This past year marks the initial establishment of the Optogenetics and Transgenic Technology Core. We have hired a group of people who are now trained in the methods to package, purify and characterize (in vitro and in vivo) both adeno-associated viral (AAV) vectors and lentiviral vectors (LV). We routinely generate vectors both for the NIDA IRP as well as other intramural and extramural collaborations. The vectors are designed primarily for studies focused on molecular mechanisms of addiction and neurodegeneration. For example, we have produced over 20 different vectors for delivery of the opsins for optogenetic manipulation of neuronal circuits. We have also been developing several novel vectors to express synaptically targeted fluorescent proteins, voltage sensitive fluorescent proteins, calcium-sensitive fluorescent proteins, infrared reporter protein and photosensitizing proteins for lesioning neurons with light. We have our first generation of transgenic rats to be characterized. We have a conditional-mCherry reporter rat that will only express the mCherry fluorescent protein where Cre recombinase is present. This rat can be used to test Cre expressing viral vectors or crossed with our other cell specific, Cre expressing rats to examine the specificity of Cre expression. We also have pups for our animals expressing the Tet repressor protein. The Tet repressor protein can bind to specific DNA elements (Tet operator) to prevent gene expression. By giving animals tetracycline or an analog we can cause the repressor to come off the DNA and allow gene expression. This will be used in combination with viruses or other rats that containing the Tet operator to allow us to activate gene expression. Constructs have also been injected into fertilized eggs for a microglia specific expression of green fluorescent protein, the dopamine transporter promoter expressing Cre recombinase and the Fos promoter, a marker of neuronal activity, expressing Cre recombinase. These animals will be characterized over the next several months. All of our characterized and published vectors are available for intramural and extramural collaborators.
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