Schematic representations of the microfluidic chambers and experimental methods.

<p>(A) Top view and cross-section of a compartmentalized microfluidic chamber. Two identical square chambers (6mm) are connected by 100 microchannels (10μm width, 450μm length and 3μm height) arranged in clusters of 10 with gaps of 20μm between the channels and 75μm between the clusters. A cutout in the silicone layer with a height of 100μm was added to allow seeding neural precursor spheres close to the channels. Differentials of volume and NGF were established between the chambers to attract axons from the soma to the axonal compartment. When the experiment was initiated by seeding VZV-infected cells in the axon compartment, the volume differential was reversed to block diffusion of virions from the axonal to cell-body compartment. (B&C) Schematic representation of experimental methods of cell-associated neuronal infection by VZV. Blue triangles represent neuronal cell-bodies and green or red ovals represent GFP or RFP-expressing non-neuronal cells. In the upper row of B, axons were infected by plating input cells into the axonal compartment of microfluidic chambers. In the lower portion of B, VZV-infected input cells were seeded onto cultures of differentiated hESC-derived neurons. (C) Diagram depicting experimental method used to demonstrate fusion of neurons with non-neuronal cells in syncytia. VZV66RFP infected ARPE cells and GFP-expressing MeWo cells were mixed together and plated on differentiated neurons in glass-bottom culture dishes.