The effect of polyunsaturated fatty acids on cardiac ryanodine and inositol triphosphate receptors

B. Honen¹, D. Saint² and D.R. Laver¹, ¹Department of Human Physiology, School of Biomedical Science, The University of Newcastle, Callaghan, NSW 2308 and ²Department of Physiology, School of Molecular and Biomedical sciences, The University of Adelaide, Adelaide, SA 5005, Australia.

It is well recognised that the consumption of fish correlates with a reduction in mortality due to cardiovascular disease (Burr et al., 1989). Whole heart studies have identified that dietary fish oil confers protection from cardiac arrhythmias (McLennan, 1993). Many studies have shown that the acute application of the polyunsaturated fatty acids present in fish oil, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) to cardiac myocytes significantly reduce the amplitude of the various sarcolemmal ion currents responsible for the cardiac action potential (Xiao et al., 1995; Xiao et al., 1997; Bogdanov et al., 1998). It is believed that this reduction in electrical excitability is the mechanism by which fish oil confers protection from cardiac arrhythmias. However some arrhythmias also arise from abnormal calcium handling by internal stores. Thus it has been suggested that the anti-arrhythmic effects of long chain polyunsaturated fatty acids (PUFAs) may be related to their ability to alter calcium handling in cardiac myocytes (Honen & Saint 2002, O'Neill et al., 2002). Therefore we investigated the effects of EPA and DHA on the kinetics of the cardiac calcium release channels (i.e. the ryanodine receptor (RyR) and the inositol triphosphate receptor (IP₃R)).

RyRs and IP₃R isolated from sheep hearts and were incorporated into artificial bilayers formed from a solution of phosphatidylethanolamine and phosphatidylcholine dissolved in either n-decane or n-tetradecane using standard techniques (O'Neill et al., 2003). Cytoplasmic solutions contained 250 mM Cs⁺ (230 mM CsCH₃O₃S, 20 mM CsCl), 10 mM TES at pH 7.4. Luminal solution contained 50 mM Cs⁺ (30 mM CsCH₃O₃S, 20 mM CsCl), 10 mM TES and 1 mM CaCl₂, pH 7.4.

Concentrations of EPA ranging between 10 and 50 μM, when applied to either the cytosolic or luminal side of the RyR, produced a dose dependent inhibition of RyR open probability with K_I =32 μM and Hill coefficient, n_I = 3.8. This inhibition typically occurred within 30 seconds of application. Inhibition was independent of the n-alkane solvent and whether RyRs were activated by ATP or Ca²⁺. Like EPA, the cytosolic application of 50 μM DHA also resulted in a reduction in channel open probability.

Like with RyR, the open probability of the IP₃R fell upon the application of 50 μM EPA. IP₃Rs were identified by their activation by IP₃ and inhibition by 10 μM heparin, a reversible IP₃R blocker.

The results suggest that both EPA and DHA affect calcium handling by directly inhibiting RyRs at micromolar concentrations. The actions of both EPA and DHA may be mediated via the membrane or by binding to a hydrophobic site on the channel itself. This provides a potential avenue by which PUFAs confer protection from cardiac arrhythmias.

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