Sulfation of Chondroitin Sulfate Regulates Neuronal Morphology via Src-Family Signaling with Likely Contribution from Fyn

Chondroitin sulfate (CS) chains are major components of the extra- and pericellular matrix in the central nervous system (CNS), and their sulfation patterns influence CNS development and function. Highly sulfated CS preparations, including CS-D- and CS-E-enriched forms, have been shown to facilitate neurite outgrowth in cultured mouse hippocampal neurons. Notably, neurons cultured on CS-D- or CS-E-enriched substrates exhibited the following distinct morphological characteristics: CS-D promoted the extension of multiple short neurites, whereas CS-E induced the formation of a single elongated neurite with a polarization-like morphology. These features are consistent with early stages of neuronal polarization. However, the specific roles of these highly sulfated CS forms in polarization-like morphology remain unclear. In this study, we demonstrate that polarization-like morphological transitions in hippocampal neurons can be modulated on mixed CS-D/CS-E substrates by varying their ratios. Compared with CS-D-enriched substrates, CS-E-enriched substrates more effectively promoted polarization-like neuronal morphology, accompanied by enhanced activation of Src-family kinases. Furthermore, forced activation of Fyn kinase induced morphological changes resembling polarization-like features in a neuroblastoma cell line, even in the absence of CS-D/CS-E mixed substrates. In conclusion, highly sulfated CS subtypes may function as extracellular cues that regulate neuronal morphology via Src-family signaling, with likely involvement of Fyn.