Intrinsic Regulation of Spatiotemporal Organization within the Suprachiasmatic Nucleus

Abstract

The mammalian pacemaker in the suprachiasmatic nucleus (SCN) contains a population of neural oscillators capable of sustaining cell-autonomous rhythms in gene expression and electrical firing. A critical question for understanding pacemaker function is how SCN oscillators are organized into a coherent tissue capable of coordinating circadian rhythms in behavior and physiology. Here we undertake a comprehensive analysis of oscillatory function across the SCN of the adult PER2::LUC mouse by developing a novel approach involving multi-position bioluminescence imaging and unbiased computational analyses. We demonstrate that there is phase heterogeneity across all three dimensions of the SCN that is intrinsically regulated and extrinsically modulated by light in a region-specific manner. By investigating the mechanistic bases of SCN phase heterogeneity, we show for the first time that phase differences are not systematically related to regional differences in period, waveform, amplitude, or brightness. Furthermore, phase differences are not related to regional differences in the expression of arginine vasopressin and vasoactive intestinal polypeptide, two key neuropeptides characterizing functionally distinct subdivisions of the SCN. The consistency of SCN spatiotemporal organization across individuals and across planes of section suggests that the precise phasing of oscillators is a robust feature of the pacemaker important for its function.

Neural correlates of individual differences in circadian behaviour

“…In agreement with the results of a previous study [24], composite maps generated using slices collected from all the mice confirm there are pronounced regional phase differences both across and within rostrocaudal SCN slices (figure 5a). As found in previous detailed analyses [24], one of the most salient organizational features is a late-peaking node located within the rostral slice. Additionally, the middle SCN contained early-peaking regions within the dorsal-most and ventral-most regions and the caudal SCN appeared to be the most homogeneous in phase.…”

“…After imaging, SCN slices were analysed for arginine vasopressin (AVP) and vasoactive intestinal polypeptide (VIP) expression as in [24]. Briefly, slices were treated with colchicine-treated medium (25 mg ml 21 ), fixed with 4% paraformaldehyde, and then cyprotected.…”

“…(ii) Criteria for slice inclusion As in [24], consistency in slice position was assessed using a metric based on the bioluminescence profile of each slice (figure 1c). For each slice, bioluminescence was summed over the first cycle in vitro, from which the width/height ratio of the bioluminescence profile was calculated ( figure 1g).…”

Topological specificity and hierarchical network of the circadian calcium rhythm in the suprachiasmatic nucleus

“…7C, and Tables S3 and S4). patterns of clock gene expression using in situ hybridization and bioluminescence imaging (2,(10)(11)(12)(13)(14). However, the autonomous expression of a clock gene is either absent or undetectable in certain regions, particularly those where neurons receive major light information from the retina (18).…”

“…Heterogeneous distribution of the circadian phase in SCN slices was shown by in situ hybridization and bioluminescence studies examining clock gene expression (e.g., Per1 or Per2) (10)(11)(12)(13)(14). Disruption of intercellular coupling by inhibiting neuronal firing with tetrodotoxin (TTX) or mechanical isolation of SCN subregions severely reduced clock gene expression and mutual synchronization (10,11).…”