Cdc42 Promotes Axonogenesis of Primary Hippocampal Neurons by Inhibiting Glycogen Synthase Kinase-3 beta

Background: Progressive axon degeneration is a common pathological feature of neurodegenerative diseases. Cdc42 is a member of the Rho GTPase family that participates in axonogenesis. GSK-3 beta is a serine/threonine kinase highly implicated in neuronal development and neurodegeneration. This study aimed to examine whether cdc42 promotes axonogenesis by regulating GSK-3 beta activity. Methods: Hippocampal neurons were isolated from neonatal Sprague-Dawley rats and transfected with designated plasmid vectors to alter the activities of cdc42 and GSK-3 beta. LiCl treatment was used to inhibit the GSK-3 beta activity in primary neurons. GSK-3 beta activity was determined by an enzyme activity assay kit. Immunofluorescence staining was used to detect axons stained with anti-Tau-1 antibody and dendrites stained with anti-MAP2 antibody. Results: Transfection with an active cdc42 mutant (cdc42F28L) decreased the activity of GSK-3 beta and induced axonogenesis in primary rat hippocampal neurons, while transfection with a negative cdc42 mutant (cdc42N17) resulted an opposite effect. Moreover, transfection with plasmid vectors carrying wild-type GSK-3 beta or a constitutively active GSK3 beta mutant (GSK-3 beta S9A) increased the activity of GSK-3 beta and attenuated axonogenesis of primary hippocampal neurons with excessive cdc42 activity, whereas inhibition of GSK-3 beta by LiCl abolished the inhibitory effect of the negative cdc42 mutant on axonogenesis. Conclusions: This study suggests that cdc42 induces axonogenesis of primary rat hippocampal neurons via inhibiting GSK-3 beta activity. These findings support further investigation into the mechanisms of cdc42/GSK-3 beta-mediated axonogenesis.