Store-Operated Ca ²⁺ Influx and Expression of TRPC Genes in Mouse Sinoatrial Node

Abstract

Abstract-Store-operated Ca 2ϩ entry was investigated in isolated mouse sinoatrial nodes (SAN) dissected from right atria and loaded with Ca 2ϩ indicators. Incubation of the SAN in Ca 2ϩ -free solution caused a substantial decrease in resting intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) and stopped pacemaker activity. Reintroduction of Ca 2ϩ in the presence of cyclopiazonic acid (CPA), a sarcoplasmic reticulum Ca 2ϩ pump inhibitor, led to sustained elevation of [Ca 2ϩ ] i , a characteristic of store-operated Ca 2ϩ channel (SOCC) activity. Two SOCC antagonists, Gd 3ϩ and SKF-96365, inhibited 72Ϯ8% and 65Ϯ8% of this Ca 2ϩ influx, respectively. SKF-96365 also reduced the spontaneous pacemaker rate to 27Ϯ4% of control in the presence of CPA. Because members of the transient receptor potential canonical (TRPC) gene family may encode SOCCs, we used RT-PCR to examine mRNA expression of the 7 known mammalian TRPC isoforms. Transcripts for TRPC1, 2, 3, 4, 6, and 7, but not TRPC5, were detected. Immunohistochemistry using anti-TRPC1, 3, 4, and 6 antibodies revealed positive labeling in the SAN region and single pacemaker cells. 4,5 In ventricular myocytes, PLC responses are generally modest with only small amounts of IP 3 being produced. 6 Thus release of Ca 2ϩ through IP 3 R is probably too small to modify excitation-contraction coupling. However, atria myocytes express functional IP 3 R at 6 to 10 times higher levels than those in ventricules, and Ca 2ϩ release from IP 3 R may be involved in the generation of arrhythmias. 7 Furthermore, recent studies have shown that activation of PLC in the heart leads to Ca 2ϩ release from perinuclear IP 3 R and thereby regulates nuclear-cytoplasmic cycling of transcription factors and alters gene expression. 8 A similar role may be played by IP 3 and the IP 3 R in skeletal muscle and SOCCs have been implicated in IGF-1 induced muscle hypertrophy. 5,9 The identity of the genes that encode SOCCs remains uncertain. Studies of the transient receptor potential (TRP) gene from Drosophila showed that it encodes a PLC-activated Ca 2ϩ permeable channel. 10 Subsequently, 7 TRP channel homologues in mammals, termed TRPC1-TRPC7, have been identified and there is considerable evidence to indicate that TRPC1 can encode a SOCC. 11 In addition, a recent study showed that overexpression of TRPC3 substantially enhanced SOCC activity in the heart. 5 The importance of Ca 2ϩ release from stores in cardiac pacemaking is now widely accepted. [12][13][14] In a previous study, we found that activation of the P2Y 1 purinergic receptor by ATP results in modulation of pacemaker firing attirbutable to receptor-coupled PLC activation and depletion of SR Ca 2ϩ stores. 15 Because activation of SOCCs also involves PLC, we speculated that SOCCs might be present in pacemaker tissue.In mammals, the sinoatrial node (SAN) is a heterogeneous tissue. The shape of the action potential and the rate of rise of the pacemaker potential change progressively from the periphery to the center, the latter being the leadin...

STIM1–Ca 2+ signaling modulates automaticity of the mouse sinoatrial node

“…SOCE is therefore an attractive candidate to link the Ca 2+ and membrane clocks that underlie SANC automaticity. Roles for STIM1-dependent SOCE have been suggested in the heart (10)(11)(12)(13)(14). Here, we show that STIM1 is selectively expressed in the SAN where it activates Ca 2+ entry via Orai1 channels and thereby modulates the Ca 2+ signals required for pacemaking activity of the SAN.…”

Ca2+-stimulated Basal Adenylyl Cyclase Activity Localization in Membrane Lipid Microdomains of Cardiac Sinoatrial Nodal Pacemaker Cells

“…Ca 2ϩ -activated AC types have also been localized to caveolin microdomains containing store-operated Ca 2ϩ -operated Ca 2ϩ channels (32). Although there is presently no evidence for functional store-operated Ca 2ϩ channels in rabbit SANC, recent evidence suggests that these channels are present in mouse SAN tissue (33).…”

Ageing-dependent remodelling of ion channel and Ca2+clock genes underlying sino-atrial node pacemaking

“…The TRPCs have been suggested to be Ca 2+ store (i.e. SR)-operated Ca 2+ channels and to be involved in SAN pacemaking (Ju et al 2007). Interestingly, TRPC1 was less abundant and TRPC6 more abundant in the SAN.…”