Fig. 3.

Representative confocal images of DRG outgrowth on astrocyte monolayers grown on stamped homogeneous LN fields (left panels) or patterned LN lanes (right panels). (A) Representative DRG outgrowth (red) on live astrocyte monolayers grown on homogeneous LN substrates (GFAP-green) showed no overall preferred directional outgrowth. (B) Similarly, no directionally biased DRG outgrowth was observed on fixed astrocytes cultures grown on homogeneous LN substrates. (C) In sharp contrast, DRG outgrowth on aligned astrocytes monolayers grown on patterned LN substrates extended in the direction of the underlying astrocytes whether alive (C) or fixed (D). Representative, 3D reconstructed confocal images of DRG outgrowth on unaligned astrocytes (E, grown on homogeneous stamped LN) or on aligned astrocyte monolayers (F) demonstrated that DRG outgrowth rarely penetrated into underlying astrocytes and did not reach the surfaces of the stamped coverslips showing that all DRG outgrowth was determined by the interaction with the astrocytes cell layers. Scale bar= 100 µm. Astrocytes were visualized with IHC against GFAP (green) while DRG outgrowth was visualized with antisera directed against neurofilament (red).