PURPOSE: To utilize power spectral analysis (PSA) of heart rate variability (HRV) as a pharmacodynamic (PD) measure of atropine parasympathetic effect, and to model the kinetics of action. METHODS: Heart rate data was collected following atropine administration to rats and was analyzed off-line for high frequency peaks by PSA of HRV as a measure of parasympathetic tone. A temporal cumulative approach (TCA) detected transient changes in parasympathetic activity. The pharmacokinetics (PK) was analyzed and linked to both direct and indirect PK-PD models. RESULTS: TCA enabled a quantitative measure of atropine parasympathetic activity. A simultaneous fit of the indirect PK-PD model to the experimental data of all three atropine doses successfully captured the experimental data. CONCLUSIONS: TCA can be used as a quantitative measure of parasympathetic tone. Our work has established a preclinical model to investigate the kinetics of drug action on the autonomic nervous system.