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The intraluminal filament Middle Cerebral Artery Occlusion (MCAO) model is widely used to study animal models of cerebral ischemia. Laser Speckle Contrast Imaging (LSCI) provides an approach to monitor changes in cortical Cerebral Blood Flow (CBF) in mice through intact skull. However, the MCAO surgery requires a supine position of animals during preparation, where the conventional LSCI systems cannot achieve to provide a real-time high spatio-temporal resolution CBF information. Herein, we implemented an optimized method to uninterruptedly measure the relative CBF during surgical procedures by designing an inverted LSCI (i-LSCI). This approach achieves a real-time high spatio-temporal resolution CBF information during the MCAO surgery, enabling a real-time monitoring of cerebral collateral circulation and assessment of the stability of the ischemic site immediately after MCAO. Moreover, this optimization allows for a detection of Cortical Spreading Depolarization (CSD) in ischemic cortex immediately after MCAO.

The quantitative measurement of the microvascular blood-flow velocity is critical to the early diagnosis of microvascular dysfunction, yet there are several challenges with the current quantitative flow velocity imaging techniques for the microvasculature. Optical flow analysis allows for the quantitative imaging of the blood-flow velocity with a high spatial resolution, using the variation in pixel brightness between consecutive frames to trace the motion of red blood cells. However, the traditional optical flow algorithm usually suffers from strong noise from the background tissue, and a significant underestimation of the blood-flow speed in blood vessels, due to the errors in detecting the feature points in optical images. Here, we propose a temporal direction filtering and peak interpolation optical flow method (TPIOF) to suppress the background noise, and improve the accuracy of the blood-flow velocity estimation. In vitro phantom experiments and in vivo animal experiments were performed to validate the improvements in our new method.