Epigenetic suppression of mouse Per2 expression in the suprachiasmatic nucleus by the inhalational anesthetic, sevoflurane

PLoS One. 2014 Jan 31;9(1):e87319. doi: 10.1371/journal.pone.0087319. eCollection 2014.


Background: We previously reported that sevoflurane anesthesia reversibly suppresses the expression of the clock gene, Period2 (Per2), in the mouse suprachiasmatic nucleus (SCN). However, the molecular mechanisms underlying this suppression remain unclear. In this study, we examined the possibility that sevoflurane suppresses Per2 expression via epigenetic modification of the Per2 promoter.

Methods: Mice were anesthetized with a gas mixture of 2.5% sevoflurane/40% oxygen at a 6 L/min flow for 1 or 4 h. After termination, brains were removed and samples of SCN tissue were derived from frozen brain sections. Chromatin immunoprecipitation (ChIP) assays using anti-acetylated-histone antibodies were performed to investigate the effects of sevoflurane on histone acetylation of the Per2 promoter. Interaction between the E'-box (a cis-element in the Per2 promoter) and CLOCK (the Clock gene product) was also assessed by a ChIP assay using an anti-CLOCK antibody. The SCN concentration of nicotinamide adenine dinucleotide (NAD(+)), a CLOCK regulator, was assessed by liquid chromatography-mass spectrometry.

Results: Acetylation of histone H4 in the proximal region of the Per2 promoter was significantly reduced by sevoflurane. This change in the epigenetic profile of the Per2 gene was observed prior to suppression of Per2 expression. Simultaneously, a reduction in the CLOCK-E'-box interaction in the Per2 promoter was observed. Sevoflurane treatment did not affect the concentration of NAD(+) in the SCN.

Conclusions: Independent of NAD(+) concentration in the SCN, sevoflurane decreases CLOCK binding to the Per2 promoter E'-box motif, reducing histone acetylation and leading to suppression of Per2 expression.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ARNTL Transcription Factors / metabolism
  • Acetylation / drug effects
  • Anesthetics, Inhalation / pharmacology
  • Animals
  • Binding Sites / genetics
  • Brain / metabolism
  • CLOCK Proteins / metabolism
  • DNA Methylation / drug effects
  • Epigenesis, Genetic / drug effects*
  • Gene Expression / drug effects
  • HSP90 Heat-Shock Proteins / genetics
  • Histones / metabolism
  • Male
  • Methyl Ethers / pharmacology*
  • Mice
  • Mice, Inbred C57BL
  • NAD / metabolism
  • Period Circadian Proteins / genetics*
  • Promoter Regions, Genetic / genetics
  • Protein Binding
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sevoflurane
  • Suprachiasmatic Nucleus / metabolism*
  • Time Factors


  • ARNTL Transcription Factors
  • Anesthetics, Inhalation
  • Arntl protein, mouse
  • HSP90 Heat-Shock Proteins
  • Histones
  • Methyl Ethers
  • Per2 protein, mouse
  • Period Circadian Proteins
  • NAD
  • Sevoflurane
  • CLOCK Proteins

Grant support

This study was supported by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) Japan, the MEXT-Supported Program for Strategic Research Foundation at Private Universities (S0801035 to H.O.), and Grants-in-Aid for Scientific Research (22590230 to H.O.; 23659125 to N.I.; 24592318 to A.S.; and 24790241 to S.H.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.