Calbindin stratifies midbrain dopaminergic neurons governing distinct aspects of locomotion

Despite advances in delineating the molecular diversity and projection patterns of midbrain dopamine (DA) neurons, subtype-specific contributions to motor learning and movement execution remain poorly defined. Here, we applied intersectional ablation and inhibitory chemogenetics to dissect the roles of calbindin-expressing (CALB1⁺) and nonexpressing (CALB1^-) DA neurons in locomotion. Using newly engineered intersectional autocleavable Caspase3 constructs, we ablated CALB1⁺ or CALB1^- DA neurons in the mouse midbrain. CALB1^- DA neuron ablation caused severe weight loss, whereas CALB1⁺ DA neuron ablation produced no overt health impairments. Nonetheless, loss of either subtype led to a bradykinetic-like phenotype on the initiation and vigor of voluntary movements. Only ablation of CALB1^- DA neurons impaired performance on the accelerated rotarod. To test if these phenotypes are the result of DA subtype activity, we silenced either population using the inhibitory DREADD hM4Di. Consistent with ablation, silencing CALB1^- DA neurons impacted the initial performance on the rotarod, whereas inhibition of CALB1⁺ DA neurons did not impact performance on the first day, but prevented across-day improvement. Silencing both populations impaired the initiation and vigor of voluntary movements. We next investigated whether this locomotor phenotype stemmed from reduced DA release in the dorsolateral striatum (DLS). While CALB1^- silencing abrogated DA transients in the DLS, CALB1⁺ silencing unexpectedly resulted in increased transients in DLS. Thus, our results demonstrate that DA transients in the DLS are not invariably coupled with movement execution. Altogether, these findings uncover both distinct and shared roles of molecularly defined DA subtypes in shaping different aspects of locomotion.