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

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 u...

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Autores principales: Wegner, Sven, Belle, Mino D.C., Hughes, Alun T.L., Diekman, Casey O., Piggins, Hugh D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Society for Neuroscience 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559760/
https://www.ncbi.nlm.nih.gov/pubmed/28698388
http://dx.doi.org/10.1523/JNEUROSCI.0691-17.2017
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author Wegner, Sven
Belle, Mino D.C.
Hughes, Alun T.L.
Diekman, Casey O.
Piggins, Hugh D.
author_facet Wegner, Sven
Belle, Mino D.C.
Hughes, Alun T.L.
Diekman, Casey O.
Piggins, Hugh D.
author_sort Wegner, Sven
collection PubMed
description 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 Afterhours mice which carry the circadian period lengthening loss-of-function Fbxl3(Afh) 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 Fbxl3(Afh/Afh) mice. At night, vSCN cells from Fbxl3(Afh/Afh) mice were more hyperpolarized, receiving more GABAergic input than their Fbxl3(+/+) counterparts. Unexpectedly, the progression to daytime hyperexcited states was slowed by Afh mutation, whereas the decline to hypoexcited states was accelerated. In long-term bioluminescence recordings, GABA(A) receptor blockade desynchronized the Fbxl3(+/+) but not the Fbxl3(Afh/Afh) vSCN neuronal network. Further, a neurochemical mimic of the light input pathway evoked larger shifts in molecular clock rhythms in Fbxl3(Afh/Afh) compared with Fbxl3(+/+) SCN slices. These results reveal unanticipated consequences of delaying CRY degradation, indicating that the Afh mutation prolongs nighttime hyperpolarized states of vSCN cells through increased GABAergic synaptic transmission. SIGNIFICANCE STATEMENT 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.
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spelling pubmed-55597602017-08-22 Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability Wegner, Sven Belle, Mino D.C. Hughes, Alun T.L. Diekman, Casey O. Piggins, Hugh D. J Neurosci Research Articles 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 Afterhours mice which carry the circadian period lengthening loss-of-function Fbxl3(Afh) 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 Fbxl3(Afh/Afh) mice. At night, vSCN cells from Fbxl3(Afh/Afh) mice were more hyperpolarized, receiving more GABAergic input than their Fbxl3(+/+) counterparts. Unexpectedly, the progression to daytime hyperexcited states was slowed by Afh mutation, whereas the decline to hypoexcited states was accelerated. In long-term bioluminescence recordings, GABA(A) receptor blockade desynchronized the Fbxl3(+/+) but not the Fbxl3(Afh/Afh) vSCN neuronal network. Further, a neurochemical mimic of the light input pathway evoked larger shifts in molecular clock rhythms in Fbxl3(Afh/Afh) compared with Fbxl3(+/+) SCN slices. These results reveal unanticipated consequences of delaying CRY degradation, indicating that the Afh mutation prolongs nighttime hyperpolarized states of vSCN cells through increased GABAergic synaptic transmission. SIGNIFICANCE STATEMENT 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. Society for Neuroscience 2017-08-16 /pmc/articles/PMC5559760/ /pubmed/28698388 http://dx.doi.org/10.1523/JNEUROSCI.0691-17.2017 Text en Copyright © 2017 Wegner et al. https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License Creative Commons Attribution 4.0 International (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.
spellingShingle Research Articles
Wegner, Sven
Belle, Mino D.C.
Hughes, Alun T.L.
Diekman, Casey O.
Piggins, Hugh D.
Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title_full Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title_fullStr Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title_full_unstemmed Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title_short Delayed Cryptochrome Degradation Asymmetrically Alters the Daily Rhythm in Suprachiasmatic Clock Neuron Excitability
title_sort delayed cryptochrome degradation asymmetrically alters the daily rhythm in suprachiasmatic clock neuron excitability
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559760/
https://www.ncbi.nlm.nih.gov/pubmed/28698388
http://dx.doi.org/10.1523/JNEUROSCI.0691-17.2017
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