Object: There is an increasing interest in preoperative diffusion tensor imaging-based fiber tracking (DTI-FT) to preserve function during surgeries in motor eloquent brain regions. However, DTI tractography is challenged by inherent presumptions during particular tracking steps [e.g., deterministic vs. probabilistic DTI, fractional anisotropy (FA) and fiber length (FL) thresholding] and the missing "ground truth" information. In the present study, we intended to establish an objective, neurophysiology-driven approach for parameter selection during DTI-FT of the corticospinal tract integrating both imaging and neurophysiological information. Methods: In ten patients with lesions in eloquent motor areas, preoperative navigated transcranial magnetic stimulation (nTMS) was performed, followed by individual deterministic DTI-FT from a grid of cortical seed points. We investigated over 300 combinations of FA and FL thresholds and applied subsequently a multidimensional mathematical modeling of this empirical data. Optimal DTI parameters were determined by the relationship between DTI-FT (i.e., number of fibers, NoF) and nTMS (i.e., amplitudes of motor-evoked potentials) results. Finally, neurophysiological DTI parameters and the resulting tractography were compared to the current standard approaches of deterministic DTI fiber tracking with a 75% and 50% FA and a FL threshold of 110 mm as well as with intraoperative direct cortical and subcortical stimulation. Results: There was a good goodness-of-fit for the mathematical model (r 2 = 0.68 ± 0 13; range: 0.59-0.97; n = 8) except of two cases. Neurophysiology-driven parameter selection showed a high correlation between DTI-FT and nTMS results (r = 0.73 ± 0.16; range: 0.38-0.93). In comparison to the standard approach, the mathematically calculated thresholds resulted in a higher NoF in 75% of patients. In 50% of patients