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Ageing related change in store-operated Ca2+ influx and TRPCs expression of sinoatrial node

Y.K. Ju, D. Lai and D.G. Allen, School of Medical Sciences, Bosch institute, University of Sydney, NSW 2006, Australia.

The sinoatrial node (SAN) is the primary pacemaker of the mammalian heart. Dysfunction of the SAN increases exponentially with age. The age-related remodelling of pacemaker currents and pacemaker action potential may contribute to age-related slowing of the heart rate and reduction in conduction velocity which result in arrhythmias (Kistler et al., 2004). We recently discovered that store-operated Ca2+ channel (SOCC) activity is present in the mouse sinoatrial node leading to Ca2+ entry (Ju et al., 2007). In the present study we tested the hypothesis that changes in SOCC activity might play a significant role in ageing related SAN dysfunction.

The experiments were carried out in intact mouse SAN preparations from either adult (2 months) or old (> 18 months) mice. Intracellular Ca2+ was detected with the fluorescent Ca2+ indicator indo-1. The SOCC activity was determined from the influx of Ca2+ following depletion of the sarcoplasmic reticulum Ca2+ store. We found that the Ca2+ influx was reduced in old mice compare to adult mice (2 month) (p < 0.05, n = 4).

It is widely believed that the transient receptor potential cannonical (TRPC) gene family are candidates for encoding SOCC. Recently we have developed one-step quantitative PCR to study gene expression in mouse SAN. This method allows us to study expression levels of up to 8 genes from one mouse SAN. Central or peripheral SAN samples were isolated from mouse hearts and stored in RNAlater solution (Ambion Inc.). One-step real-time PCRs were performed using the Cells Direct kit (Invitrogen, Australia). We tested 4 housekeeping genes including 18s rRNA, β-actin, HPRT1, B2m as reference genes, to normalise the changes mRNA expression of TRPCs or other genes of interest. We also used TRPC1 DNA as sample input reference. HCN4 used as a positive control for the central SAN region. We quantified the mRNA expression of TRPCs (TRPC1, 3, and 4) and stromal interacting molecule 1 (STIM1), an endoplasmic reticulum-Ca2+ sensor protein thought to associate with TRPCs. We found a reduction of the TRPC3 mRNA expression in the SANs from old mice (20-24 months) compared to young mice (1 month). Our preliminary data suggests that changes in SOCC activity and expression of TRPC genes might play a role in SAN dysfunction, specifically in the ageing heart.

Ju YK, Chu Y, Chaulet H, Lai D, Gervasio OL, Graham RM, Cannell MB, Allen DG. (2007) Circulation Research, 100: 1605-14.

Kistler PM, Sanders P, Fynn SP, Stevenson IH, Spence SJ, Vohra JK, Sparks PB, Kalman JM. (2004) Journal of American College of Cardiology, 44:109-16.

This work was supported by NH&MRC, Australia.