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A diversity of paracrine signals sustains molecular circadian cycling in suprachiasmatic nucleus circuits.

ABSTRACT: The suprachiasmatic nucleus (SCN) is the principal circadian pacemaker of mammals, coordinating daily rhythms of behavior and metabolism. Circadian timekeeping in SCN neurons revolves around transcriptional/posttranslational feedback loops, in which Period (Per) and Cryptochrome (Cry) genes are negatively regulated by their protein products. Recent studies have revealed, however, that these "core loops" also rely upon cytosolic and circuit-level properties for sustained oscillation. To characterize interneuronal signals responsible for robust pacemaking in SCN cells and circuits, we have developed a unique coculture technique using wild-type (WT) "graft" SCN to drive pacemaking (reported by PER2::LUCIFERASE bioluminescence) in "host" SCN deficient either in elements of neuropeptidergic signaling or in elements of the core feedback loop. We demonstrate that paracrine signaling is sufficient to restore cellular synchrony and amplitude of pacemaking in SCN circuits lacking vasoactive intestinal peptide (VIP). By using grafts with mutant circadian periods we show that pacemaking in the host SCN is specified by the genotype of the graft, confirming graft-derived factors as determinants of the host rhythm. By combining pharmacological with genetic manipulations, we show that a hierarchy of neuropeptidergic signals underpins this paracrine regulation, with a preeminent role for VIP augmented by contributions from arginine vasopressin (AVP) and gastrin-releasing peptide (GRP). Finally, we show that interneuronal signaling is sufficiently powerful to maintain circadian pacemaking in arrhythmic Cry-null SCN, deficient in essential elements of the transcriptional negative feedback loops. Thus, a hierarchy of paracrine neuropeptidergic signals determines cell- and circuit-level circadian pacemaking in the SCN.

SUBMITTER: Maywood ES

PROVIDER: S-EPMC3161534 | biostudies-other | 2011 Aug

REPOSITORIES: biostudies-other

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A diversity of paracrine signals sustains molecular circadian cycling in suprachiasmatic nucleus circuits.

Maywood Elizabeth S ES Chesham Johanna E JE O'Brien John A JA Hastings Michael H MH

Proceedings of the National Academy of Sciences of the United States of America 20110725 34

The suprachiasmatic nucleus (SCN) is the principal circadian pacemaker of mammals, coordinating daily rhythms of behavior and metabolism. Circadian timekeeping in SCN neurons revolves around transcriptional/posttranslational feedback loops, in which Period (Per) and Cryptochrome (Cry) genes are negatively regulated by their protein products. Recent studies have revealed, however, that these "core loops" also rely upon cytosolic and circuit-level properties for sustained oscillation. To character ...[more]

PMID: 21788520

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Project description:The suprachiasmatic nucleus (SCN) of the hypothalamus is the principal circadian pacemaker of the brain. It co-ordinates the daily rhythms of sleep and wakefulness, as well as physiology and behaviour, that set the tempo to our lives. Disturbance of this daily pattern, most acutely with jet-lag but more insidiously with rotational shift-work, can have severely deleterious effects for mental function and long-term health. The present review considers recent developments in our understanding of the properties of the SCN that make it a robust circadian time-keeper. It first focuses on the intracellular transcriptional/ translational feedback loops (TTFL) that constitute the cellular clockwork of the SCN neurone. Daily timing by these loops pivots around the negative regulation of the Period (Per) and Cryptochrome (Cry) genes by their protein products. The period of the circadian cycle is set by the relative stability of Per and Cry proteins, and this can be controlled by both genetic and pharmacological interventions. It then considers the function of these feedback loops in the context of cytosolic signalling by cAMP and intracellular calcium ([Ca(2+) ]i ), which are both outputs from, and inputs to, the TTFL, as well as the critical role of vasoactive intestinal peptide (VIP) signalling in synchronising cellular clocks across the SCN. Synchronisation by VIP in the SCN is paracrine, operating over an unconventionally long time frame (i.e. 24 h) and wide spatial domain, mediated via the cytosolic pathways upstream of the TTFL. Finally, we show how intersectional pharmacogenetics can be used to control G-protein-coupled signalling in individual SCN neurones, and how manipulation of Gq/[Ca(2+) ]i -signalling in VIP neurones can re-programme the circuit-level encoding of circadian time. Circadian pacemaking in the SCN therefore provides an unrivalled context in which to understand how a complex, adaptive behaviour can be organised by the dynamic activity of a relatively few gene products, operating in a clearly defined neuronal circuit, with both cell-autonomous and emergent, circuit-level properties.

