Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability

• Published in: The Journal of neuroscience Vol. 37; no. 33; pp. 7824 - 7836
• Main Authors: Wegner, Sven, Belle, Mino D C, Hughes, Alun T L, Diekman, Casey O, Piggins, Hugh D
• Format: Journal Article
• Language: English
• Published: United States Society for Neuroscience 16-08-2017
• Subjects: Animals; Bioluminescence; Brain; Brain slice preparation; Circadian Clocks - physiology; Circadian rhythm; Circadian Rhythm - physiology; Circadian rhythms; Cryptochromes; Cryptochromes - genetics; Cryptochromes - metabolism; Degradation; Excitability; Female; Male; Mice; Mice, Transgenic; Mutation; Mutation - physiology; Neurons; Neurons - metabolism; Neurophysiology; Night; Organ Culture Techniques; Post-translation; Repressors; Suprachiasmatic Nucleus - metabolism; Synaptic transmission; Synchronization; Time Factors; Transcription factors; Ubiquitin; Ubiquitin-protein ligase; Ubiquitination; γ-Aminobutyric acid A receptors; brain slice; circadian; electrophysiology; cryptochrome; fbxl3; post-translational modification; suprachiasmatic
• ISSN: 0270-6474; 1529-2401
• DOI: 10.1523/JNEUROSCI.0691-17.2017

Suprachiasmatic nuclei (SCN) neurons contain an intracellular molecular circadian clock and the Cryptochromes (CRY1/2), key transcriptional repressors of this molecular apparatus, are subject to post-translational modification through ubiquitination and targeting for proteosomal degradation by the ubiquitin E3 ligase complex. Loss-of-function point mutations in a component of this ligase complex, Fbxl3, delay CRY1/2 degradation, reduce circadian rhythm strength, and lengthen the circadian period by ∼2.5 h. The molecular clock drives circadian changes in the membrane properties of SCN neurons, but it is unclear how alterations in CRY1/2 stability affect SCN neurophysiology. Here we use male and female
mice which carry the circadian period lengthening loss-of-function
mutation and perform patch-clamp recordings from SCN brain slices across the projected day/night cycle. We find that the daily rhythm in membrane excitability in the ventral SCN (vSCN) was enhanced in amplitude and delayed in timing in
mice. At night, vSCN cells from
mice were more hyperpolarized, receiving more GABAergic input than their
counterparts. Unexpectedly, the progression to daytime hyperexcited states was slowed by
mutation, whereas the decline to hypoexcited states was accelerated. In long-term bioluminescence recordings, GABA
receptor blockade desynchronized the
but not the
vSCN neuronal network. Further, a neurochemical mimic of the light input pathway evoked larger shifts in molecular clock rhythms in
compared with
SCN slices. These results reveal unanticipated consequences of delaying CRY degradation, indicating that the
mutation prolongs nighttime hyperpolarized states of vSCN cells through increased GABAergic synaptic transmission.
The intracellular molecular clock drives changes in SCN neuronal excitability, but it is unclear how mutations affecting post-translational modification of molecular clock proteins influence the temporal expression of SCN neuronal state or intercellular communication within the SCN network. Here we show for the first time, that a mutation that prolongs the stability of key components of the intracellular clock, the cryptochrome proteins, unexpectedly increases in the expression of hypoexcited neuronal state in the ventral SCN at night and enhances hyperpolarization of ventral SCN neurons at this time. This is accompanied by increased GABAergic signaling and by enhanced responsiveness to a neurochemical mimic of the light input pathway to the SCN. Therefore, post-translational modification shapes SCN neuronal state and network properties.