Cyber-attacks can compromise the reliability of onboard digital systems, navigation, sensing, control and decision support. An undetected attack may mislead bridge teams and increase the risk of collisions and groundings. Building maritime cyber resilience is therefore essential to the safety of navigation. In this study, 61 certificated seafarers participated in an experimental full-mission bridge simulator study replicating an Istanbul Strait transit. Participants were allocated to a test group or a control group and were exposed to a timed GNSS (Global Navigation Satellite) System spoofing event in a single continuous run. A multimodal dataset was acquired using physiological measures, including EEG and eye-tracking, together with objective and subjective instruments. Relative to the routine briefing, the cyber-situational awareness briefing produced statistically significant improvements across modalities. The test group exhibited higher EEG-derived vigilance (p = 0.027) and lower stress and workload than the control group. Participants’ self-reports indicated higher anomaly awareness, more frequent noticing of unusual device behaviour and greater suspicion (p = 0.006; p = 0.004). Eye-tracking metrics revealed more independent cross-verification, with reduced reliance on the primary electronic display. Objective performance, summarised by the composite performance index, was greater in the test group (p < 0.001), and the proportion of safe passages was higher. This study investigated whether a situational awareness intervention favourably shifts human factors, trust calibration, and performance under cyber-attacks, specifically GNSS spoofing. To our knowledge, this research provides the first empirical evidence from seafarers exposed to cyberattacks in a simulator environment and offers new insights into maritime human factors.