Project description:In mammals, the suprachiasmatic nucleus (SCN) of the hypothalamus coordinates daily rhythms including sleep-wake, hormone release, and gene expression. The cells of the SCN must synchronize to each other to drive these circadian rhythms in the rest of the body. The ontogeny of circadian cycling and intercellular coupling in the SCN remains poorly understood. Recent in vitro studies have recorded circadian rhythms from the whole embryonic SCN. Here, we tracked the onset and precision of rhythms in PERIOD2 (PER2), a clock protein, within the SCN isolated from embryonic and postnatal mice of undetermined sex. We found that a few SCN cells developed circadian periodicity in PER2 by 14.5 d after mating (E14.5) with no evidence for daily cycling on E13.5. On E15.5, the fraction of competent oscillators increased dramatically corresponding with stabilization of their circadian periods. The cells of the SCN harvested at E15.5 expressed sustained, synchronous daily rhythms. By postnatal day 2 (P2), SCN oscillators displayed the daily, dorsal-ventral phase wave in clock gene expression typical of the adult SCN. Strikingly, vasoactive intestinal polypeptide (VIP), a neuropeptide critical for synchrony in the adult SCN, and its receptor, VPAC2R, reached detectable levels after birth and after the onset of circadian synchrony. Antagonists of GABA or VIP signaling or action potentials did not disrupt circadian synchrony in the E15.5 SCN. We conclude that endogenous daily rhythms in the fetal SCN begin with few noisy oscillators on E14.5, followed by widespread oscillations that rapidly synchronize on E15.5 by an unknown mechanism.SIGNIFICANCE STATEMENT We recorded the onset of PER2 circadian oscillations during embryonic development in the mouse SCN. When isolated at E13.5, the anlagen of the SCN expresses high, arrhythmic PER2. In contrast, a few cells show noisy circadian rhythms in the isolated E14.5 SCN and most show reliable, self-sustained, synchronized rhythms in the E15.5 SCN. Strikingly, this synchrony at E15.5 appears before expression of VIP or its receptor and persists in the presence of blockers of VIP, GABA or neuronal firing. Finally, the dorsal-ventral phase wave of PER2 typical of the adult SCN appears ?P2, indicating that multiple signals may mediate circadian synchrony during the ontogeny of the SCN.

Project description:The suprachiasmatic nuclei (SCN) host a robust, self-sustained circadian pacemaker that coordinates physiological rhythms with the daily changes in the environment. Neuronal clocks within the SCN form a heterogeneous network that must synchronize to maintain timekeeping activity. Coherent circadian output of the SCN tissue is established by intercellular signaling factors, such as vasointestinal polypeptide. It was recently shown that besides coordinating cells, the synchronization factors play a crucial role in the sustenance of intrinsic cellular rhythmicity. Disruption of intercellular signaling abolishes sustained rhythmicity in a majority of neurons and desynchronizes the remaining rhythmic neurons. Based on these observations, the authors propose a model for the synchronization of circadian oscillators that combines intracellular and intercellular dynamics at the single-cell level. The model is a heterogeneous network of circadian neuronal oscillators where individual oscillators are damped rather than self-sustained. The authors simulated different experimental conditions and found that: (1) in normal, constant conditions, coupled circadian oscillators quickly synchronize and produce a coherent output; (2) in large populations, such oscillators either synchronize or gradually lose rhythmicity, but do not run out of phase, demonstrating that rhythmicity and synchrony are codependent; (3) the number of oscillators and connectivity are important for these synchronization properties; (4) slow oscillators have a higher impact on the period in mixed populations; and (5) coupled circadian oscillators can be efficiently entrained by light-dark cycles. Based on these results, it is predicted that: (1) a majority of SCN neurons needs periodic synchronization signal to be rhythmic; (2) a small number of neurons or a low connectivity results in desynchrony; and (3) amplitudes and phases of neurons are negatively correlated. The authors conclude that to understand the orchestration of timekeeping in the SCN, intracellular circadian clocks cannot be isolated from their intercellular communication components.

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A diversity of paracrine signals sustains molecular circadian cycling in suprachiasmatic nucleus circuits.

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A diversity of paracrine signals sustains molecular circadian cycling in suprachiasmatic nucleus circuits.

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