ObjectiveTo preliminary study on the feasibility of constructing three-dimensional (3D) hippocampal neural network in vitro by using microfluidic technology.MethodsA network patterned microfluidic chip was designed and fabricated by standard wet etching process. The primary hippocampal neurons of neonatal Sprague Dawley rats were isolated and cultured, and then inoculated on microfluidic chip for culture. Immunofluorescence staining was used to observe the growth of hippocampal neurons at 3, 5, and 7 days of culture and electrophysiological detection of hippocampal neuron network at 7 days of culture.ResultsThe results showed that the number of hippocampal neurons increased gradually with the prolongation of culture time, and the neurite of neurons increased accordingly, and distributed uniformly and regularly in microfluidic chip channels, suggesting that the 3D hippocampal neuron network was successfully constructed in vitro. Single and multi-channel spontaneous firing signals of hippocampal neuronal networks could be detected at 7 days of culture, suggesting that neuronal networks had preliminary biological functions.ConclusionPatterned microfluidic chips can make hippocampal neurons grow along limited paths and form 3D neuron networks with corresponding biological functions such as signal transduction, which lays a foundation for further exploring the function of neuron networks in vitro.