Separate optogenetic manipulation of Nerve/glial antigen 2 (NG2) glia and mural cells using the NG2 promoter

Mitsuhiro Oishi, Stefan Passlick, Yoshihiko Yamazaki, Miyuki Unekawa, Ruka Adachi, Mayumi Yamada, Itaru Imayoshi, Yoshifumi Abe, Christian Steinhäuser, Kenji F. Tanaka

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)


Nerve/glial antigen 2 (NG2) is a protein marker of NG2 glia and mural cells, and NG2 promoter activity is utilized to target these cells. However, the NG2 promoter cannot target NG2 glia and mural cells separately. This has been an obstacle for NG2 glia-specific manipulation. Here, we developed transgenic mice in which either cell type can be targeted using the NG2 promoter. We selected a tetracycline-controllable gene induction system for cell type-specific transgene expression, and generated NG2-tetracycline transactivator (tTA) transgenic lines. We crossed tTA lines with the tetO-ChR2 (channelrhodopsin-2)-EYFP line to characterize tTA-dependent transgene induction. We isolated two unique NG2-tTA mouse lines: one that induced ChR2-EYFP only in mural cells, likely due to the chromosomal position effect of NG2-tTA insertion, and the other that induced it in both cell types. We then applied a Cre-mediated set-subtraction strategy to the latter case and eliminated ChR2-EYFP from mural cells, resulting in NG2 glia-specific transgene induction. We further demonstrated that tTA-dependent ChR2 expression could manipulate cell function. Optogenetic mural cell activation decreased cerebral blood flow, as previously reported, indicating that tTA-mediated ChR2 expression was sufficient to impact cellular function. ChR2-mediated depolarization was observed in NG2 glia in acute hippocampal slices. In addition, ChR2-mediated depolarization of NG2 glia inhibited their proliferation but promoted their differentiation in juvenile mice. Since the tTA–tetO combination is expandable, the mural cell–specific NG2-tTA line and the NG2 glia-specific NG2-tTA line will permit us to conduct observational and manipulation studies to examine in vivo function of these cells separately.

Original languageEnglish
Pages (from-to)317-333
Number of pages17
Issue number2
Publication statusPublished - 2023 Feb


  • DM20
  • cAMP
  • mural cell
  • oligodendrocyte precursor cell
  • tet-system

ASJC Scopus subject areas

  • Neurology
  • Cellular and Molecular Neuroscience


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