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Introduction: Atrial fibrillation (AF) is the most common arrhythmia, associated with significant burdens to patients and the healthcare system. The atrioventricular (AV) node plays a vital role in regulating heart rate during AF by filtering electrical impulses from the atria. However, it is often insufficient in regards to maintaining a healthy heart rate, thus the AV node properties are modified using rate-control drugs. Moreover, treatment selection during permanent AF is currently done empirically. Quantifying individual differences in diurnal and short-term variability of AV-nodal function could aid in personalized treatment selection. Methods: This study presents a novel methodology for estimating the refractory period (RP) and conduction delay (CD) trends, and their uncertainty in the two pathways of the AV node during 24 h using non-invasive data. This was achieved by utilizing a network model together with a problem-specific genetic algorithm and an approximate Bayesian computation algorithm. Diurnal variability in the estimated RP and CD was quantified by the difference between the daytime and nighttime estimates, and short-term variability was quantified by the Kolmogorov-Smirnov distance between adjacent 10-min segments in the 24-h trends. Additionally, the predictive value of the derived parameter trends regarding drug outcome was investigated using several machine learning tools. Results: Holter electrocardiograms from 51 patients with permanent AF during baseline were analyzed, and the predictive power of variations in RP and CD on the resulting heart rate reduction after treatment with four rate control drugs was investigated. Diurnal variability yielded no correlation to treatment outcome, and no prediction of drug outcome was possible using the machine learning tools. However, a correlation between the short-term variability for the RP and CD in the fast pathway and resulting heart rate reduction during treatment with metoprolol (ρ = 0.48, p < 0.005 in RP, ρ = 0.35, p < 0.05 in CD) were found. Discussion: The proposed methodology enables non-invasive estimation of the AV node properties during 24 h, which--indicated by the correlation between the short-term variability and heart rate reduction--may have the potential to assist in treatment selection. [ABSTRACT FROM AUTHOR]