Introduction: Veteran athletes have a higher incidence of pacemaker implantation due to an increased susceptibility to bradyarrhythmias and heart block. Previously we showed that training-induced bradycardia is due to a downregulation of the funny current If (a key pacemaker current) and the corresponding channel HCN4. Here we investigated If and HCN4 expression in the atrioventricular node (AVN) following training. Methods: 10-week-old male C57BL/6J mice were trained by swimming for 60 min twice/day for 28 days and compared to sedentary littermates. The AVN was dissected and cells were dissociated by enzymatic digestion for whole cell patch clamp. If was recorded from AVN cells during hyperpolarising pulses from a holding potential of -35 mV in -20 mV increments till -135 mV. Activation and deactivation time constants were measured at -100, -40 and 0 mV using suitable protocols. In another set of experiments, 10-week-old male C57BL/6J mice were trained by swimming for 60 min/day, 5 days/week for 5 months following which AVN preparations were harvested and frozen. Total RNA was isolated from serial 20 μm sections of the AVN, identified by HCN4 immunolabelling and collected by laser capture microdissection. Expression of HCN channels was measured by Taqman qPCR. Results: The AVN preparations encompassed the compact node, penetrating bundle, His bundle and tricuspid valve. There was considerable heterogeneity in isolated AVN cells, including classical spindle-shaped nodal cells and small atrial-like nodal cells. Whole cell patch clamp was only performed on spindle-shaped cells. If was present in 40% of control (n=15) and 28% of trained (n=21) AVN myocytes. The cell capacitance of AVN cells exhibiting If was increased from 13.7±1.1 pF (n=9) in control to 16.6±1.2 pF (n=15) in trained AVN cells (P=0.06). The density of If was reduced by 18.2% from 33.6±2.5 pA/pF (n=9) in control to 27.5±1.6 pA/pF (n=14) in trained AVN cells (P<0.05). Consistent with this, the trained AVN presented with a striking reduction in HCN4 mRNA (8.6±2.1 in control versus 0.7±0.1 in trained AVN, P<0.05, n=5-6). Expression levels of HCN1 and HCN2 were unaltered. Conclusion: Endurance exercise results in electrophysiological remodelling of the AVN characterised by a reduction in If and HCN4. If has been implicated in AVN conduction and the downregulation in If and HCN4 following training might contribute to the occurrence of heart block in veteran athletes.