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We exploited two-photon microscopy and Doppler optical coherence tomography to examine the cerebral blood flow and tissue pO₂ response to forced treadmill exercise in awake mice. To our knowledge, this is the first study performing both direct measure of brain tissue pO₂ during acute forced exercise and underlying microvascular response at capillary and non-capillary levels. We observed that cerebral perfusion and oxygenation are enhanced during running at 5 m/min compared to rest. At faster running speeds (10 and 15 m/min), decreasing trends in arteriolar and capillary flow speed were observed, which could be due to cerebral autoregulation and constriction of arterioles in response to blood pressure increase. However, tissue pO₂ was maintained, likely due to an increase in RBC linear density. Higher cerebral oxygenation at exercise levels 5–15 m/min suggests beneficial effects of exercise in …