Background: Analysis of wavelet thresholds and energy components can be used to differentiate patients with congestive heart failure (CHF) from healthy subjects and to demonstrate the improvement of heart rate variability after treatment with beta-blockers in the patient group. Methods: Nine patients with CHF without concurrent beta-blocker therapy and 15 healthy volunteers were recruited and received 24-hour Holter recording. Wavelet thresholds and energy components were calculated. The patients with CHF were then treated with atenolol for 3 months. Another Holter recording was repeated and associated wavelet parameters were estimated. Results: Wavelet thresholds were statistically higher in normal subjects than those derived from patients with CHF (62.40 ± 20.65 vs. 16.90 ± 9.72, p < 0.001). The difference was still significant after control of heart rate and the total number of heart beats (74.85 ± 22.68 vs. 32.51 ± 18.73, p < 0.001). The energy component of HRV in the finest wavelet domain was also significantly higher in the normal subjects (243.01 ± 237.69 × 10<sup>–6</sup> vs. 40.65 ± 39.65 × 10<sup>–6</sup>, p = 0.001). These two measures increased after treatment with beta-blockers in the patients with CHF, but not to the extent of those of normal subjects. Conclusion: Analysis of wavelet thresholds and energy components has provided robust and accurate measurements for background HRV.