Astrocytes Sustain Circadian Oscillation and Bidirectionally Determine Circadian Period, But Do Not Regulate Circadian Phase in the Suprachiasmatic Nucleus

Abstract

The suprachiasmatic nucleus (SCN) is the master circadian clock of mammals, generating and transmitting an internal representation of environmental time that is produced by the cell-autonomous transcriptional/post-translational feedback loops (TTFLs) of the 10,000 neurons and 3500 glial cells. Recently, we showed that TTFL function in SCN astrocytes alone is sufficient to drive circadian timekeeping and behavior, raising questions about the respective contributions of astrocytes and neurons within the SCN circuit. We compared their relative roles in circadian timekeeping in mouse SCN explants, of either sex. Treatment with the glial-specific toxin fluorocitrate revealed a requirement for metabolically competent astrocytes for circuit-level timekeeping. Recombinase-mediated genetically complemented Cryptochrome (Cry) proteins in Cry1-deficient and/or Cry2-deficient SCNs were used to compare the influence of the TTFLs of neurons or astrocytes in the initiation of de novo oscillation or in pacemaking. While neurons and astrocytes both initiated de novo oscillation and lengthened the period equally, their kinetics were different, with astrocytes taking twice as long. Furthermore, astrocytes could shorten the period, but not as potently as neurons. Chemogenetic manipulation of Gi- and Gq-coupled signaling pathways in neurons acutely advanced or delayed the ensemble phase, respectively. In contrast, comparable manipulations in astrocytes were without effect. Thus, astrocytes can initiate SCN rhythms and bidirectionally control the SCN period, albeit with lower potency than neurons. Nevertheless, their activation does not influence the SCN phase. The emergent SCN properties of high-amplitude oscillation, initiation of rhythmicity, pacemaking, and phase are differentially regulated: astrocytes and neurons sustain the ongoing oscillation, but its phase is determined by neurons. SIGNIFICANCE STATEMENT The hypothalamic suprachiasmatic nucleus (SCN) encodes and disseminates time-of-day information to allow mammals to adapt their physiology to daily environmental cycles. Recent investigations have revealed a role for astrocytes, in addition to neurons, in the regulation of this rhythm. Using pharmacology, genetic complementation, and chemogenetics, we compared the abilities of neurons and astrocytes in determining the emergent SCN properties of high-amplitude oscillation, initiation of rhythmicity, pacemaking, and determination of phase. These findings parameterize the circadian properties of the astrocyte population in the SCN and reveal the types of circadian information that astrocytes and neurons can contribute within their heterogeneous cellular network.

Astrocytic control of extra-cellular GABA drives circadian time-keeping in the suprachiasmatic nucleus

“…Due to the fact that the GAT3 transporter is integral to neuropeptide release from SCN neurons to ensure correct network function, and that it appears to be an astrocyte-enriched transporter and circadian-regulated, we next explored whether the circadian clock of astrocytes could, autonomously, control [GABA] e . We have previously used CRY1-complementation targeted to astrocytes to control circadian network function by initiating oscillations in arrhythmic CRY1,2-null SCN (Brancaccio et al, 2019; Patton et al, 2022). Using this approach, however, initiation takes >7 days (Patton et al ., 2022).…”

A role for caveolar proteins in regulation of the circadian clock

“…Recent evidence supports an important role of the astrocytes in the SCN. The circadian clock in astrocytes is required for normal circadian rhythms (Barca-Mayo et al, 2017, Brancaccio et al, 2017, Tso et al, 2017) and astrocytes are important to generate the rhythm and keep the synchrony of surrounding neurons, particularly through Ca 2+ signalling (Brancaccio et al, 2017, Patton et al, 2022, Brancaccio et al, 2019). The response of astrocytes to external stimuli involves the intricate relationship between caveolae and fatty acid-binding protein 7 (FABP7) through the formation and function of lipid microdomains (‘rafts’) (Kagawa et al, 2015).…”

Circadian Control of Glial Cell Homeodynamics

“…Moreover, in the absence of Cry1-2 expression in neurons, astrocyte clocks alone can autonomously initiate and modulate self-sustained circadian oscillations in the SCN (Brancaccio et al, 2019). However, astrocytes alone cannot influence SCN phase, as this is only regulated by neurons (Patton et al, 2022). Notably, as opposed to eliminating Bmal1 , the lack of Per2 only in GFAP + cells does not alter circadian period or locomotor activity (Martini et al, 2021), further emphasizing the importance of extra-circadian signaling roles of these clock genes.…”