Abstract Cerebrovascular reactivity (CVR) represents an important marker of brain vascular health, particularly in the context of small and large vessel diseases. However, an undesired feature of this measure is that there exist large variations in CVR values across individuals, which is mainly attributed to physiological factors. Here, we test the hypothesis that caffeine, a widely consumed neurostimulant, has a significant effect on CVR measured with MRI. Sixteen young healthy participants were enrolled and categorized into caffeine-naive (N = 8) and caffeine-habituated (N = 8) groups based on their caffeine consumption habits. CVR was assessed via CO2 inhalation using two different MRI methods, phase-contrast (PC) cerebral blood flow (CBF), and T2*-EPI Blood-Oxygenation-Level-Dependent (BOLD)-MRI. Each participant underwent two MRI sessions, one before and the other after an oral administration of 200 mg of caffeine. Additionally, venous oxygenation (Yv) was measured using T2-Relaxation-Under-Spin-Tagging (TRUST) MRI. For basal physiological parameters, a significant caffeine-induced CBF decrease was observed in both naive (p = 0.002) and habituated (p < 0.001) groups. The caffeine-naive group exhibited a 31.2 ± 14.1% reduction in basal CBF, whereas the caffeine-habituated group showed a 16.7 ± 5.0% reduction, revealing significant differences between groups (p = 0.04). A similar observation was seen in basal Yv, with caffeine-naive participants showing a greater (p = 0.02) reduction (21.5 ± 8.9%) than the habituated participants (7.6 ± 10.1%). CBF-CVR decreased significantly in both groups: from 4.5 ± 0.9 to 3.0 ± 0.9 %CBF/mmHg of CO2 (33.3 ± 14.1%, p < 0.001) in the caffeine-naive group, and from 5.1 ± 1.5 to 3.7 ± 1.3 %CBF/mmHg of CO2 (27.3 ± 16.0%, p = 0.009) in the caffeine-habituated group. No significant differences were observed between groups in terms of the extent of CVR reduction (p = 0.23). BOLD-CVR showed modest reduction after caffeine administration, from 0.17 ± 0.04 %/mmHg to 0.15 ± 0.05 %/mmHg (14.1 ± 16.8%, p = 0.02). There was no difference between the participant groups in terms of BOLD-CVR reduction following caffeine consumption. This study suggests that investigations using CVR as a disease marker may benefit from accounting for the caffeine consumption and/or its blood concentration in the participants.