Hippocampal neurons of neonatal rats were cultured in serum-free culture medium for 5 days in vitro, and treated with the Rho-kinase inducer lysophosphatidic acid.Atomic force microscopy revealed that the numbers of level-1, -2 and -3 neurites protruding from rat hippocampal neurons was significantly reduced.After treatment with the Rho kinase inhibitor Y27632, a significant increase in the numbers of these neurites was observed.Our experimental findings indicate that the Rho-kinase pathway is closely associated with the neurites of hippocampal neurons.