Expression Patterns of Inducible Cre Recombinase Driven by Differential Astrocyte-Specific Promoters in Transgenic Mouse Lines

  • Neng-Yuan Hu
  • Ya-Ting Chen
  • Qian Wang
  • Wei Jie
  • Yi-Si Liu
  • Qiang-Long You
  • Ze-Lin Li
  • Xiao-Wen Li
  • Sophie Reibel
  • Frank W. Pfrieger
  • Jian-Ming YangEmail author
  • Tian-Ming GaoEmail author


Astrocytes are the most abundant cell type in the central nervous system (CNS). They provide trophic support for neurons, modulate synaptic transmission and plasticity, and contribute to neuronal dysfunction. Many transgenic mouse lines have been generated to obtain astrocyte-specific expression of inducible Cre recombinase for functional studies; however, the expression patterns of inducible Cre recombinase in these lines have not been systematically characterized. We generated a new astrocyte-specific Aldh1l1-CreERT2 knock-in mouse line and compared the expression pattern of Cre recombinase between this and five widely-used transgenic lines (hGfap-CreERT2 from The Jackson Laboratory and The Mutant Mouse Resource and Research Center, Glast-CreERT2, Cx30-CreERT2, and Fgfr3-iCreERT2) by crossing with Ai14 mice, which express tdTomato fluorescence following Cre-mediated recombination. In adult Aldh1l1-CreERT2:Ai14 transgenic mice, tdTomato was detected throughout the CNS, and five novel morphologically-defined types of astrocyte were described. Among the six evaluated lines, the specificity of Cre-mediated recombination was highest when driven by Aldh1l1 and lowest when driven by hGfap; in the latter mice, co-staining between tdTomato and NeuN was observed in the hippocampus and cortex. Notably, evident leakage was noted in Fgfr3-iCreERT2 mice, and the expression level of tdTomato was low in the thalamus when Cre recombinase expression was driven by Glast and in the capsular part of the central amygdaloid nucleus when driven by Cx30. Furthermore, tdTomato was clearly expressed in peripheral organs in four of the lines. Our results emphasize that the astrocyte-specific CreERT2 transgenic lines used in functional studies should be carefully selected.


Astrocytes Cre recombinase Expression pattern Aldh1l1 Morphology 



This work was supported by Grants from the National Natural Science Foundation of China (31430032, 31830033, 81971080, and 81671356), the Program for Changjiang Scholars and Innovative Research Teams in University (IRT_16R37), the Science and Technology Program of Guangdong (2018B030334001), and the Guangzhou Science and Technology Project (201707020027, 201704020116). Thanks to Professor William D. Richardson (University College London, UK) for the Fgfr3-iCreERT2 line.

Conflict of interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Supplementary material

12264_2019_451_MOESM1_ESM.pdf (5.5 mb)
Supplementary material 1 (PDF 5617 kb)


