Corsi, S. | bioRxiv Science

bioRxiv2025

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

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.

neuroscience↗

bioRxiv2025

Cografting strategies uncover cell type dependent regulation of dopamine neuron specification and functional maturation in a pre clinical model of Parkinson`s Disease

Parkinsons disease (PD), the second most common neurodegenerative disorder, is characterized by the progressive loss of A9 dopaminergic neurons in the substantia nigra, leading to dopamine (DA) depletion in the striatum and subsequent motor symptoms. Transplantation of ventral midbrain-patterned DA (vmDA) progenitors derived from human pluripotent stem cells, aimed at restoring DA neurotransmission in the striatum, is being developed and currently explored in ongoing clinical trials. One factor that may improve the maturation and fate determination of DA neurons in vivo is the intercellular communication within the graft environment, ultimately affecting the therapeutic outcome. In this study, we co-transplanted vmDA progenitors with either glial, ventral forebrain or striatal progenitors into a preclinical xenograft PD model to investigate how these interactions shape the development, maturation, and function of therapeutic DA neurons. Our findings show that co-grafts with ventral forebrain progenitors increase the yield of DA neurons and also promote their functional maturation. Furthermore, we demonstrated that co-grafts with striatal neurons promote functional maturation and the acquisition of DA subtype identity. From these data, we identified EBF3 and PBX3 as candidate transcription factors directing DA neuron maturation and subtype specification, and then functionally validated their role in brain organoids. Taken together, our data highlight that the cellular microenvironment, including specific interactions with neighbouring cells, guides in vivo DA neuron specification and maturation. These findings provide a foundation for developing more refined and effective cell preparations for replacement therapy in PD, and define a conceptual framework that could inform stem cell-based strategies for other neurodegenerative diseases.

neuroscience↗