Supplementary Figure 5: ArchT expression in L5 neurons is similar using virus or transgenic mice.

a, Whole-cell recordings were obtained from layer 5 neurons in acute brain slices. ArchT was expressed by injecting Rbp4-cre mice with an ArchT virus (see Methods) or by breeding Rbp4-cre and Ai40D mice. Micrograph shows a 2-photon maximal intensity z-stack of a virally infected layer 5 neuron filled with Alexa594, along with neighboring infected cells (green). Scale bar = 100 µm. A fluorescence ratio of recorded to neighboring cells in the same field of view was 2.9 ± 0.5 (mean and sem), suggesting that our visual targeting approach may have biased our recordings to neurons exhibiting a relatively high expression level. b, Top: Example traces showing that 590 nm light pulses (300 ms) via full-field illumination hyperpolarizes the membrane potential of somatosensory cortex layer 5 neurons expressing ArchT via transgenic or viral methods (black and red traces, respectively). c, Summary data showing the effect of light intensity on membrane potential hyperpolarization for n=9 and n=10 neurons in transgenic and virus groups, respectively. Solid lines represent individual experiments and circles are mean ± SEM. A two-way repeated measures ANOVA reveals a main effect of light intensity (F(5,85)=96.17, p = 0.000073), but the membrane potential hyperpolarization was not significantly different between transgenic and virus groups at any light intensity (p=0.239 for each light intensity after correcting for multiple comparisons, Bonferonni post-hoc test).