Molecular and electrical remodeling of ion channels determining action potential duration has been proposed as a major mechanism in chronic atrial fibrillation. We investigated the mRNA expression of the cardiac L-type Ca²⁺-channel subunits α_1c, α₂/δ_1a, and β_1b/c in atrial tissue of patients with chronic atrial fibrillation compared to patients in sinus rhythm. In addition, the mRNA expression of the 5-hydroxytryptamine type 4-, β₁-, and β₂-adrenergic receptors, which are known to stimulate the L-type Ca²⁺-current in human atrium, was analyzed and the effect of chronic β-blocker treatment on the mRNA expression of these receptors and of the L-type Ca²⁺-channel subunits was assessed. Total RNA was isolated from right atrial appendages of patients in sinus rhythm and of patients with chronic atrial fibrillation. Then, semiquantitative RT-PCR using 18 S RNA as the “housekeeping gene” was performed. In patients with chronic atrial fibrillation, there were only mild reductions in mRNA expression of the α_1c-subunit (−15.5%, p = 0.13), and of the β₁-subunit isoforms a and c (−13.3%, p = 0.14 and −16.6%, p = 0.18, respectively). However, mRNA expression of the α₂/δ₁-subunit (−31.5%, p < 0.01) and of the β₁-subunit isoform b (−39.9%, p < 0.0005) was significantly reduced in patients with chronic AF. Taken together, the mRNA expression of the β₁-subunit isoforms b and c, which are splice variants, was significantly down-regulated by 26.5% (p < 0.05) in these patients. The analysis of the β_1c/β_1b ratio resulted in a significant shift by 39.2% (p < 0.0001) in favor of β_1c in patients with chronic atrial fibrillation. In the AF patients, the abundance of the 5-HT₄-receptor transcript was significantly reduced by 36% (p < 0.05). The β-adrenoreceptor transcription was unchanged. In both SR and AF patients, chronic β-blocker treatment did neither significantly effect the mRNA expression of the L-type Ca²⁺channel subunits, the β-adrenoreceptor subtypes 1 and 2, nor that of the 5-HT₄-receptor. Our data show that chronic AF is associated with a decrease in the atrial mRNA amount of auxiliary subunits of the L-type Ca²⁺-channel and of the 5-HT₄-receptor. This supports the hypothesis that the observed alterations in mRNA transcription in AF patients may lead to a decrease in the availability of functional L-type Ca²⁺-channels and 5-HT₄-receptors and/or reduce L-type Ca²⁺-current amplitude and density, thus, promoting and stabilizing the arrhythmia.