Reconstruction of the human nigrostriatal pathway in vitro reveals target-dependent dopamine neuron maturation

Abstract

The human nigrostriatal pathway, comprising dopaminergic neurons in the ventral midbrain (vMB) projecting to the dorsolateral striatum, is essential for motor control and selectively vulnerable in Parkinsons disease (PD). How this circuit assembles during development and how it degenerates under pathological conditions remains poorly understood in a human context and in vitro models capturing its long-range connectivity and spatial organization have been lacking. Here, we introduce the connectoid, a compartmentalized, human stem cell-based model of the nigrostriatal pathway that integrates vMB and striatal organoids within a custom-engineered microfluidic device, confining cell bodies while guiding axonal growth, mimicking the in vivo topography. Functional connectivity was confirmed by retrograde rabies tracing, and optogenetic and pharmacological stimulation, while 6-hydroxydopamine-induced selective degeneration of dopamine neurons, recapitulating a key feature of PD. Additionally, single-cell transcriptomics revealed that interaction with striatal targets enhances dopaminergic neuron maturation and activates transcriptional programs linked to synaptic signaling. Thus, connectoids uniquely allow spatial segregation of regionalized organoids while preserving long-range communication, providing a scalable and physiologically relevant platform for studying human circuit assembly, selective vulnerability, and therapeutic interventions in PD.