Aim: 

The extracellular Calcium Sensing Receptor (CaR) is a pleiotropic calcium sensor recently identified in the heart. Although scattered reports have suggested that CaR may have a role in the pathogenesis of cardiac hypertrophy and ischemia, the signalling pathways activated by this receptor still remain elusive and therefore represent the aim of this study.

Methods: 

Cytoplasmic [Ca²⁺] and [cAMP] were imaged in real-time in rat neonatal myocytes and fibroblasts loaded with fluo-4 (3 mM) or transiently transfected with an EPAC-based fluorescent probe (Ponsioen et al, 2004).

Results: 

Stimulation of fluo-4 loaded cardiac fibroblasts with spermine (1 mM), L-leucine and L-arginine (5 mM) and neomycin (250 mM) induced cytosolic [Ca²⁺] peaks that were respectively 75.9±9.7% (n=5, p<0.03), 45.1±6.3% (n=3, p<0.01), 39.4±5.8% (n=4, p<0.01;) and 46.2±5.4% (n=3; p<0.01) of control-ATP response. Parallel experiments performed on low-confluence spontaneously-beating cardiomyocytes showed that stimulation with spermine and NPS-R-467 (5 mM) reduced the frequency of Ca²⁺ oscillations by 41.3±13.6% (p<o.05; n=5) and 40.6±7.7% (p<0.05; n=3), respectively. The same effect was apparent on isoproterenol-induced increase of Ca²⁺ oscillation frequency. Direct [cAMP]i measurements showed that spermine was able to reduce isoproterenol-induced increase of [cAMP] by 31.3±2.0% (p<0.001; n=5).

Conclusions: 

The data collected strongly suggest that different physiological CaR-agonists activate PLC/IP₃ pathway in cardiac fibroblasts. On the contrary, the CaR agonist spermine decreased Ca²⁺ oscillation frequency in spontaneously beating cardiomyocytes, most probably via modulation of cAMP levels. Studies to ascertain the role of these cell-specific intracellular signalling pathways in cardiac hypertrophy are in progress.