  1. 1.
    Herculano-Houzel S. The glia/neuron ratio: how it varies uniformly across brain structures and species and what that means for brain physiology and evolution. Glia 2014, 62: 1377–1391.PubMedCrossRefPubMedCentralGoogle Scholar
  2. 2.
    Rowitch DH, Kriegstein AR. Developmental genetics of vertebrate glial-cell specification. Nature 2010, 468: 214–222.PubMedCrossRefPubMedCentralGoogle Scholar
  3. 3.
    Verkhratsky AN, Butt A. Glial Physiology and Pathophysiology. Chichester, West Sussex, UK; Hoboken, NJ, USA: Wiley-Blackwell, 2013.CrossRefGoogle Scholar
  4. 4.
    Bailey MS, Shipley MT. Astrocyte subtypes in the rat olfactory bulb: morphological heterogeneity and differential laminar distribution. J Comp Neurol 1993, 328: 501–526.PubMedCrossRefPubMedCentralGoogle Scholar
  5. 5.
    Emsley JG, Macklis JD. Astroglial heterogeneity closely reflects the neuronal-defined anatomy of the adult murine CNS. Neuron Glia Biol 2006, 2: 175–186.PubMedPubMedCentralCrossRefGoogle Scholar
  6. 6.
    Buosi AS, Matias I, Araujo APB, Batista C, Gomes FCA. Heterogeneity in synaptogenic profile of astrocytes from different brain regions. Mol Neurobiol 2018, 55: 751–762.PubMedCrossRefPubMedCentralGoogle Scholar
  7. 7.
    Ben Haim L, Rowitch DH. Functional diversity of astrocytes in neural circuit regulation. Nat Rev Neurosci 2017, 18: 31–41.PubMedCrossRefPubMedCentralGoogle Scholar
  8. 8.
    Morel L, Men Y, Chiang MSR, Tian Y, Jin S, Yelick J, et al. Intracortical astrocyte subpopulations defined by astrocyte reporter mice in the adult brain. Glia 2019, 67: 171–181.PubMedCrossRefPubMedCentralGoogle Scholar
  9. 9.
    Zhang Y, Barres BA. Astrocyte heterogeneity: an underappreciated topic in neurobiology. Curr Opin Neurobiol 2010, 20: 588–594.PubMedCrossRefPubMedCentralGoogle Scholar
  10. 10.
    Wang Q, Jie W, Liu JH, Yang JM, Gao TM. An astroglial basis of major depressive disorder? An overview. Glia 2017, 65: 1227–1250.PubMedPubMedCentralGoogle Scholar
  11. 11.
    Cui Y, Yang Y, Ni Z, Dong Y, Cai G, Foncelle A, et al. Astroglial Kir4.1 in the lateral habenula drives neuronal bursts in depression. Nature 2018, 554: 323–327.PubMedCrossRefPubMedCentralGoogle Scholar
  12. 12.
    Tso CF, Simon T, Greenlaw AC, Puri T, Mieda M, Herzog ED. Astrocytes regulate daily rhythms in the suprachiasmatic nucleus and behavior. Curr Biol 2017, 27: 1055–1061.PubMedPubMedCentralCrossRefGoogle Scholar
  13. 13.
    Niu B, Zhang T, Hu HQ, Cao BZ. Transcriptome sequencing reveals astrocytes as a therapeutic target in heat-stroke. Neurosci Bull 2017, 33: 627–640.PubMedPubMedCentralCrossRefGoogle Scholar
  14. 14.
    Jahn HM, Scheller A, Kirchhoff F. Genetic control of astrocyte function in neural circuits. Front Cell Neurosci 2015, 9: 310.PubMedPubMedCentralCrossRefGoogle Scholar
  15. 15.
    Guo ZB, Su YY, Lou HF. GFAP-positive progenitor cell production is concentrated in specific encephalic regions in young adult mice. Neurosci Bull 2018, 34: 769–778.PubMedPubMedCentralCrossRefGoogle Scholar
  16. 16.
    Casper KB, McCarthy KD. GFAP-positive progenitor cells produce neurons and oligodendrocytes throughout the CNS. Mol Cell Neurosci 2006, 31: 676–684.PubMedCrossRefPubMedCentralGoogle Scholar
  17. 17.
    Hirrlinger PG, Scheller A, Braun C, Hirrlinger J, Kirchhoff F. Temporal control of gene recombination in astrocytes by transgenic expression of the tamoxifen-inducible DNA recombinase variant CreERT2. Glia 2006, 54: 11–20.PubMedCrossRefPubMedCentralGoogle Scholar
  18. 18.
    Slezak M, Goritz C, Niemiec A, Frisen J, Chambon P, Metzger D, et al. Transgenic mice for conditional gene manipulation in astroglial cells. Glia 2007, 55: 1565–1576.PubMedCrossRefPubMedCentralGoogle Scholar
  19. 19.
    Casper KB, Jones K, McCarthy KD. Characterization of astrocyte-specific conditional knockouts. Genesis 2007, 45: 292–299.PubMedCrossRefPubMedCentralGoogle Scholar
  20. 20.
    Young KM, Mitsumori T, Pringle N, Grist M, Kessaris N, Richardson WD. An Fgfr3-iCreER(T2) transgenic mouse line for studies of neural stem cells and astrocytes. Glia 2010, 58: 943–953.PubMedPubMedCentralGoogle Scholar
  21. 21.
