Neurexins contribute to inhibitory connectivity and dopamine neuron resilience in culture

: Midbrain dopamine (DA) neurons are essential regulators of basal ganglia functions. Their axonal structure is intricate, with numerous non-synaptic release sites and fewer synaptic terminals that notably release glutamate or GABA. Despite their significance, the molecular mechanisms governing DA neuron connectivity and neurochemical identity remain poorly understood. We hypothesize that trans-synaptic cell adhesion molecules such as neurexins (Nrxns) regulate the interactions of DA neuron axons with target cells and thereby influence axonal branching and synapse formation by DA neurons. We therefore examined neuronal survival, axonal growth and synapse formation in cultured DA neurons lacking all neurexins (DAT::NrxnsKO). Conditional deletion of all Nrxns in DA neurons revealed that loss of Nrxns does not disrupt the basic development of these neurons or the structure of their axonal terminals, including normal expression of the vesicular monoamine transporter (VMAT2) and the calcium sensor synaptotagmin 1 (Syt1). However, we observed a reduction in the number of TH-positive DA neurons in culture, suggesting altered resilience or increased vulnerability under in vitro conditions. In addition, loss of Nrxns selectively reduced the proportion of DA neuron terminals associated with the inhibitory postsynaptic marker gephyrin, while the association with excitatory synaptic markers was preserved.