The structural and functional organization of brain networks subserving basic daily activities (i.e. language, visuo-spatial cognition, movement, semantics, etc.) are not completely understood to date. Here, we report the first probabilistic cortical and subcortical atlas of critical structures mediating human brain functions based on direct electrical stimulation (DES), a well-validated tool for the exploration of cerebral processing and for performing safe surgical interventions in eloquent areas. Methods: We collected 1162 cortical and 659 subcortical DES responses during testing of 16 functional domains in 256 patients undergoing awake surgery. Spatial coordinates for each functional response were calculated, and probability distributions for the entire patient cohort were mapped onto a standardized three-dimensional brain template using a multinomial statistical analysis. In addition, matching analyses were performed against prior established anatomy-based cortical and white matter (WM) atlases. Results: The probabilistic maps for each functional domain were provided. The topographical analysis demonstrated a wide spatial distribution of cortical functional responses, while subcortical responses were more restricted, localizing to known WM pathways. These DES-derived data showed reliable matching with existing cortical and WM atlases as well as recent neuroimaging and neurophysiological data. Conclusions: We present the first integrated and comprehensive cortical-subcortical atlas of structures essential for humans' neural functions based on highly-specific DES mapping during real-time neuropsychological testing. This novel atlas can serve as a complementary tool for neuroscientists, along with data obtained from other modalities, to improve and refine our understanding of the functional anatomy of critical brain networks.