Neuron labeling by GFP expression using Cre-activated AAV vectors. Two strategies have been used to render GFP expression conditional on Cre-mediated recombination. (A) A loxP-STOP-loxP cassette is inserted between the promoter and the GFP gene. (B) A GFP gene flanked by a pair of incompatible antiparallel loxP sites is cloned in the opposite orientation to an upstream cytomegalovirus (CMV) promoter; Cre recombination results in an inversion of this flip-excision (FLEX) switch and, thus, expression of the GFP gene. (C) Injection of the AAV-lox-STOP-lox-GFP virus into the neocortex of a Pv-cre mouse specifically labeled a subclass of GABAergic neurons (S. Kuhlman and Z.J. Huang, unpubl.). (D–F) In vivo two-photon imaging of GFP-labeled cortical basket interneurons in Pv-Cre mice. (D) z-Projections of an image stack 85–90 µm below the pia mater. Note the smooth, aspiny dendrites (blue arrow) and the dense cluster of boutons of varying size (yellow arrowheads). (E) Projection of a z-series 60–170 µm below the pia mater from a sparsely labeled area, showing an isolated basket interneuron. Dendrites (blue arrows) could be traced back to the soma. Axonal boutons (yellow arrowheads) appear as a cloudy signal at this magnification. (F) Examples of a well-isolated basket cell axon (yellow arrow) with distinct boutons (yellow arrowheads). Scale bars, 100 µm in C, 20 µm in E, 5 µm in D and F. (D–F, Adapted from Kuhlman and Huang 2008.)