Car driving is considered a very complex activity, consisting of different concomitant tasks and subtasks, thus it is crucial to understand the impact of different factors, such as road complexity, traffic, dashboard devices, and external events on the driver’s behavior and performance. For this reason, in particular situations the cognitive demand experienced by the driver could be very high, inducing an excessive experienced mental workload and consequently an increasing of error commission probability. In this regard, it has been demonstrated that human error is the main cause of the 57% of road accidents and a contributing factor in most of them. In this study, 20 young subjects have been involved in a real driving experiment, performed under different traffic conditions (rush hour and not) and along different road types (main and secondary streets). Moreover, during the driving tasks different specific events, in particular a pedestrian crossing the road and a car entering the traffic flow just ahead of the experimental subject, have been acted. A Workload Index based on the Electroencephalographic (EEG), i.e., brain activity, of the drivers has been employed to investigate the impact of the different factors on the driver’s workload. Eye-Tracking (ET) technology and subjective measures have also been employed in order to have a comprehensive overview of the driver’s perceived workload and to investigate the different insights obtainable from the employed methodologies. The employment of such EEG-based Workload index confirmed the significant impact of both traffic and road types on the drivers’ behavior (increasing their workload), with the advantage of being under real settings. Also, it allowed to highlight the increased workload related to external events while driving, in particular with a significant effect during those situations when the traffic was low. Finally, the comparison between methodologies revealed the higher sensitivity of neurophysiological measures with respect to ET and subjective ones. In conclusion, such an EEG-based Workload index would allow to assess objectively the mental workload experienced by the driver, standing out as a powerful tool for research aimed to investigate drivers’ behavior and providing additional and complementary insights with respect to traditional methodologies employed within road safety research.