Purpose <p>Hypotension, or low mean arterial blood pressure (MAP), has been associated with adverse outcomes in perioperative patients. A primary goal of treating hypotension during surgery is to preserve vital organ perfusion by maintaining intravascular volume and the use of vasoactive medications, including phenylephrine (PE), to support MAP. Phenylephrine, a vasoconstrictor with α₁-adrenergic agonist activity, acts on resistance arterioles and veins to increase vascular resistance and reduce venous capacitance, thereby increasing MAP. It predominantly acts on skeletal muscle resistance arterioles, but concerns have been raised about its potential negative impact on brain and revascularized muscle flap perfusion.</p> Methods <p>We conducted an <i>in vivo</i> animal study using translational rodent models. Measuring microvascular blood flow (laser Doppler) and partial pressure of oxygen (PO₂) (phosphorescence quenching of oxygen) in rats (total <i>N</i> = 48), we sought to test the hypothesis that PE produces differential effects on brain, skeletal muscle, and skeletal muscle flap perfusion.</p> Results <p>Treatment of hypotension with PE increased MAP, brain microvascular blood flow, and brain tissue PO₂, at the expense of reduced skeletal muscle microvascular blood flow. Escalating doses of PE reduced skeletal muscle microvascular blood flow without reducing tissue PO₂. Tissue blood flow and PO₂ were severely reduced in skeletal muscle free flaps at baseline, without any further reduction after exposure to escalating doses of PE. Elevation of the hypoxic cellular protein hypoxia-inducible factor 1α (HIF-1α) in muscle free flaps provided evidence of severe tissue hypoxia in viable muscle flap tissue.</p> Conclusions <p>These data may inform the optimal use of PE to restore MAP to ensure optimal brain tissue perfusion. Use of tissue oximetry may ensure the adequacy of tissue perfusion in perioperative patients.</p>