The Cre/loxP system has been used extensively for conditional mutagenesis in mice. Reporters of Cre activity are important for defining the spatial and temporal extent of Cre-mediated recombination. Here we describe mT/mG, a double-fluorescent Cre reporter mouse that expresses membrane-targeted tandem dimer Tomato (mT) prior to Cre-mediated excision and membrane-targeted green fluorescent protein (mG) after excision. We show that reporter expression is nearly ubiquitous, allowing visualization of fluorescent markers in live and fixed samples of all tissues examined. We further demonstrate that mG labeling is Cre-dependent, complementary to mT at single cell resolution, and distinguishable by fluorescence-activated cell sorting. Both membrane-targeted markers outline cell morphology, highlight membrane structures, and permit visualization of fine cellular processes. In addition to serving as a global Cre reporter, the mT/mG mouse may also be used as a tool for lineage tracing, transplantation studies, and analysis of cell morphology in vivo.

SUMMARY Dopamine (DA) neurons in the midbrain ventral tegmental area (VTA) integrate complex inputs to encode multiple signals that influence motivated behaviors via diverse projections. Here we combine axon-initiated viral transduction with rabies-mediated transsynaptic tracing and Cre-based cell type-specific targeting to systematically map input–output relationships of VTA-DA neurons. We found that VTA-DA (and VTA-GABA) neurons receive excitatory, inhibitory, and modulatory input from diverse sources. VTA-DA neurons projecting to different forebrain regions exhibit specific biases in their input selection. VTA-DA neurons projecting to lateral and medial nucleus accumbens innervate largely non-overlapping striatal targets, with the latter also sending extra-striatal axon collaterals. Using electrophysiology and behavior, we validated new circuits identified in our tracing studies, including a previously unappreciated top-down reinforcing circuit from anterior cortex to lateral nucleus accumbens via VTA-DA neurons. This study highlights the utility of our viral-genetic tracing strategies to elucidate the complex neural substrates that underlie motivated behaviors.

Top-down modulation of sensory processing allows the animal to select inputs most relevant to current tasks. We found that the cingulate (Cg) region of mouse frontal cortex powerfully influences sensory processing in primary visual cortex (V1) through long-range projections that activate local GABAergic circuits. Optogenetic activation of Cg neurons enhanced V1 neuron responses and improved visual discrimination. Focal activation of Cg axons in V1 caused a response increase at the activation site but decrease at nearby locations (center-surround modulation). While somatostatin-positive GABAergic interneurons contributed preferentially to surround suppression, vasoactive intestinal peptide-positive interneurons were crucial for center facilitation. Long-range cortico-cortical projections thus act through local microcircuits to exert spatially specific top-down modulation of sensory processing.