Impulse conduction and gap junctional remodelling by endothelin-1 in cultured neonatal rat ventricular myocytes |
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Authors: | Y. Reisner G. Meiry N. Zeevi-Levin D. Y. Barac I. Reiter Z. Abassi N. Ziv S. Kostin J. Schaper M. R. Rosen O. Binah |
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Affiliation: | Rappaport Family Institute for Research in the Medical Sciences, Ruth and Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel;Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany;The Departments of Pharmacology and Pediatrics and Center for Molecular Therapeutics, College of Physicians &Surgeons of Columbia University, New York, NY, USA |
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Abstract: | Endothelin-1 (ET-1) is an important contributor to ventricular hypertrophy and failure, which are associated with arrhythmogenesis and sudden death. To elucidate the mechanism(s) underlying the arrhythmogenic effects of ET-1 we tested the hypothesis that long-term (24 hrs) exposure to ET-1 impairs impulse conduction in cultures of neonatal rat ventricular myocytes (NRVM). NRVM were seeded on micro-electrode-arrays (MEAs, Multi Channel Systems, Reutlingen, Germany) and exposed to 50 nM ET-1 for 24 hrs. Hypertrophy was assessed by morphological and molecular methods. Consecutive recordings of paced activation times from the same cultures were conducted at baseline and after 3, 6 and 24 hrs, and activation maps for each time period constructed. Gap junctional Cx43 expression was assessed using Western blot and confocal microscopy of immunofluorescence staining using anti-Cx43 antibodies. ET-1 caused hypertrophy as indicated by a 70% increase in mRNA for atrial natriuretic peptide ( P < 0.05), and increased cell areas ( P < 0.05) compared to control. ET-1 also caused a time-dependent decrease in conduction velocity that was evident after 3 hrs of exposure to ET-1, and was augmented at 24 hrs, compared to controls ( P < 0.01). ET-1 increased total Cx43 protein by ∼40% ( P < 0.05) without affecting non- phosphorylated Cx43 (NP-Cx43) protein expression. Quantitative confocal microscopy showed a ∼30% decrease in the Cx43 immunofluorescence per field in the ET-1 group ( P < 0.05) and a reduced field stain intensity ( P < 0.05), compared to controls. ET-1-induced hypertrophy was accompanied by reduction in conduction velocity and gap junctional remodelling. The reduction in conduction velocity may play a role in ET-1 induced susceptibility to arrhythmogenesis. |
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Keywords: | Endothelin-1 connexin 43 conduction velocity hypertrophy neonatal rat ventricular myocytes |
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