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Attenuation of acetylcholine activated potassium current (I KACh) by simvastatin, not pravastatin in mouse atrial cardiomyocyte: possible atrial fibrillation preventing effects of statin.

Cho KI, Cha TJ, Lee SJ, Shim IK, Zhang YH, Heo JH, Kim HS, Kim SJ, Kim KL, Lee JW - PLoS ONE (2014)

Bottom Line: Supplementation of substrates for the synthesis of cholesterol (mevalonate, geranylgeranyl pyrophosphate or farnesyl pyrophosphate) did not reverse the effect of simvastatin on IKACh, suggesting a cholesterol-independent effect on IKACh.Furthermore, supplementation of phosphatidylinositol 4,5-bisphosphate, extracellular perfusion of phospholipase C inhibitor or a protein kinase C (PKC) inhibitor had no effect on the inhibitory activity of simvastatin on IKACh.Simvastatin also inhibits adenosine activated IKACh, however, simvastatin does not inhibit IKACh after activated by intracellular loading of GTP gamma S.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Institute, Department of Internal Medicine, Kosin University College of Medicine, Busan, South Korea.

ABSTRACT
Statins, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors, are associated with the prevention of atrial fibrillation (AF) by pleiotropic effects. Recent clinical trial studies have demonstrated conflicting results on anti-arrhythmia between lipophilic and hydrophilic statins. However, the underlying mechanisms responsible for anti-arrhythmogenic effects of statins are largely unexplored. In this study, we evaluated the different roles of lipophilic and hydrophilic statins (simvastatin and pravastatin, respectively) in acetylcholine (100 µM)-activated K+ current (IKACh, recorded by nystatin-perforated whole cell patch clamp technique) which are important for AF initiation and maintenance in mouse atrial cardiomyocytes. Our results showed that simvastatin (1-10 µM) inhibited both peak and quasi-steady-state IKACh in a dose-dependent manner. In contrast, pravastatin (10 µM) had no effect on IKACh. Supplementation of substrates for the synthesis of cholesterol (mevalonate, geranylgeranyl pyrophosphate or farnesyl pyrophosphate) did not reverse the effect of simvastatin on IKACh, suggesting a cholesterol-independent effect on IKACh. Furthermore, supplementation of phosphatidylinositol 4,5-bisphosphate, extracellular perfusion of phospholipase C inhibitor or a protein kinase C (PKC) inhibitor had no effect on the inhibitory activity of simvastatin on IKACh. Simvastatin also inhibits adenosine activated IKACh, however, simvastatin does not inhibit IKACh after activated by intracellular loading of GTP gamma S. Importantly, shortening of the action potential duration by acetylcholine was restored by simvastatin but not by pravastatin. Together, these findings demonstrate that lipophilic statins but not hydrophilic statins attenuate IKACh in atrial cardiomyocytes via a mechanism that is independent of cholesterol synthesis or PKC pathway, but may be via the blockade of acetylcholine binding site. Our results may provide important background information for the use of statins in patients with AF.

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Acetylcholine-activated K+ current (IKACh) after simvastatin with A. phosphatidylinositol 4,5-bisphosphate (PIP2), B. phospholipase C inhibitor (PLC inhibitor, neomycin 50 µM), and C. protein kinase C inhibitor (PKC inhibitor, calphostin C 1 µM).D. Peak amplitudes of baseline IKACh (I1, peak) and the second IKACh (I2, peak) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. E. Quasi-steady state amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. F. Simvastatin did not inhibit the activated IKACh by intracellular loading of GTP gamma S (100 µM/L). G. Simvastatin inhibit activated IKACh by adenosine. H. Peak amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) activated by GTP gamma S and adenosine after treatment with simvastatin. NS; no significant change, *; p<0.05 compared to controls.
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pone-0106570-g005: Acetylcholine-activated K+ current (IKACh) after simvastatin with A. phosphatidylinositol 4,5-bisphosphate (PIP2), B. phospholipase C inhibitor (PLC inhibitor, neomycin 50 µM), and C. protein kinase C inhibitor (PKC inhibitor, calphostin C 1 µM).D. Peak amplitudes of baseline IKACh (I1, peak) and the second IKACh (I2, peak) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. E. Quasi-steady state amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. F. Simvastatin did not inhibit the activated IKACh by intracellular loading of GTP gamma S (100 µM/L). G. Simvastatin inhibit activated IKACh by adenosine. H. Peak amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) activated by GTP gamma S and adenosine after treatment with simvastatin. NS; no significant change, *; p<0.05 compared to controls.

