Lower body negative pressure (LBNP) chambers can be utilized to experimentally-elicit reduction in blood pressure, cerebral blood flow (CBF), and associated presyncope symptoms. There is high variability in tolerance to LBNP between individuals, however, the underlying physiological responses affecting tolerance remain unclear. Pulsatility in CBF may affect LBNP tolerance, as more pulsatile flow may protect the delivery of oxygen to cerebral tissues during hypotension. We aimed to assess the pulsatility index (PI) in anterior and posterior cerebral circulation during LBNP superimposed head-up tilt (HUT), where gravitational-induced blood volume redistribution augments volume unloading during LBNP. We recruited and included 12 healthy male participants and instrumented them in a custom-built integrated 45° HUT-LBNP chamber. All experiments and protocols were reviewed and approved by the Mount Royal University Human Research Ethics Board (Ethics ID 2011-91Sa) and conformed to the Declaration of Helsinki and the Canadian Government Tri-Council Policy Statement on research ethics (TCPS2). All participants provided written informed consent prior to participation in this study. Participants were instrumented for heart rate (HR; ECG), end-tidal CO2 (PETCO2; mouthpiece and gas analyzer), beat-by-beat mean arterial pressure (MAP; finometer), and middle and posterior cerebral artery velocity (MCAv, PCAv; transcranial Doppler ultrasound). All measures were recorded during baseline (BL) and during -50 mmHg LBNP exposure up to a maximum of 10-minutes (600s). Presyncope was defined as a 30% reduction in systolic blood pressure (i.e., investigator stop) or onset subjective symptoms (i.e., participant stop). We quantified all variables as a 30-sec mean bin during BL and the final 30-sec of LBNP prior to presyncope using a paired t-test. MAP, MCAv, and PCAv PI were calculated as systolic-diastolic/mean. LBNP tolerance time was 480.9±134.3 sec. HR increased from 72.3±10.3 (BL) to 103.0±18.2 bpm (P<0.01) during LBNP, suggesting a baroreflex response, and PETCO2 was reduced from 32.0±2.9 (BL) to 28.7±2.9 Torr (P<0.01) during LBNP, suggesting mild hyperventilation. MAP decreased from 84.8±11.3 (BL) to 76.2±14.7 mmHg (P<0.01) during LBNP, with associated reductions in mean MCAv from 52.1±15.0 to 44.3±14.5 cm/s (P<0.01), and mean PCAv from 35.5±12.6 to 29.0±9.9 cm/s (P<0.01) during LBNP. MAP PI was reduced from 0.76±0.2 to 0.59±0.02 (P<0.01) during LBNP. However, MCAv and PCAv PI were unchanged during (P=0.6 and P=0.9, respectively). Our results suggest that although MAP and MAP PI were both significantly reduced during LBNP at pre-syncope, with associated reductions in mean MCAv and PCAv, MCAv and PCAv PI remained stable with LBNP in HUT LBNP likely due to cerebral autoregulation. These findings suggest that PI in CBF variables at the cardiac frequency are not related to LBNP tolerance in healthy men.