Aim: 

We created a mice line in which the exon 2 of HCN4 gene could be eliminated in a conditional and inducible manner to evaluate the role of the cardiac pacemaker current in an intact animal model.

Methods: 

The experimental strategy adopted to induce the knockout is based on a recombination process controlled by the MerCreMer-loxP system. The knockout event was induced by the administration of the estrogen agonist (tamoxifen) which was able to induce the elimination of the exon 2 of the HCN4 gene selectively in the cardiac region.

Results: 

The first striking evidence is that KO induction is lethal: 15 animals were treated with tamoxifen and all of them died within 3–8 days after tamoxifen administration; 17 control animals were exposed to the same treatment and all of them survived. Immunofluorescence experiments on single SAN cells and on SAN tissue slices obtained from tamoxifen-treated KO mice confirmed the successful elimination of the HCN4 protein. Telemetric ECG recordings revealed the development of a strong level of bradycardia upon KO induction; heart rate dropped to 42 21% (n = 5) of the control values (pre-tamoxifen) 6 days after the beginning of tamoxifen treatment. Similar experiments carried out in control animals did not result in any rate modifications.

Conclusion: 

Our results indicate that the elimination of HCN4 cardiac channels leads to a marked bradycardia and to the death of the animals. Our experiments thus highlight the relevance of the pacemaker current in maintaining the heart rate to levels compatible with life in an intact animal. These data markedly differ from those reported in the literature for global HCN4 KO mice and a clear understanding of these differences must be investigated.