Introduction Sex-related differences in microvascular reactivity may be influenced by the magnitude of muscle desaturation (ischemic stimulus) during the blood flow occlusion period. Thus, this study used near-infrared spectroscopy (NIRS) to assess skeletal muscle microvascular reactivity in three distinct metabolic and mechanical conditions. Methods Nineteen participants (10 males, 9 females) completed a randomized parallel design including a vascular occlusion test (VOT), maximal voluntary isometric contraction (MVIC), and a dynamic fatigue exercise protocol (FEx) to elicit ischemic, mechanic and metabolic insults, respectively. Morphological muscle quality was also assessed using ultrasonography. NIRS-derived indices of tissue oxygenation and blood volume were continuously monitored. Results Males exhibited a greater post-exercise total hemoglobin reperfusion slope ([tHb] slope) across protocols compared with females (p textless 0.05). In males, higher morphological muscle quality was associated with a steeper reperfusion slope (p textless 0.05), whereas no such relationships were observed in females. Conclusion Sex differences in microvascular reactivity were evident across ischemic, isometric, and dynamic exercise conditions, with oxygen extraction only partially accounting for sex differences in microvascular reactivity in highly metabolic demand condition such as FEx. Our findings also suggest that the greater microvascular reactivity in males may be linked to better morphological muscle quality compared to females, thus providing novel insights into the mechanisms driving the accelerated reperfusion rates observed in males compared to females.