Aim: To investigate the changes in the spontaneous neuronal excitability induced by astragaloside IV (AGS-IV) in the cultured hippocampal network. Methods: Hippocampal neurons in culture for 9-11 d were used for this study. The spontaneous synaptic activities of these hippocampal neurons were examined by Ca2+ imaging and whole-cell patch-clamp techniques. In total, 40 mg/L AGS-IV dissolved in DMSO and 2 mL/L DMSO were applied to the neurons under a microscope while the experiments were taking place. Results: AGS-IV inhibited the frequencies of synchronized spontaneous Ca2+ oscillations to 59.39%±3.25% (mean±SEM), the spontaneous postsynaptic currents to 43.78%±7.72% (mean±SEM), and the spontaneous excitatory postsynaptic currents to 49.25%±7.06% (mean±SEM) of those of the control periods, respectively, at 16 min after the AGS-IV applications. AGS-IV also decreased the peak values of the voltage-gated K+ and Na+ channel currents at that time point. Conclusion: These results indicate that AGS-IV suppresses the spontaneous neuronal excitabilities effectively. Such a modulation of neuronal activity could represent new evidence for AGS-IV as a neuroprotector.