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Transcutaneous electrical nerve stimulation (TENS) is a noninvasive modality used to control pain. Animal models show that repeated TENS application produces analgesic tolerance and cross-tolerance at spinal opioid receptors. The aim of the present investigation was to examine whether repeated application of TENS produces analgesic tolerance in humans. One hundred healthy subjects were randomly assigned to 1 of 4 groups: control, placebo, low-frequency (4Hz) or high-frequency (100Hz) TENS. TENS was applied daily for 5days to the nondominant upper limb; pressure-pain threshold (PPT) measurements were recorded before and after TENS. Temporal summation to mechanical stimulation was recorded on days 1 and 5, before and after TENS. Diffuse noxious inhibitory control (DNIC) was tested on day 5 using the cold pressor test and PPT measurements. There was an initial increase in PPTs in both low- and high-frequency TENS groups when compared with placebo or control groups. However, by day 5 this TENS-induced increase in PPT did not occur, and there was no difference between active TENS and placebo or control groups. High-frequency TENS decreased temporal summation on day 1 when compared with day 5. DNIC increased the PPT similarly in all groups. These data suggest that repeated daily application of TENS results in a decrease in its hypoalgesic effect by the fifth day and that the tolerance-like effect to repeated TENS results from tolerance at centrally located opioid receptors. The lack of change in DNIC response suggests that TENS and DNIC utilize separate pathways to produce analgesia. Repeated high-frequency and low-frequency transcutaneous electrical nerve stimulation produce analgesic tolerance in humans by the fourth and fifth day of treatment, respectively.