The amplitude of respiratory sinus arrhythmia (RSA) was measured in eight healthy young male students with special reference to the effect of tidal volume (Vt). Under simultaneously controlled respiratory frequency and Vt, the heart rate variability (HRV) of the subjects was measured. While the respiratory frequency was adjusted to either 0.25 or 0.10 Hz, the Vt was controlled at 13 different volumes for each frequency. Linear relationships between RSA amplitude and Vt were observed and close correlations were obtained for 0.25 Hz compared with 0.10 Hz. However, regression equations showed a marked variation among subjects. Furthermore, RSA amplitude was related to vital capacity. Subjects who had lower vital capacity tended to show higher RSA amplitudes at the same Vt. Therefore, the ratio (% Vt) of Vt to vital capacity is a more effective index in normalizing RSA than raw tidal volume. From these results, we have proposed a normalized RSA (RSA amplitude/% Vt) as a new index of autonomic activity that provides a constant value regardless of Vt.