Akin to cardiac stress testing, functional integrity of the peripheral vasculature can be interrogated by measuring the response to a stimulus. Recent reports suggest that the reactive hyperemia response, the physiologic reaction following induced ischemia, is associated with disease presence, correlated with disease severity, and may be a sensitive biomarker of pre-clinical disease. In this dissertation, an innovative, interleaved magnetic resonance imaging method is developed, termed Perfusion, Intravascular Venous Oxygen saturation, and T2* (PIVOT), which simultaneously measures microvascular perfusion, venous oxygen saturation (SvO2), and the blood-oxygen-level dependent (BOLD) signal. PIVOT is first applied in healthy subjects to demonstrate its ability to measure reactive hyperemia response dynamics. Next, reactive hyperemia perfusion is compared between the more temporally efficient pulsed arterial spin labeling (PASL) used in PIVOT and the more recently developed and preferred method for the brain, pseudo-continuous ASL (pCASL). Assessment of the impact of blood flow variability throughout the ischemia-reperfusion paradigm on pCASL perfusion quantification is investigated. Then, both PASL and pCASL sequences are used to measure reactive hyperemia perfusion in healthy subjects. No significant differences were detected between perfusion measured with PASL or pCASL despite different labeling strategies, temporal resolutions, and perfusion quantification models. Subsequently, PIVOT is combined with a velocity-encoded dual-echo GRE to create an interleaved three-slice sequence that provides quantification of bulk blood flow in the arteries and veins in addition to the traditional PIVOT measures. This new sequence, termed Velocity and PIVOT (vPIVOT) is used to investigate the relationship of blood flow in the macro- and microvasculature and muscle oxygen consumption during the transition from exercise to rest. Finally, PIVOT is applied clinically in a cohort of patients with varying degrees of severity of peripheral artery disease. Increasing disease severity was correlated with a prolongation of the hyperemic response time, measured as a lengthening of time to peak perfusion, SvO2 washout time, and time to peak T2*. In addition, peak perfusion and SvO2 upslope were significantly different between patients with PAD and healthy controls. These results suggest the potential for PIVOT to evaluate disease severity and may present a tool to assess response to therapeutic intervention.