AIMS: Atrial fibrillation (AF) is the most frequent arrhythmia. Screening of SCN5A-the gene encoding the a-subunit of the cardiac sodium channel-has indicated that disturbances of the sodium current may play a central role in the mechanism of lone AF. We tested the hypothesis that lone AF in young patients is associated with genetic mutations in SCN3B and SCN4B, the genes encoding the two ß-subunits of the cardiac sodium channel. METHODS AND RESULTS: In 192 unrelated lone AF patients, the entire coding sequence and splice junctions of SCN3B and SCN4B were bidirectionally sequenced. Three non-synonymous mutations were found in SCN3B (R6K, L10P, and M161T). Two mutations were novel (R6K and M161T). None of the mutations were present in the control group (n = 432 alleles), nor have any been previously reported in conjunction with AF. All SCN3B mutations affected residues that are evolutionarily conserved across species. Electrophysiological studies on the SCN3B mutation were carried out and all three SCN3B mutations caused a functionally reduced sodium channel current. One synonymous variant was found in SCN4B. CONCLUSION: In 192 young lone AF patients, we found three patients with suspected disease-causing non-synonymous mutations in SCN3B, indicating that mutations in this gene contribute to the mechanism of lone AF. The three mutations in SCN3B were investigated electrophysiologically and all led to loss of function in the sodium current, supporting the hypothesis that decreased sodium current enhances AF susceptibility.