The effects of sympathetic activity on vasoconstriction are dampened in active skeletal muscle during exercise, a phenomenon termed functional sympatholysis. Limited work has examined the influence of sex on the magnitude of sympatholysis or the test-retest reliability of measurements. In 16 women and 15 men, forearm blood flow (FBF; Doppler ultrasound), muscle oxygenation (near-infrared spectroscopy; NIRS) and beat-to-beat mean arterial pressure (MAP; photoplethysmography) were measured during lower-body negative pressure (LBNP; -20mmHg) at rest and simultaneously during rhythmic handgrip exercise (30% maximum contraction). Measures were taken twice within the same visit (separated by 15-min) and repeated on a second visit. Forearm vascular conductance (FVC) was calculated as FBF/MAP. The magnitude of sympatholysis was calculated as the difference of LBNP-induced changes between handgrip and rest. LBNP decreased FBF (∆-45±15%), FVC (∆-45±16%), and muscle oxygenation (∆-14±11%); however, these responses were attenuated when LBNP was applied during rhythmic handgrip exercise (∆-7±9%, ∆-9±10%, and ∆-6±9%, respectively). The magnitude of sympatholysis was not different between men and women (FBF: 40±16% vs. 35±9%, P=0.37; FVC: 38±16% vs. 35±11%, P=0.53; muscle oxygenation: 5±9% vs. 11±11%, P=0.11). Furthermore, sympatholysis measurements demonstrated good to excellent intra-day (intraclass-correlation coefficients; ICC≥0.85) and inter-day (ICC≥0.72) test-retest reliability (all P£0.01) in both sexes. The coefficients of variation were larger with NIRS (68-91%) than Doppler ultrasound (16-22%) assessments of functional sympatholysis. Collectively, these findings demonstrate that assessments of functional sympatholysis are not impacted by biological sex and that Doppler ultrasound-derived measures of sympatholysis have better within-subject reliability than NIRS-derived measures in young healthy adults.