Our previous study examined the neural effects of extremely low frequency electric field (ELF-EF) using electroencephalogram (EEG) and heart rate variability (HRV) measurement and revealed that the theta power of EEG increases during 1-min EF treatment (30 kV and 50 Hz). In the present study, the effects of a longer treatment duration were assessed. EF was applied between two electrodes, one above the head and the other under the feet, for 20 min to 10 healthy men sitting on a chair. Occipital EEG and HRV were measured in the eyes-open condition. Power spectrum analysis showed that during the EF treatment, the power of alpha and theta rhythms as well as the low frequency component of HRV increased, and the heart rate decreased. These effects did not persist after the end of EF treatment. It is suggested that ELF-EF treatment alters not only the brain activity but also the autonomic activity reflected in HRV.