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Welcome to the Designer Receptors Exclusively Activated by Designer Drugs DREADD wiki

DREADDs are engineered G-protein coupled receptors which are activated by otherwise inert drug-like small molecules. In the example shown below, an evolved M3-muscarinic receptor is activated by the small molecule clozapine-N-oxide (CNO; see Armbruster et al, PNAS 2007 for details) http://www.ncbi.nlm.nih.gov/pubmed/17360345

http://pharmrev.aspetjournals.org/content/63/2/291/F3.small.gif

DREADDs are currently used by many labs to remotely and non-invasively control neuronal signaling

http://davecharest.com/wp-content/uploads/2009/12/neurons1-240x180.jpg

Helpful reviews on the technology:

Rogan and Roth, Pharmacol Rev 2011 Jun;63(2):291-315. Epub 2011 Mar 17. http://www.ncbi.nlm.nih.gov/pubmed/21415127

Dong et al, Mol Biosystems 2010 Aug;6(8):1376-80. Epub 2010 Jun 7. http://www.ncbi.nlm.nih.gov/pubmed/20532295


Nomenclature

Gq-DREADD = hM3Dq = D-q ; clozapine-N-oxide activates Gq-mediated signaling. In neurons activation of hM3Dq induces burst firing

Gi-DREADD=hM4Di= D-i; clozapine-N-oxide induces Gi activation. In neurons, activation of hM4Di induces neuronal silencing

Gs-DREADD = Gs-D = D-s; clozapine-N-oxide induces Gs activation. In neurons this leads to an increase in cAMP.

Detailed Protocols for Creating DREADDs

Dong et al, Nature Protocols 2010;5(3):561-73. Epub 2010 Feb 25. http://www.ncbi.nlm.nih.gov/pubmed/20203671


Listed below are various DREADD constructs currently available with links to VectorNTI files:

available in pcDNA3.0:

HA-tagged hM3D in pcDNA3: this is coupled to Gq and induces neuronal burst firinghttp://www.ncbi.nlm.nih.gov/pubmed/19607790

HA-tagged hM4D in pcDNA3: this is coupled to Gi and induces neuronal silencing http://www.ncbi.nlm.nih.gov/pubmed/17360345

HA-tagged Gs-DREADD in pcDNA3: this is coupled to Gs and elevates cAMP production http://www.ncbi.nlm.nih.gov/pubmed/19858481



Available as FLEX-AAV Constructs using Synapsin-1 promotor (also known as DIO Vectors) (See Krashes et al, JCI 2011) http://www.ncbi.nlm.nih.gov/pubmed/21364278)
mCherry Fusion Proteins

hM3Dq-mCherry fusion protein construct http://www.ncbi.nlm.nih.gov/pubmed/21364278: Causes burst firing of neurons

hM4Di-mCherry fusion protein construct http://www.ncbi.nlm.nih.gov/pubmed/21364278: Silences neuronal firing


AAV-FLEX Constructs with Syanpsin-1 promotor; also contain IRES and either mCherry or mCitrine reporters: Validated in HEK-293 cells and neurons but not yet published.

HIGH TITER VIRUS NOW AVAILABLE FROM THE UNC VECTOR CORE FOR mCITRENE VERSIONS
HA-hM3Dq-IRES-mCherry

HA-hM3Dq-IRES-mCitrine


HA-hM4Di-IRES-mCherry

HA-hM4Di-IRES-mCitrine



How to obtain high-titer virus stocks: Each aliquot is ~$220 and no MTA needed. How to obtain:

  1. Please go to http://genetherapy.unc.edu/services.htm
  1. Fill out the order form for serotype 5, 2 or 8 virus. We find that AAV serotype 8 yields highest levels of neuronal expression and greatest spread of infection and is ideal for studies using DIO vectors. Serotypes 2 and 5 are best where localized expression is key.

Available in Lentiviral vectors

HA-hM3Dq-IRES-mCherry http://www.ncbi.nlm.nih.gov/pubmed/19607790

HA-hM4Dq-IRES-mCherry

DREADDs targeted to neuronal subdomains (available for collaborative studies only):

Somato-dendritically-targeted DREADDs (hM4Di, hM3Dq)

Axonally-targeted DREADD (hM4Di, hM3Dq)

Glial-specific expression: AAV-GFAP minimal promotor available via UNC Vector core

Glial-specific Gs and Gq DREADDs (GsD and hM3Dq)


Mice currently available:

tet-HA-tagged hM3Dq mice: http://www.ncbi.nlm.nih.gov/pubmed/19607790
tet-HA-tagged hM4Dq mice: http://www.ncbi.nlm.nih.gov/pubmed/24525710

Adora2A-GsD mice:http://www.ncbi.nlm.nih.gov/pubmed/23303063

http://www.antibodyreview.com/article_images/19893765/Front%20Mol%20Neurosci/2-_/p16-2773177/fnmol-02-016-g002.jpg



The Jackson lab offers tet- HA-tagged hM4Di mice http://jaxmice.jax.org/strain/024114.html

The Jackson Laboratory offers tet-HA-tagged hM3Dq mice: http://jaxmice.jax.org/strain/014093.html

Adora2A-rM3Ds-mCherry mice: http://jaxmice.jax.org/strain/017863.html

Detailed protocol for HA-staining now available (see attachments below)

Detailed protocol for dissolving CNO now available (see attachments below)

Vector maps available for common vectors (see Attachments)

Last modified 3 months ago Last modified on 06/24/14 12:04:53

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