MOLECULAR PHYSIOLOGY OF CIRCADIAN PACEMAKING

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Many critical daily rhythms are endogenous, driven by an internal master pacemaker or biological clock, with a free-running period of about a day (i.e. circadian). The suprachiasmatic nuclei (SCN) of the hypothalamus are the apparent site of the master biological clock in mammals. Recent research has revealed that (i) the ability to generate neurophysiological circadian rhythms resides in single SCN neurons, and (ii) the fundamental mechanisms for rhythms generation within these neurons involve transcriptional-translational negative feedback loops among circadian clock genes. A critical challenge for circadian neurobiology is to define the links between the intracellular molecular clock with its neurophysiological inputs and outputs. Toward this end we have created transgenic mice incorporating a circadian reporter construct which expresses a degradable form GFP under the control of the mPer1 promoter. We will combine our ability to visualize the cellular dynamics of per1 promoter activation in the living SCN with double label immunocytochemistry, patch clamp electrophysiology, gene expression profiling and multielectrode array recording to (i) identify light- entrainment-transducing SCN neurons and the physiological and molecular events which occur in them during the phase-resetting process, (ii) measure individual neuronal rhythms in SCN slices en masse to provide a novel view of pacemaker structure, (iii) identify specific populations of rhythmic SCN neurons and examine the mechanisms by which the molecular clock loop is output to the cell membrane. Successful completion of these aims will elucidate critical links between gene regulation and circadian function of the SCN. This will increase our understanding of the fundamental mechanisms underlying normal biological clock function and our understanding of disease processes in which the clock is deranged.