Figure 2.

Cortical layer and cell type-specific expression of Cre through lentivector encoding GFP and Cre driven by the CamKII promoter. (A) pCMVΔR8.92 encoding virion components, pRSV-Rev encoding HIV-1 Rev protein, pMD.G, encoding the VSV-G glycoprotein, and the transfer vector plasmid, pFC1.3Cre (αCaMKII GFP IRES Cre or the control αCaMKII GFP) are used for lentivector production by transient transfection of 293T cells. (B) While the mouse is under anesthesia, a small craniotomy is made in the skull to allow a glass micropipette containing the lentivector solution to be introduced directly into the neocortex at a specific location using micromani pulators. A small volume ∼50 nl is then slowly injected into the neocortex. The micropipette is slowly retracted, the skin sutured, and the mouse quickly recovers from the surgical procedure. (C) Three weeks after lentivector injection, the brain was fixed and coronal sections were prepared for confocal imaging. GFP-positive cells are found in a small volume of neocortex (diameter ∼200 μm) at the location of the injection site (left). The vast majority of GFP expressing cells are pyramidal neurons (right). Thus, lentivector injection can allow GFP expression that is column, layer, and cell type specific.