Abstract

Despite rhythmic clock gene expression being found throughout the central nervous system, very little is known about their function outside of the suprachiasmatic nucleus. Understanding how clock genes are expressed across a variety of neural cell types is important to elucidating how these clock genes are regulated and how they may influence the function of each brain region. Using immunofluorescence and confocal microscopy, we quantified the co-expression of the clock proteins BMAL1 and PER2 with Substance P (SubP) and Enkephalin (Enk). Regions examined included the limbic forebrain (the dorsal striatum, ventral striatum, amygdala, stria terminalis), the thalamus (medial habenula), the hypothalamus (paraventricular nucleus, arcuate nucleus) and the olfactory bulb. In most regions examined, PER2 and BMAL1 were homogenously expressed in nearly all cells (~90%), despite very different expression profiles of SubP or Enk in each nucleus. In nuclei that expressed both SubP and Enk, PER2 and BMAL1 were not preferentially co-expressed in one cell type or the other. The olfactory bulb was unique and expressed PER2 or BMAL1 in a much smaller percentage of cells, and Enk was rarely found in the same cells that expressed the clock proteins (SubP was undetectable). This indicates that clock genes are not unique to specific cell types and further studies will be required to address which factors contribute to clock gene rhythmicity. A focus on network interactions is recommended in order to determine how different cell types contribute to various aspects of rhythmicity, such as rhythm generation, amplitude and synchrony.