Adult male and female mice (>P60) were used for all surgeries. Bilateral viral injections were performed using previously described procedures (54) at the following stereotaxic coordinates: 1.94 mm from bregma, 0.34 mm lateral from midline, and 0.70 mm vertical from cortical surface for dorsomedial prefrontal cortex (dmPFC) and −3.08 mm from bregma, 1.25 mm lateral from midline, and 4.0 mm vertical from cortical surface for SNc. For glutamatergic corticostriatal axon stimulation experiments, mice were bilaterally injected with 0.5 μl of CAG-ChR2-EYFP (enhanced YFP) virus into dmPFC. For nigrostriatal dopaminergic axon stimulation experiments, DAT-Cre mice were injected with 0.5 μl of DIO-ChR2-EYFP virus bilaterally. For all optogenetic experiments, we waited at least 3 weeks from viral injection to experimental stimulation to allow for sufficient ChR2 gene expression.

To confirm that DA neurons were transfected with ChR2 in animals used for optogenetic DA stimulation experiments, we perfused DAT-Cre mice that had been injected into the SNc with Cre-dependent ChR2-EYFP virus with 4% paraformaldehyde in PBS and post-fixed brains overnight. Coronal sections that included the injection site (SNc) and imaging site (dorsal striatum) were cut at 50 μM and immunolabeled using antibody against tyrosine hydroxylase [TH (1:1000); rabbit anti-TH, Millipore], the rate-limiting enzyme for catecholamine synthesis. Goat anti-rabbit DyLight 594 secondary antibody (1:1000; Invitrogen) was used to visualize TH. Image acquisition was performed on a Zeiss Axio ScanZ.1 using a 5× objective.

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