    Mori T, Tanaka K, Buffo A, Wurst W, Kuhn R, Gotz M. Inducible gene deletion in astroglia and radial glia–a valuable tool for functional and lineage analysis. Glia 2006, 54: 21–34.PubMedCrossRefPubMedCentralGoogle Scholar
  22. 22.
    Winchenbach J, Duking T, Berghoff SA, Stumpf SK, Hulsmann S, Nave KA, et al. Inducible targeting of CNS astrocytes in Aldh1l1-CreERT2 BAC transgenic mice. F1000Research 2016, 5: 2934.PubMedPubMedCentralCrossRefGoogle Scholar
  23. 23.
    Srinivasan R, Lu TY, Chai H, Xu J, Huang BS, Golshani P, et al. New transgenic mouse lines for selectively targeting astrocytes and studying calcium signals in astrocyte processes in situ and in vivo. Neuron 2016, 92: 1181–1195.PubMedPubMedCentralCrossRefGoogle Scholar
  24. 24.
    Lee KM, Lan Q, Kricker A, Purdue MP, Grulich AE, Vajdic CM, et al. One-carbon metabolism gene polymorphisms and risk of non-Hodgkin lymphoma in Australia. Hum Genet 2007, 122: 525–533.PubMedCrossRefPubMedCentralGoogle Scholar
  25. 25.
    Lim U, Wang SS, Hartge P, Cozen W, Kelemen LE, Chanock S, et al. Gene-nutrient interactions among determinants of folate and one-carbon metabolism on the risk of non-Hodgkin lymphoma: NCI-SEER case-control study. Blood 2007, 109: 3050–3059.PubMedPubMedCentralCrossRefGoogle Scholar
  26. 26.
    Neymeyer V, Tephly TR, Miller MW. Folate and 10-formyltetrahydrofolate dehydrogenase (FDH) expression in the central nervous system of the mature rat. Brain Res 1997, 766: 195–204.PubMedCrossRefPubMedCentralGoogle Scholar
  27. 27.
    Cahoy JD, Emery B, Kaushal A, Foo LC, Zamanian JL, Christopherson KS, et al. A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci 2008, 28: 264–278.PubMedPubMedCentralCrossRefGoogle Scholar
  28. 28.
    Zhang Y, Chen K, Sloan SA, Bennett ML, Scholze AR, O’Keeffe S, et al. An RNA-sequencing transcriptome and splicing database of glia, neurons, and vascular cells of the cerebral cortex. J Neurosci 2014, 34: 11929–11947.PubMedPubMedCentralCrossRefGoogle Scholar
  29. 29.
    Ganat YM, Silbereis J, Cave C, Ngu H, Anderson GM, Ohkubo Y, et al. Early postnatal astroglial cells produce multilineage precursors and neural stem cells in vivo. J Neurosci 2006, 26: 8609–8621.PubMedPubMedCentralCrossRefGoogle Scholar
  30. 30.
    Shen B, Zhang J, Wu H, Wang J, Ma K, Li Z, et al. Generation of gene-modified mice via Cas9/RNA-mediated gene targeting. Cell Res 2013, 23: 720–723.PubMedPubMedCentralCrossRefGoogle Scholar
  31. 31.
    Ma Y, Zhang X, Shen B, Lu Y, Chen W, Ma J, et al. Generating rats with conditional alleles using CRISPR/Cas9. Cell Res 2014, 24: 122–125.PubMedCrossRefPubMedCentralGoogle Scholar
  32. 32.
    Cui L, Zhang Z, Sun F, Duan X, Wang M, Di K, et al. Transcervical embryo transfer in mice. J Am Assoc Lab Anim Sci 2014, 53: 228–231.PubMedPubMedCentralGoogle Scholar
  33. 33.
    Wei C, Liu J, Yu Z, Zhang B, Gao G, Jiao R. TALEN or Cas9 - rapid, efficient and specific choices for genome modifications. J Genet Genomics 2013, 40: 281–289.PubMedCrossRefPubMedCentralGoogle Scholar
  34. 34.
    Chen YH, Lan YJ, Zhang SR, Li WP, Luo ZY, Lin S, et al. ErbB4 signaling in the prelimbic cortex regulates fear expression. Transl Psychiatry 2017, 7: e1168.PubMedPubMedCentralCrossRefGoogle Scholar
  35. 35.
    Madisen L, Zwingman TA, Sunkin SM, Oh SW, Zariwala HA, Gu H, et al. A robust and high-throughput Cre reporting and characterization system for the whole mouse brain. Nat Neurosci 2010, 13: 133–140.PubMedCrossRefPubMedCentralGoogle Scholar
  36. 36.
    Liu Y, Wu Y, Lee JC, Xue H, Pevny LH, Kaprielian Z, et al. Oligodendrocyte and astrocyte development in rodents: an in situ and immunohistological analysis during embryonic development. Glia 2002, 40: 25–43.PubMedCrossRefPubMedCentralGoogle Scholar
  37. 37.
    Molofsky AV, Krencik R, Ullian EM, Tsai HH, Deneen B, Richardson WD, et al. Astrocytes and disease: a neurodevelopmental perspective. Genes Dev 2012, 26: 891–907.PubMedPubMedCentralCrossRefGoogle Scholar
  38. 38.
    Yang Y, Vidensky S, Jin L, Jie C, Lorenzini I, Frankl M, et al. Molecular comparison of GLT1 + and ALDH1L1 + astrocytes in vivo in astroglial reporter mice. Glia 2011, 59: 200–207.PubMedPubMedCentralCrossRefGoogle Scholar
  39. 39.
