Department of Neuroscience, Physiology & Pharmacology, University College London, London, UK.
Department of Biochemistry, University of Cambridge, Cambridge, UK.
Pflugers Arch. 2023 May;475(5):583-593. doi: 10.1007/s00424-023-02798-9. Epub 2023 Mar 14.
Atrial fibrillation (AF) occurs from disordered atrial action potential conduction and is associated with reduced gap junction electrical conductance (G). The Ca and calmodulin-dependent phosphatase, calcineurin, reduces G in ventricular myocardium via a protein phosphatase-1 (PP1)-dependent pathway culminating in phosphorylation of serine368 on connexin43 (pSer368-Cx43). However, characterisation of corresponding pathways in left atrial myocardium, which have a more complex connexin subtype profile, is undefined and was the aim of this study. G was measured in guinea-pig left atrium from the frequency-dependent variation of intracellular impedance; intracellular [Ca], ([Ca]) in low-Na solution was measured by Fura-2 fluorescence. Phosphorylation of guinea-pig Ser368-Cx43 residues was measured by Western blot; Cx40 was immunoprecipitated and probed for serine/threonine residue phosphorylation. Low-Na solution reversibly reduced G, in turn attenuated or prevented by calcineurin inhibitors cyclosporin-A or CAIP, respectively. Moreover, Ser368-Cx43 phosphorylation in low-Na solution was also prevented by CAIP. Changes were partially prevented by fostreicin (FST), a protein phosphatase-2A (PP2A) inhibitor; but not by tautomycin, a PP1 inhibitor. Serine/threonine residues on Cx40 were also phosphorylated in low-Na solution; prevented by CAIP and attenuated by FST. Reduced G with raised [Ca] is paralleled by a changed Cx43/Cx40 phosphorylation status; changes mediated by calcineurin and PP2A-dependent pathways, but not PP1. The pharmacological profile underlying changes to guinea-pig atrial gap junction electrical conductance with raised intracellular [Ca] is fundamentally different from that in ventricular myocardium. This provides a targeted drug model whereby atrial and ventricular myocardium can be selectively targeted to correct conduction defects.
心房颤动（AF）是由于心房动作电位传导紊乱引起的，与间隙连接电导率（G）降低有关。钙和钙调蛋白依赖性磷酸酶钙调神经磷酸酶通过蛋白磷酸酶-1（PP1）依赖性途径降低心室心肌中的 G，最终导致连接蛋白 43（Cx43）上丝氨酸 368 的磷酸化（pSer368-Cx43）。然而，左心房心肌中对应途径的特征尚不明确，而左心房心肌具有更复杂的连接蛋白亚型谱，这也是本研究的目的。通过测量细胞内阻抗的频率依赖性变化来测量豚鼠左心房的 G；通过 Fura-2 荧光测量低钠溶液中的细胞内[Ca]（[Ca]）。通过 Western blot 测量豚鼠 Ser368-Cx43 残基的磷酸化；免疫沉淀 Cx40 并探测丝氨酸/苏氨酸残基磷酸化。低钠溶液可逆性降低 G，分别被钙调神经磷酸酶抑制剂环孢素 A 或 CAIP 减弱或阻止。此外，低钠溶液中的 Ser368-Cx43 磷酸化也被 CAIP 阻止。变化部分被蛋白磷酸酶-2A（PP2A）抑制剂 fostreicin（FST）阻止；但不被蛋白磷酸酶-1（PP1）抑制剂 tautomycin 阻止。低钠溶液中的 Cx40 丝氨酸/苏氨酸残基也发生磷酸化；被 CAIP 阻止并被 FST 减弱。升高细胞内[Ca]时 G 降低伴随着 Cx43/Cx40 磷酸化状态的改变；变化由钙调神经磷酸酶和 PP2A 依赖性途径介导，但不被 PP1 介导。升高细胞内[Ca]时豚鼠心房缝隙连接电导率变化的药理学特征与心室心肌根本不同。这提供了一种有针对性的药物模型，可选择性靶向心房和心室心肌以纠正传导缺陷。
