were examined in vitro following a light pulse during early subjective night. Golden hamsters (n = 91) were housed under a 14:10 LD cycle and then moved to constant dim red light for 3 days. Hamsters were exposed to a 5-min light pulse previously shown to induce phase shifts and prepared for in vitro application of NPY. Hypothalamic slices containing the SCN were maintained in vitro for 40 min to 4 h after the light pulse, then quick-frozen. Sections were evaluated by in situ hybridization with [35S]-labeled cRNA probes for per mRNA. Rapid light induction of both per1 and per2 by 40 min and 1 h after the light pulse, respectively, was apparent, with NPY inhibition of this response significant by at least these same time points. However, although striking suppression of per2 mRNA by the NPY continued through the peak for per2 at 2 h, per1 mRNA levels rebounded quickly to equal the per1 induction peak at 1 h and mirrored the control light induction pattern for per1 thereafter. Delaying NPY to 30 min after slice preparation demonstrated that NPY is capable of suppressing peak per1 levels. These results confirm the feasibility of measuring light-induced gene expression in the SCN in vitro. A differential regulation of per1 and per2 transcription might be of critical importance for the modulation of circadian responses to light.