With the increasing number of manned space missions and human exploration of extreme environments such as deep-sea and polar regions, researching the environmental design of confined working spaces, particularly the impact of lighting on personnel comfort and work performance, has become important. Lighting not only affects visual perception but also exerts effects through non-visual pathways on aspects such as pupil constriction, hormone levels, cognitive abilities, and alertness, which in turn influence work performance. The combination of lighting layout and illuminance levels in confined spaces can affect on operators’ visual perception and fatigue levels. This study simulates different lighting scenarios through experiments, integrating non-visual effects and task load, to assess the impact of lighting layout types and illuminance levels on both physiological and psychological fatigue. Six distinct lighting layout schemes were designed, and their effects on operator fatigue levels during neurobehavioral tasks were analyzed. Fatigue was assessed based on multimodal fusion, including task reaction time, eye aspect ratio, blink frequency, and task load. The results indicate that the combination of illuminance level and layout type has a significant impact on operator fatigue. Low illuminance environments (300 lux) lead to higher levels of fatigue, while high illuminance environments (700 lux) can alleviate fatigue. The differences in layout types also play an important role under different illuminance conditions, especially under low illuminance, where linear layouts showed better results in terms of reaction time, blink frequency, and task load perception. This study provides theoretical support for optimizing lighting design in confined spaces.