We investigated whether the eccrine sweat glands must actively produce sweat during heat acclimation if they are to adapt and increase their capacity to sweat. Eight volunteers received intradermal injections of BOTOX, to prevent neural stimulation and sweat production of the sweat glands during heat acclimation, and saline injections as a control in the contralateral forearm. Subjects performed 90 min of moderate-intensity exercise in the heat (35 degrees C, 40% relative humidity) on 10 consecutive days. Heat acclimation decreased end-exercise heart rate (156 +/- 22 vs. 138 +/- 17 beats/min; P = 0.0001) and rectal temperature (38.2 +/- 0.3 vs. 37.9 +/- 0.3 degrees C; P = 0.0003) and increased whole body sweat rate (0.70 +/- 0.29 vs. 1.06 +/- 0.50 l/h; P = 0.030). During heat acclimation, there was no measurable sweating in the BOTOX-treated forearm, but the control forearm sweat rate during exercise increased 40% over the 10 days (P = 0.040). Peripheral sweat gland function was assessed using pilocarpine iontophoresis before and after heat acclimation. Before heat acclimation, the pilocarpine-induced sweat rate of the control and BOTOX-injected forearms did not differ (0.65 +/- 0.20 vs. 0.66 +/- 0.22 mg x cm(-2) x min(-1)). However, following heat acclimation, the pilocarpine-induced sweat rate in the control arm increased 18% to 0.77 +/- 0.21 mg x cm(-2) x min(-1) (P = 0.021) but decreased 52% to 0.32 +/- 0.18 mg x cm(-2) x min(-1) (P < 0.001) in the BOTOX-treated arm. Using complete chemodenervation of the sweat glands, coupled with direct cholinergic stimulation via pilocarpine iontophoresis, we demonstrated that sweat glands must be active during heat acclimation if they are to adapt and increase their capacity to sweat.