Distribution and Functional Role of Inositol 1,4,5-trisphosphate Receptors in Mouse Sinoatrial Node

Rationale: Inositol 1,4,5-trisphosphate receptors (IP₃Rs) have been implicated in the generation of arrhythmias and cardiac muscle nuclear signaling. However, in the mammalian sinoatrial node (SAN), where the heart beat originates, the expression and functional activity of IP₃Rs have not been investigated.

Objectives: To determine whether SAN express IP₃Rs and which isoforms are present. To examine the response of the SAN to IP₃Rs analogues and agonists and the potential role played by IP₃Rs in cardiac pacemaking.

Methods and Results: The expression and distribution of IP₃Rs, were studied by reverse-transcription polymerase chain reaction, Western blotting, and immunolabeling. Ca²⁺ signaling and electric activity in intact mouse SAN were measured with Ca²⁺-sensitive fluorescent dyes. We found that although the entire SAN expressed three IP₃R mRNA isoforms, IP₃R2 was the predominant protein isoform detected by Western blot using the protein extracts from the SAN, AV node, and atrial tissue. Immunohistochemistry studies also showed that IP₃R2 was expressed in the central SAN region. Studies using isolated single pacemaker cells revealed that IP₃R2 (but not IP₃R1) was located with a similar distribution to the sarcoplasmic reticulum marker protein SERCA2a with some labeling adjacent to the surface membrane. The application of membrane-permeable IP₃ (IP₃-butyryloxymethyl ester) increased Ca²⁺ spark frequency and the pacemaker firing rate in single isolated pacemaker cells. In intact SAN preparations, endothelin-1–IP₃R agonist and IP₃-butyryloxymethyl ester both increased intracellular Ca²⁺ and the pacemaker firing rate, whereas the IP₃R antagonist, 2-aminoethoxydiphenyl borate decreased Ca²⁺ and the firing rate. Both of these effects were absent in the SAN from transgenic IP₃R2 knockout mice.

Conclusions: This study provides new evidence that functional IP₃R2s are expressed in the mouse SAN and could serve as an additional Ca²⁺-dependent mechanism in modulating cardiac pacemaker activity as well as other Ca²⁺-dependent processes.