Aim: Pacemaking in the sino-atrial node (SAN) is modulated by hyperpolarization-activated cyclic nucleotide (HCN) gated channel, I_f. Several isoforms of the HCN family have been identified in murine SAN which show vast differences in time kinetics. This computer modelling study evaluates the functional role of individual HCN isoforms on murine pacemaker activity. Methods: Murine SAN I_f consists of HCN1, HCN2 and HCN4 [1]. All isoforms have similar steady state activtion [2] with a half-activation of approximately -63.7 mV. The molecular expression of HCN1, HCN2 and HCN4 in SAN cells has been shown to be 5:25:70 respectively [3]. The primary biophysical differences in the individual isoform function have been identified to be a quantitative difference in time kinetics and cAMP sensitivities [4]. Based on these biophysical properties, Hodgkin-Huxley models for each isoformal component of I_f were developed and incorporated into our recently developed murine SAN mathematical cell model. The contributions of individual isoforms to model action potential (AP) were evaluated by blocking the isoformal channels individually. Model responses were defined as AP features including minimum diastolic potential (MDP), over shoot potential (OS), AP duration at 50% (APD₅₀) and at 90% repolarisation (APD₉₀), cycle length (CL) and diastolic depolarisation rate (DDR) [5]. Blocking of total I_f and Control cases were also simulated. Results: Blocking of I_f or its isoforms had marked effects on CL and DDR with other AP features largely unaffected. The results are summarised in the table. Blocking of total I_f increased the CL by 26% and the DDR reduced by 5%. Selectively blocking HCN1 isoform gave a small prolongation of CL by 2%. Blocking of HCN2 however, prolonged CL by 28% and DDR was reduced by 37%. Blocking of HCN4 gave a relatively modest prolongation of CL by 2.5%. Conclusions: I_f regulates murine SAN pacemaking during the slow diastolic depolarisation. Altough the slowly activating HCN4 is the most extensively expressed, HCN2 is the isoform majorly contributing to murine pacemaking electrical activity. On the other hand, the contribution of HCN1 is small due to its small conductance. The HCN isoforms have different functional impacts on murine pacemaking.