Mentions: To investigate the association between simvastatin-induced IKACh inhibition and inhibition of cholesterol synthesis, substrates for cholesterol synthesis consisting of mevalonate (MVA, Fig. 4A), geranylgeranyl pyrophosphate (GGPP, Fig. 4B), or farnesyl pyrophosphate (FPP, Fig. 4C) were added with simvastatin in the bath solution. However, the reductions in peak amplitude and quasi-steady-state current of IKACh by simvastatin were not prevented by supplementation with any of these substrates (p = 0.28, p = 0.37 and p = 0.41 for MVA, GGPP and FPP, respectively, each n = 7, Figs. 4D,E). Moreover, to investigate if the modulation of simvastatin-induced IKACh inhibition may happen through the phospholipase C (PLC), protein kinase C (PKC) pathway or depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) [19], [20], PLC inhibitor, PKC inhibitor, and PIP2 were tested. Loading the patch pipette with PIP2 via whole cell ruptured patch clamp did not alter simvastatin-mediated inhibition of IKACh (Fig. 5A), implying that simvastatin did not limit the availability of these agents. Similarly, application of the PLC inhibitor neomycin (50 µM, Fig. 5B) or the PKC inhibitor calphostin C extracellular solution (1 µM, Fig. 5C) failed to alter simvastatin-inhibition of IKACh (each n = 7, Figs. 5D,E). When we activate IKACh by intracellular loading of GTP gamma S (100 µM/L) via whole cell patch, simvastatin did not inhibit IKACh (n = 5, Figs. 5F,H). However, when we activate IKACh by extracellular application of adenosine, simvastatin also inhibit adenosine activated IKACh (n = 5, Figs. 5G,H), which suggest that simvastatin influence on the adenosine binding site as well as acetylcholine binding sites. This result suggests that acute administration of simvastatin may inhibit the IKACh by blockade of acetylcholine binding site.


Attenuation of acetylcholine activated potassium current (I KACh) by simvastatin, not pravastatin in mouse atrial cardiomyocyte: possible atrial fibrillation preventing effects of statin.

Cho KI, Cha TJ, Lee SJ, Shim IK, Zhang YH, Heo JH, Kim HS, Kim SJ, Kim KL, Lee JW - PLoS ONE (2014)

Acetylcholine-activated K+ current (IKACh) after simvastatin with A. phosphatidylinositol 4,5-bisphosphate (PIP2), B. phospholipase C inhibitor (PLC inhibitor, neomycin 50 µM), and C. protein kinase C inhibitor (PKC inhibitor, calphostin C 1 µM).D. Peak amplitudes of baseline IKACh (I1, peak) and the second IKACh (I2, peak) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. E. Quasi-steady state amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. F. Simvastatin did not inhibit the activated IKACh by intracellular loading of GTP gamma S (100 µM/L). G. Simvastatin inhibit activated IKACh by adenosine. H. Peak amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) activated by GTP gamma S and adenosine after treatment with simvastatin. NS; no significant change, *; p<0.05 compared to controls.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4199526&req=5

