Siebenmann C, Ryrsø CK, Oberholzer L, Fisher JP, Hilsted LM, Rasmussen P, Secher NH, Lundby C. Hypoxia-induced vagal withdrawal is independent of the hypoxic ventilatory response in men. J Appl Physiol 126: 124 –131, 2019. First published November 29, 2018; doi:10.1152/japplphysiol.00701.2018.—Hypoxia increases heart rate (HR) in humans by sympathetic activation and vagal withdrawal. However, in anaesthetized dogs hypoxia increases vagal activity and reduces HR if pulmonary ventilation does not increase and we evaluated whether that observation applies to awake humans. Ten healthy males were exposed to 15 min of normoxia and hypoxia (10.5% O2), while respiratory rate and tidal volume were volitionally controlled at values identified during spontaneous breathing in hypoxia. End-tidal CO2 tension was clamped at 40 mmHg by CO2 supplementation. -Adrenergic blockade by intravenous propranolol isolated vagal regulation of HR. During spontaneous breathing, hypoxia increased ventilation by 3.2 2.1 l/min (P 0.0033) and HR by 8.9 5.5 beats/min (P 0.001). During controlled breathing, respiratory rate (16.3 3.2 vs. 16.4 3.3 breaths/min) and tidal volume (1.05 0.27 vs. 1.06 0.24 l) were similar for normoxia and hypoxia, whereas the HR increase in hypoxia persisted without (8.6 10.2 beats/min) and with (6.6 5.6 beats/min) propranolol. Neither controlled breathing (P 0.80), propranolol (P 0.64), nor their combination (P 0.89) affected the HR increase in hypoxia. Arterial pressure was unaffected (P 0.48) by hypoxia across conditions. The hypoxia-induced increase in HR during controlled breathing and -adrenergic blockade indicates that hypoxia reduces vagal activity in humans even when ventilation does not increase. Vagal withdrawal in hypoxia seems to be governed by the arterial chemoreflex rather than a pulmonary inflation reflex in humans. NEW & NOTEWORTHY Hypoxia accelerates the heart rate of humans by increasing sympathetic activity and reducing vagal activity. Animal studies have indicated that hypoxia-induced vagal withdrawal is governed by a pulmonary inflation reflex that is activated by the increased pulmonary ventilation in hypoxia. The present findings, however, indicate that humans experience vagal withdrawal in hypoxia even if ventilation does not increase, indicating that vagal withdrawal is governed by the arterial chemoreflex rather than a pulmonary inflation reflex.