Virus-Mediated Expression of DREADDs for In Vivo Metabolic Studies

  • Mario Rossi
  • Zhenzhong Cui
  • Ken-ichiro Nakajima
  • Jianxin Hu
  • Lu Zhu
  • Jürgen Wess
Part of the Methods in Molecular Biology book series (MIMB, volume 1335)

Abstract

During the past few years, CNO-sensitive designer G protein-coupled receptors (GPCRs) known as DREADDs (designer receptors exclusively activated by designer drugs) have emerged as powerful new tools for the study of GPCR physiology. In this chapter, we present protocols employing adeno-associated viruses (AAVs) to express a Gq-coupled DREADD (Dq) in two metabolically important cell types, AgRP neurons of the hypothalamus and hepatocytes of the liver. We also provide examples dealing with the metabolic analysis of the Dq mutant mice after administration of CNO in vivo. The approaches described in this chapter can be applied to other members of the DREADD family and, of course, different cell types. It is likely that the use of DREADD technology will identify physiologically important signaling pathways that can be targeted for therapeutic purposes.

Key words

G-protein-coupled receptors Designer GPCRs G proteins Signal transduction Glucose homeostasis Food intake 

References

  1. 1.
    Regard JB, Sato IT, Coughlin SR (2008) Anatomical profiling of G protein-coupled receptor expression. Cell 135:561–571PubMedCentralPubMedCrossRefGoogle Scholar
  2. 2.
    Armbruster BN, Li X, Pausch MH, Herlitze S, Roth BL (2007) Evolving the lock to fit the key to create a family of G protein-coupled receptors potently activated by an inert ligand. Proc Natl Acad Sci U S A 104:5163–5168PubMedCentralPubMedCrossRefGoogle Scholar
  3. 3.
    Wess J, Nakajima K, Jain S (2013) Novel designer receptors to probe GPCR signaling and physiology. Trends Pharmacol Sci 34:385–392PubMedCentralPubMedCrossRefGoogle Scholar
  4. 4.
    Urban DJ, Roth BL (2014) DREADDs (designer receptors exclusively activated by designer drugs): chemogenetic tools with therapeutic utility. Annu Rev Pharmacol Toxicol. doi:10.1146/annurev-pharmtox-010814-124803 PubMedGoogle Scholar
  5. 5.
    Guettier JM, Gautam D, Scarselli M, Ruiz de Azua I, Li JH, Rosemond E, Ma X, Gonzalez FJ, Armbruster BN, Lu H, Roth BL, Wess J (2009) A chemical-genetic approach to study G protein regulation of beta cell function in vivo. Proc Natl Acad Sci U S A 106:19197–19202PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Zincarelli C, Soltys S, Rengo G, Rabinowitz JE (2008) Analysis of AAV serotypes 1-9 mediated gene expression and tropism in mice after systemic injection. Mol Ther 16:1073–1080PubMedCrossRefGoogle Scholar
  7. 7.
    Aponte Y, Atasoy D, Sternson SM (2011) AGRP neurons are sufficient to orchestrate feeding behavior rapidly and without training. Nat Neurosci 14:351–355PubMedCentralPubMedCrossRefGoogle Scholar
  8. 8.
    Krashes MJ, Koda S, Ye C, Rogan SC, Adams AC, Cusher DS, Maratos-Flier E, Roth BL, Lowell BB (2011) Rapid, reversible activation of AgRP neurons drives feeding behavior in mice. J Clin Invest 121:1424–1428PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Cansell C, Denis RG, Joly-Amado A, Castel J, Luquet S (2012) Arcuate AgRP neurons and the regulation of energy balance. Front Endocrinol (Lausanne) 3:169, eCollection 2012Google Scholar
  10. 10.
    Krashes MJ, Shah BP, Koda S, Lowell BB (2013) Rapid versus delayed stimulation of feeding by the endogenously released AgRP neuron mediators GABA, NPY, and AgRP. Cell Metab 18:588–595PubMedCrossRefGoogle Scholar
  11. 11.
    Atasoy D, Betley JN, Su HH, Sternson SM (2012) Deconstruction of a neural circuit for hunger. Nature 488:172–177PubMedCentralPubMedCrossRefGoogle Scholar
  12. 12.
    Krashes MJ, Shah BP, Madara JC, Olson DP, Strochlic DE, Garfield AS, Vong L, Pei H, Watabe-Uchida M, Uchida N et al (2014) An excitatory paraventricular nucleus to AgRP neuron circuit that drives hunger. Nature 507:238–242PubMedCentralPubMedCrossRefGoogle Scholar
  13. 13.
    Cardin JA, Carlen M, Meletis K, Knoblich U, Zhang F, Deisseroth K, Tsai LH, Moore CI (2009) Driving fast-spiking cells induces gamma rhythm and controls sensory responses. Nature 459:663–667PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Atasoy D, Aponte Y, Su HH, Sternson SM (2008) A FLEX switch targets Channelrhodopsin-2 to multiple cell types for imaging and long-range circuit mapping. J Neurosci 28:7025–7030PubMedCentralPubMedCrossRefGoogle Scholar
  15. 15.
    Grieger JC, Choi VW, Samulski RJ (2006) Production and characterization of adeno-associated viral vectors. Nat Protocols 1:1412–1428PubMedCrossRefGoogle Scholar
  16. 16.
    Tong Q, Ye CP, Jones JE, Elmquist JK, Lowell BB (2008) Synaptic release of GABA by AgRP neurons is required for normal regulation of energy balance. Nat Neurosci 11:998–1000PubMedCentralPubMedCrossRefGoogle Scholar
  17. 17.
    Lin HV, Accili D (2011) Hormonal regulation of hepatic glucose production in health and disease. Cell Metab 14:9–19PubMedCentralPubMedCrossRefGoogle Scholar
  18. 18.
    Nakajima K, Wess J (2012) Design and functional characterization of a novel, arrestin-biased designer G protein-coupled receptor. Mol Pharmacol 82:575–582PubMedCentralPubMedCrossRefGoogle Scholar
  19. 19.
    Li JH, Jain S, McMillin SM, Cui Y, Gautam D, Sakamoto W, Lu H, Jou W, McGuinness OP, Gavrilova O, Wess J (2013) A novel experimental strategy to assess the metabolic effects of selective activation of a Gq-coupled receptor in hepatocytes in vivo. Endocrinology 154:3539–3551PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Mario Rossi
    • 1
  • Zhenzhong Cui
    • 1
  • Ken-ichiro Nakajima
    • 1
  • Jianxin Hu
    • 1
  • Lu Zhu
    • 1
  • Jürgen Wess
    • 1
  1. 1.Molecular Signaling Section, Laboratory of Bioorganic ChemistryNIH-NIDDKBethesdaUSA

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