AAV with eSYN promoter driven Cre Inducible hChR2(H134R)-EGFP

This AAV expresses DIO-hChR2(H134R)-EGFP driven by a neuron eSYN promoter.

The eSYN is a hybrid promoter consisting of a 0.4Kb CMV enhancer (E) and the 0.45Kb human Synapsin I promoter fragment (hSYN1). eSYN hybrid promoter gives about 2-4 fold higher expression than that of the parental hSYN1 promoter. However, the neuronal specificity of eSYN is slightly decreased, compared to that of the parental hSYN1 promoter.

Channelrhodopsins are light-gated ion channels that exist naturally in microalgae. hChR2 is a humanized version of ChR2 for mammalian expression. It is maximally excited by 470 nm light. The wild-type, as well as a few mutations, provide the fastest excitation of the channelrhodopsins offered, and are widely used in optogenetics techniques in neuroscience. Due to delayed channel closure, hChR2(H134R) is a gain-of-function mutation that produces larger photocurrents than wild-type hChR2, but slows down channel kinetics.

In the DIO scenario, the transgene of interest is inserted in reverse orientation relative to the 5' promoter and is flanked by oppositely oriented loxP and lox2272 sites. In the absence of Cre expression, the transgene will not be produced. In the presence of Cre expression, the transgene will be "FLip-EXchanged" or FLEXed, leading to expression of the transgene. This is due to a permanent Cre-mediated recombination/inversion of the flanked transgene. This arrangement is called DIO (double-floxed inverse ORF), Cre-ON, Flex-rev (reverse), Flex-ON/FlexON, or DIO-AAV/AAV-DIO (double-floxed inverse ORF in AAV).