Modulation of DRG neurons response to semaphorin 3A via substrate stiffness

Abstract

Semaphorin 3A (Sema3a) is a chemotropic protein that acts as a neuronal guidance cue and plays a major role in dorsal root ganglion (DRG) sensory neurons projection during embryo development. The present study evaluated the impact of stiffness in the repulsive response of DRG neurons to Sema3a when cultured over substrates of variable stiffness. Stiffness modified DRG neurons morphology and regulated their response to Sema3a, reducing the collapse of growth cones when they were cultured on softer substrates. Sema3a receptors expression was also regulated by stiffness, neuropilin-1 was overexpressed and plexin A4 mRNA was downregulated in stiffer substrates. Cytoskeleton distribution was also modified by stiffness. In softer substrates, βIII-tubulin and actin co-localized up to the leading edge of the growth cones, and as the substrate became stiffer, βIII-tubulin was confined to the transition and peripheral domains of the growth cone. Moreover, a decrease in the α-actinin adaptor protein was also observed in softer substrates. Our results show that substrate stiffness plays an important role in regulating the collapse response to Sema3a and that the modulation of cytoskeleton distribution and Sema3a receptors expression are related to the differential collapse responses of the growth cones.