Intraoperative neurophysiological monitoring is used to assess neurological function during surgeries placing the spinal cord at risk. Transcranial electrical stimulation muscle motor evoked potentials (Tc-mMEPs) are used to monitor motors tracts, but their interpretation is complicated by the large temporal variability which can result in false-positive warnings. Although the choice of anesthetic drug regimen and drug dose are often claimed to be factors causing this variability, the relationship between depth of anesthesia, quantified by processed electroencephalogram (pEEG) parameters, and Tc-mMEPs characteristics in upper and lower extremity muscles have not yet been rigorously investigated in patients receiving propofol/remifentanil-based anesthesia. Twenty-five patients were included in this prospective observational study. All received propofol/remifentanil-based total intravenous anesthesia. Depth of anesthesia was quantified by the bispectral index (BIS). After induction of anesthesia, the target propofol concentration was altered to sequentially achieve BIS values of either 30, 40, and 50, or the reverse (direction randomly determined). At each depth of anesthesia Tc-mMEP thresholds were determined, and arterial blood samples were collected. Supramaximal Tc-mMEP signals were recorded every 2 minutes and amplitudes, latencies and area under the curve (AUC) were subsequently calculated. Effects of depth of anesthesia on Tc-mMEP outcomes were analyzed using linear mixed effects modeling. The median (range) age of the study population was 18 (14-66) years (n = 25). In the leg muscles, a decrease of 10 BIS points was associated with a decrease in Tc-mMEP amplitude of 11%-12% (all P < .001; mean [95% confidence interval \CI], 12% [7.1-16], 11% [6.8-16], and 12% [7.5-16], for the AH, TA, and GAS muscles, respectively). In contrast, no significant amplitude or AUC change was found in the hand muscles (P = .201, 2.8% [-1.5 to 7.1] and P = .076, 4.0% [-0.4 to 7.6], respectively). Latencies changed <0.5% per 10 BIS points decrease (0.03% [-0.3 to 0.2], -0.2% [-0.5 to 0.1], -0.2% [-0.5 to 0.04], 0.3% [0.04-0.6] for the AH, TA, GAS, and hand muscles, respectively), and thresholds increased 3.6% (0.8-7) when BIS decreased from 50 to 30 (P = .037). Our findings challenge some commonly held beliefs. First, our findings suggest that deeper anesthesia has differential effects on the different muscle groups, with little effect on the hand muscles. The current practice of using the hand signals as reference values during procedures below C8/T1 may therefore need re-evaluation. Second, the paucity of effect of depth of anesthesia on Tc-mMEP thresholds and latencies suggests that Tc-mMEP generation is not influenced by deep anesthesia in a clinically relevant way. Therefore, the threshold level monitoring method may provide a more reliable indicator of motor pathway integrity during surgery. This could reduce the likelihood of false-positive warnings and unnecessary interventions.