The aim of this study was to assess hemodynamic changes by near-infrared spectroscopy (NIRS) during acute focal cerebral ischemia and reperfusion. The study also has evaluated the therapeutic effects of estrogen against vascular dysfunction. Focal cerebral ischemia was induced in nine bilaterally ovariectomized rats, using an endovascular occlusion technique of the middle cerebral artery (MCA). Four out of nine rats had estrogen pretreatment before MCA occlusion (MCAO). The other five rats had MCAO with no pretreatment. The occlusion time was 60 min, followed by 40-60 min of reperfusion. Real-time monitoring of changes in hemoglobin concentrations was performed by a steady-state, two-channel, NIRS system through the period of occlusion and reperfusion. Both changes in total and oxygenated hemoglobin concentrations (Δ[HbT] and Δ[HbO2]) display apparent periodic fluctuations during occlusion for the rats without estrogen pretreatment, while no rhythmic fluctuation was observed in the rats with the pretreatment. This rhythmic fluctuation is a microvascular dysfunction. Fourier power spectral analysis was performed on the Δ[HbO2] profiles in both rat groups. The results show that the cumulative frequency power of Δ[HbO 2] in the range of 0.0025-0.01 Hz for the rats without pretreatment is significantly higher than that with pretreatment. The study implies that the dysfunctional fluctuations disappear in the rats with estrogen pretreatment, demonstrating a new application of NIRS, i.e., to detect focal cerebral ischemia and to monitor cerebral responses to therapy against vascular dysfunction in animal models.