    Matyash V, Kettenmann H. Heterogeneity in astrocyte morphology and physiology. Brain Res Rev 2010, 63: 2–10.PubMedCrossRefPubMedCentralGoogle Scholar
  40. 40.
    Tran CHT, Peringod G, Gordon GR. Astrocytes integrate behavioral state and vascular signals during functional hyperemia. Neuron 2018, 100: 1133–1148 e1133.CrossRefGoogle Scholar
  41. 41.
    Chaboub LS, Deneen B. Developmental origins of astrocyte heterogeneity: the final frontier of CNS development. Dev Neurosci 2012, 34: 379–388.PubMedPubMedCentralCrossRefGoogle Scholar
  42. 42.
    Bayraktar OA, Fuentealba LC, Alvarez-Buylla A, Rowitch DH. Astrocyte development and heterogeneity. Cold Spring Harb Perspect Biol 2014, 7: a020362.PubMedCrossRefPubMedCentralGoogle Scholar
  43. 43.
    Obernier K, Alvarez-Buylla A. Neural stem cells: origin, heterogeneity and regulation in the adult mammalian brain. Development 2019, 146: dev156059.PubMedCrossRefPubMedCentralGoogle Scholar
  44. 44.
    Foo LC, Dougherty JD. Aldh1L1 is expressed by postnatal neural stem cells in vivo. Glia 2013, 61: 1533–1541.PubMedPubMedCentralCrossRefGoogle Scholar
  45. 45.
    Oleinik NV, Krupenko NI, Krupenko SA. Epigenetic silencing of ALDH1L1, a metabolic regulator of cellular proliferation, in cancers. Genes Cancer 2011, 2: 130–139.PubMedPubMedCentralCrossRefGoogle Scholar
  46. 46.
    Boesmans W, Rocha NP, Reis HJ, Holt M, Vanden Berghe P. The astrocyte marker Aldh1L1 does not reliably label enteric glial cells. Neurosci Lett 2014, 566: 102–105.PubMedCrossRefPubMedCentralGoogle Scholar
  47. 47.
    Beil J, Buch T. Generation of bacterial artificial chromosome (BAC) transgenic mice. Methods Mol Biol 2014, 1194: 157–169.PubMedCrossRefPubMedCentralGoogle Scholar
  48. 48.
    Yang XW, Gong S. An overview on the generation of BAC transgenic mice for neuroscience research. Curr Protoc Neurosci 2005, Chapter 5: 5–20.Google Scholar
  49. 49.
    Kesavan G, Chekuru A, Machate A, Brand M. CRISPR/Cas9-mediated zebrafish knock-in as a novel strategy to study midbrain-hindbrain boundary development. Front Neuroanat 2017, 11: 52.PubMedPubMedCentralCrossRefGoogle Scholar
  50. 50.
    Platt RJ, Chen S, Zhou Y, Yim MJ, Swiech L, Kempton HR, et al. CRISPR-Cas9 knockin mice for genome editing and cancer modeling. Cell 2014, 159: 440–455.PubMedPubMedCentralCrossRefGoogle Scholar
  51. 51.
    Reichenbach A, Wolburg H. Neuroglia, 2nd ed. Oxford, UK: Oxford University Press, 2004.Google Scholar
  52. 52.
    Pannasch U, Dossi E, Ezan P, Rouach N. Astroglial Cx30 sustains neuronal population bursts independently of gap-junction mediated biochemical coupling. Glia 2019. 67: 1104–1112.PubMedPubMedCentralCrossRefGoogle Scholar
  53. 53.
    Clasadonte J, Haydon PG. Connexin 30 controls the extension of astrocytic processes into the synaptic cleft through an unconventional non-channel function. Neurosci Bull 2014, 30: 1045–1048.PubMedPubMedCentralCrossRefGoogle Scholar
  54. 54.
    Yoshiki A, Moriwaki K. Mouse phenome research: implications of genetic background. ILAR J 2006, 47: 94–102.PubMedCrossRefPubMedCentralGoogle Scholar
  55. 55.
    Srinivas S, Watanabe T, Lin CS, William CM, Tanabe Y, Jessell TM, et al. Cre reporter strains produced by targeted insertion of EYFP and ECFP into the ROSA26 locus. BMC Dev Biol 2001, 1: 4.PubMedPubMedCentralCrossRefGoogle Scholar
  56. 56.
    Su M, Hu H, Lee Y, d’Azzo A, Messing A, Brenner M. Expression specificity of GFAP transgenes. Neurochem Res 2004, 29: 2075–2093.PubMedCrossRefPubMedCentralGoogle Scholar

Copyright information

© Shanghai Institutes for Biological Sciences, CAS 2019

Authors and Affiliations

  1. 1.State Key Laboratory of Organ Failure Research, Key Laboratory of Mental Health of the Ministry of Education, Guangdong–Hong Kong–Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong Key Laboratory of Psychiatric Disorders, Collaborative Innovation Center for Brain Science, Department of Neurobiology, School of Basic Medical SciencesSouthern Medical UniversityGuangzhouChina
  2. 2.Chronobiotron – UMS 3415University of StrasbourgStrasbourgFrance
  3. 3.Institute of Cellular and Integrative Neurosciences, CNRS UPR 3212University of StrasbourgStrasbourgFrance

Personalised recommendations