Viral vector de novo

face:!5.7 mm,!5.5 mm,!5.3 mm). A micromanipulator (Narishige) was used to control injection speed at 30 nl/min, and the pipette was withdrawn 15 min af-ter the final injection. For injection, rAAV-FLEX-rev-ChR2mCherry titer was 3.4 # 1013 genomic copies (GC)/ml; rAAV-ChR2EGFP titer was 1.8 # 1012 GC/ml. Imaging Mice were deeply anesthetized and then perfused with cold saline followed by 4% paraformaldehyde (PFA) in PBS and fixed overnight in PFA. Brain sections (60!m) were cut, mounted on glass slides, and coverslipped for imaging. Neuron images were collected by confocal microscopy (Zeiss). Electrophysiology and photostimulation Experimental techniques were similar to those reported previously (Petreanu et al., 2007), and only the differences are de-scribed here. Mice [postnatal day 21 (P21)–P25] were infected as above, and 5–8 d later they were deeply anesthetized with isoflurane and decapitated. Coronal brain slices were prepared in chilled cut-ting solution containing the followin