pone-0106570-g005: Acetylcholine-activated K+ current (IKACh) after simvastatin with A. phosphatidylinositol 4,5-bisphosphate (PIP2), B. phospholipase C inhibitor (PLC inhibitor, neomycin 50 µM), and C. protein kinase C inhibitor (PKC inhibitor, calphostin C 1 µM).D. Peak amplitudes of baseline IKACh (I1, peak) and the second IKACh (I2, peak) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. E. Quasi-steady state amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) after treatment with PIP2, PLC inhibitor, and PKC inhibitor. F. Simvastatin did not inhibit the activated IKACh by intracellular loading of GTP gamma S (100 µM/L). G. Simvastatin inhibit activated IKACh by adenosine. H. Peak amplitudes of baseline IKACh (qss I1) and second IKACh (qss I2) activated by GTP gamma S and adenosine after treatment with simvastatin. NS; no significant change, *; p<0.05 compared to controls.
Mentions: To investigate the association between simvastatin-induced IKACh inhibition and inhibition of cholesterol synthesis, substrates for cholesterol synthesis consisting of mevalonate (MVA, Fig. 4A), geranylgeranyl pyrophosphate (GGPP, Fig. 4B), or farnesyl pyrophosphate (FPP, Fig. 4C) were added with simvastatin in the bath solution. However, the reductions in peak amplitude and quasi-steady-state current of IKACh by simvastatin were not prevented by supplementation with any of these substrates (p = 0.28, p = 0.37 and p = 0.41 for MVA, GGPP and FPP, respectively, each n = 7, Figs. 4D,E). Moreover, to investigate if the modulation of simvastatin-induced IKACh inhibition may happen through the phospholipase C (PLC), protein kinase C (PKC) pathway or depletion of phosphatidylinositol 4,5-bisphosphate (PIP2) [19], [20], PLC inhibitor, PKC inhibitor, and PIP2 were tested. Loading the patch pipette with PIP2 via whole cell ruptured patch clamp did not alter simvastatin-mediated inhibition of IKACh (Fig. 5A), implying that simvastatin did not limit the availability of these agents. Similarly, application of the PLC inhibitor neomycin (50 µM, Fig. 5B) or the PKC inhibitor calphostin C extracellular solution (1 µM, Fig. 5C) failed to alter simvastatin-inhibition of IKACh (each n = 7, Figs. 5D,E). When we activate IKACh by intracellular loading of GTP gamma S (100 µM/L) via whole cell patch, simvastatin did not inhibit IKACh (n = 5, Figs. 5F,H). However, when we activate IKACh by extracellular application of adenosine, simvastatin also inhibit adenosine activated IKACh (n = 5, Figs. 5G,H), which suggest that simvastatin influence on the adenosine binding site as well as acetylcholine binding sites. This result suggests that acute administration of simvastatin may inhibit the IKACh by blockade of acetylcholine binding site.

Bottom Line: Supplementation of substrates for the synthesis of cholesterol (mevalonate, geranylgeranyl pyrophosphate or farnesyl pyrophosphate) did not reverse the effect of simvastatin on IKACh, suggesting a cholesterol-independent effect on IKACh.Furthermore, supplementation of phosphatidylinositol 4,5-bisphosphate, extracellular perfusion of phospholipase C inhibitor or a protein kinase C (PKC) inhibitor had no effect on the inhibitory activity of simvastatin on IKACh.Simvastatin also inhibits adenosine activated IKACh, however, simvastatin does not inhibit IKACh after activated by intracellular loading of GTP gamma S.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Institute, Department of Internal Medicine, Kosin University College of Medicine, Busan, South Korea.

ABSTRACT
Statins, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors, are associated with the prevention of atrial fibrillation (AF) by pleiotropic effects. Recent clinical trial studies have demonstrated conflicting results on anti-arrhythmia between lipophilic and hydrophilic statins. However, the underlying mechanisms responsible for anti-arrhythmogenic effects of statins are largely unexplored. In this study, we evaluated the different roles of lipophilic and hydrophilic statins (simvastatin and pravastatin, respectively) in acetylcholine (100 µM)-activated K+ current (IKACh, recorded by nystatin-perforated whole cell patch clamp technique) which are important for AF initiation and maintenance in mouse atrial cardiomyocytes. Our results showed that simvastatin (1-10 µM) inhibited both peak and quasi-steady-state IKACh in a dose-dependent manner. In contrast, pravastatin (10 µM) had no effect on IKACh. Supplementation of substrates for the synthesis of cholesterol (mevalonate, geranylgeranyl pyrophosphate or farnesyl pyrophosphate) did not reverse the effect of simvastatin on IKACh, suggesting a cholesterol-independent effect on IKACh. Furthermore, supplementation of phosphatidylinositol 4,5-bisphosphate, extracellular perfusion of phospholipase C inhibitor or a protein kinase C (PKC) inhibitor had no effect on the inhibitory activity of simvastatin on IKACh. Simvastatin also inhibits adenosine activated IKACh, however, simvastatin does not inhibit IKACh after activated by intracellular loading of GTP gamma S. Importantly, shortening of the action potential duration by acetylcholine was restored by simvastatin but not by pravastatin. Together, these findings demonstrate that lipophilic statins but not hydrophilic statins attenuate IKACh in atrial cardiomyocytes via a mechanism that is independent of cholesterol synthesis or PKC pathway, but may be via the blockade of acetylcholine binding site. Our results may provide important background information for the use of statins in patients with AF.

Show MeSH
Related in: MedlinePlus