Cargando…

Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes

Experiments in isolated ventricular cardiomyocytes have greatly facilitated the study of cellular and subcellular physiology in the heart. However, the isolation and culture of high‐quality adult murine ventricular cardiomyocytes can be technically challenging. In most experimental protocols, the cu...

Descripción completa

Detalles Bibliográficos
Autores principales: Hall, Andrew R., Hausenloy, Derek J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4760411/
https://www.ncbi.nlm.nih.gov/pubmed/26733241
http://dx.doi.org/10.14814/phy2.12606
_version_ 1782416871948025856
author Hall, Andrew R.
Hausenloy, Derek J.
author_facet Hall, Andrew R.
Hausenloy, Derek J.
author_sort Hall, Andrew R.
collection PubMed
description Experiments in isolated ventricular cardiomyocytes have greatly facilitated the study of cellular and subcellular physiology in the heart. However, the isolation and culture of high‐quality adult murine ventricular cardiomyocytes can be technically challenging. In most experimental protocols, the culture of viable adult murine cardiomyocytes for prolonged time periods is achieved with the addition of the myosin II ATPase inhibitors blebbistatin and/or 2,3‐butanedione monoxime (BDM). These drugs are added to increase cell viability and life span by inhibiting spontaneous cardiomyocyte contraction, thereby preventing calcium overload, cell hypercontracture, and cell death. While the addition of BDM has been reported to prolong the life span of isolated adult murine cardiomyocytes, it is also associated with several off‐target effects. Here, we report a novel off‐target effect, in which BDM inhibits mitochondrial respiration by acting directly on the electron transport chain to reduce cell viability. In contrast, when cells were cultured with blebbistatin alone, cells survived for longer, and no metabolic off‐target effects were observed. Based on these novel observations, we recommend that culture media for isolated mouse ventricular cardiomyocytes should be supplemented with blebbistatin alone, as BDM has the potential to affect mitochondrial respiration and cell viability, effects which may impact adversely on subsequent experiments.
format Online
Article
Text
id pubmed-4760411
institution National Center for Biotechnology Information
language English
publishDate 2016
publisher John Wiley and Sons Inc.
record_format MEDLINE/PubMed
spelling pubmed-47604112016-02-22 Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes Hall, Andrew R. Hausenloy, Derek J. Physiol Rep Original Research Experiments in isolated ventricular cardiomyocytes have greatly facilitated the study of cellular and subcellular physiology in the heart. However, the isolation and culture of high‐quality adult murine ventricular cardiomyocytes can be technically challenging. In most experimental protocols, the culture of viable adult murine cardiomyocytes for prolonged time periods is achieved with the addition of the myosin II ATPase inhibitors blebbistatin and/or 2,3‐butanedione monoxime (BDM). These drugs are added to increase cell viability and life span by inhibiting spontaneous cardiomyocyte contraction, thereby preventing calcium overload, cell hypercontracture, and cell death. While the addition of BDM has been reported to prolong the life span of isolated adult murine cardiomyocytes, it is also associated with several off‐target effects. Here, we report a novel off‐target effect, in which BDM inhibits mitochondrial respiration by acting directly on the electron transport chain to reduce cell viability. In contrast, when cells were cultured with blebbistatin alone, cells survived for longer, and no metabolic off‐target effects were observed. Based on these novel observations, we recommend that culture media for isolated mouse ventricular cardiomyocytes should be supplemented with blebbistatin alone, as BDM has the potential to affect mitochondrial respiration and cell viability, effects which may impact adversely on subsequent experiments. John Wiley and Sons Inc. 2016-01-05 /pmc/articles/PMC4760411/ /pubmed/26733241 http://dx.doi.org/10.14814/phy2.12606 Text en © 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Original Research
Hall, Andrew R.
Hausenloy, Derek J.
Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title_full Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title_fullStr Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title_full_unstemmed Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title_short Mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (BDM): implications for culturing isolated mouse ventricular cardiomyocytes
title_sort mitochondrial respiratory inhibition by 2,3‐butanedione monoxime (bdm): implications for culturing isolated mouse ventricular cardiomyocytes
topic Original Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4760411/
https://www.ncbi.nlm.nih.gov/pubmed/26733241
http://dx.doi.org/10.14814/phy2.12606
work_keys_str_mv AT hallandrewr mitochondrialrespiratoryinhibitionby23butanedionemonoximebdmimplicationsforculturingisolatedmouseventricularcardiomyocytes
AT hausenloyderekj mitochondrialrespiratoryinhibitionby23butanedionemonoximebdmimplicationsforculturingisolatedmouseventricularcardiomyocytes