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Showing posts from September, 2014

DREADDs in rats redux

An elegant paper in Cell today demonstrating (again) that local infusions of CNO can modulate terminal release in rats.  Again, TH-Cre rats were used and both excitatory and inhibitory DREADDs were used to modulate norepinephrine release and thereby modify choice decisions.

Where can I obtain CAV-Cre for 'retro-DREADD'-esque studies

I see that this facility in France may provide CAV-Cre

http://www.biocampus.cnrs.fr/index.php/en/plateformes?id=78


How much virus should I inject?

Q: "I have some Gi-DREADDs that I got from UNC and am preparing to inject an aliquot of them into the cortex.

Can you tell me if the sample I got from UNC should be diluted or should it be used as is?  If diluted, what do you recommend and how much?"

A1: "We look at the titer and generally inject 10^9/uL--so it may need to be diluted to 10^9/ul"
Explanation: we try to inject virus with a titer generally ~ 10^9/ul
A2:  "It depends on what kind of experiments you are going to do. Some experiments you need to dilute the virus just like Bryan said. Some are not.
You can aliquot the virus into 5ul/tube after thawing the virus on ice, and store it in -80. Try your injection with different dilutions of virus. Check the expression and then decide what dilution is good for you. "

DREADDs using cell-type specific promotors

Q: "Doyou know of anyone who has put DREADD into locus coeruleus?"

A: yes a very nice paper recently using PRSx8 and lentivirus.  Per the authors:

 "PRSx8 is based on an upstream regulatory site in the human DBH promoter and drives high levels of expression in adrenergic neurons"


Use of AAV for therapy of human CNS diseases: relevance for use of DREADDs in humans

The recent posts on the use of DREADDs in primates and novel lentiviral approaches in primate brainhas raised the issue of the suitability of virally-mediated transduction of human neurons for delivery of various cargos (including DREADDs).

Clinicaltrials.gov shows many ongoing/completed trials of AAV-based gene delivery for a variety of CNS diseases in humans indicating that this is certainly feasible.

Targeting primate neurons with lentiviral vectors

A very nice technical paper has appearedwhich describes many of the parameters which can be optimized to achieve cell-type specific expression of many cargos (including DREADDs) in primates using lentiviral vectors.