A Cross-Species Enhancer AAV Toolkit for Cell Type-Specific Targeting Across the Basal Ganglia

The mammalian basal ganglia (BG) orchestrate diverse motor, cognitive, and affective functions, yet efforts to dissect their cellular circuitry have been limited by a lack of scalable, cell type-specific tools, especially in nonhuman primates. Moreover, key basal ganglia structures outside the striatum, including those critically involved in neuropsychiatric and movement disorders, remain difficult to access due to a lack of high-specificity viral reagents. Here, we present a comprehensive enhancer-AAV library enabling targeted labeling and manipulation of the major neuronal populations across the BG, including striatal projection neuron subtypes, pallidal and subthalamic populations, and dopaminergic and GABAergic midbrain neurons. Using an evolutionarily informed enhancer discovery pipeline, we identified and validated functional enhancers derived from both mouse and macaque, including tools for both canonical and rare subtypes. Systematic testing reveals robust cross-species activity and highlights both conserved and divergent features of enhancer-driven specificity. Bioinformatic modeling of the full enhancer library identifies key sequence features predictive of tool success, including motif content, chromatin accessibility, and evolutionary conservation. We conclude by identifying class-specific enhancer architectures, with glial, projection, and interneuron BG lineages each exhibiting unique regulatory codes. By uniting cross-species enhancer design with deep mechanistic insight, this work establishes the first comprehensive, cross-species viral toolkit for the basal ganglia, unlocking previously inaccessible cell types and setting a new standard for regulatory element-driven circuit targeting.

Data citation

No data citation listed

Funding agency

NIMH

Grant numbers

1RF1MH121274

Access

open

License

CC BY 4.0