Rhythmic clock and neuropeptide gene expression in hypothalamic mHypoE-44 neurons

Abstract

The rhythmic expression of specific clock genes: Bmal1, Per2, Clock and Rev-Erbalpha; and specific hypothalamic neuropeptides: NPY, Crh, AgRP, neurotensin and preproghrelin, expressed in clonal hypothalamic neuronal cell lines, was assayed and analyzed using a novel non-linear least squares statistical analysis to determine rhythmicity in an in vitro milieu. In silico analysis of the neuropeptide promoter regions identified putative E-box motifs and a motif in the NPY promoter is bound to in an oscillatory fashion. Within the mHypoE-44 neurons, we demonstrate that mRNA of four core circadian components: Bmal1, Clock, Per2 and Rev-Erbalpha, oscillate with approximate 24h rhythms. NPY and NT demonstrated significant 24h gene expression. However, CRH and preproghrelin mRNA cycled significantly in an ultradian fashion, oscillating approximately every 18h. AgRP mRNA did not show a significant rhythm. We speculate that endogenous rhythmic neuropeptide expression contributes to neuroendocrine homeostasis, which may include energy balance.