Tracing Neuron-astrocyte Connections With Retrograde Monosynaptic Rabies Virus

In response to various stimulations,astrocytes have been reported to release signaling molecules to modulate neuronal activities.These results,together with the finding that most CNS synapses are closely associated with astrocyte processes suggest that astrocytes constitute an integrative part of synapses and crosstalk with neurons at the synaptic level.However,there is still lack of an effective tool to directly trace the connections between astrocytes and neurons.Retrograde monosynaptic rabies virus(RV)has been widely used for tracing neurons with monosynaptic connections.In the present study we found surprisingly that RV can be also used to trace astrocyte-neuron connections.We found that retrograde monosynaptic RV,G-deleted rabies(RVdG),spread from starter neurons into astrocytes,but not into NG2 glial cells or microglia.The RVdG transfected into astrocytes can also spread into neurons.The process of bidirection spread between neurons and astrocytes was dependent on RV G.The trans-cell RVdG spreading depends on synapse formation and maturation,suggesting the critical role of synapses in mediating virus spreading.Calcium waves evoked in starter astrocytes or infected astrocytes were found to propagate to connected neurons revealed by retrograde monosynaptic RV.Our results indicate that retrograde monosynaptic RV tracing approach can be also applied to identify neuron-astrocyte network with direct functional interactions.In addition,our studies also suggest that astrocytes may play a certain role in the process of RV infection and its spread in CNS,and provide the basis for future study about the detailed role of astrocytes in rabies.