@article{553f0ff9dcc641799c532444b9cb7403,
title = "Protocol to record and quantify the intracellular pH in contracting cardiomyocytes",
abstract = "Intracellular pH (pHi) plays critical roles in the regulation of cardiac function. Methods and techniques for cardiac pHi measurement have continued to evolve since early 1960s. Fluorescent microscopy is the most recently developed technique with several advantages over other techniques including higher spatial and temporal resolutions, and feasibility for contracting cell measurement. Here, we describe detailed methods for mouse cardiomyocyte isolation, and simultaneous measurement and quantification of pHi and sarcomere length in contracting cardiomyocytes. For complete details on the use and execution of this protocol, please refer to Lyu et al. (2022).",
keywords = "Cell isolation, Cell-based Assays, Metabolism, Microscopy, Single Cell",
author = "Yankun Lyu and Valeriy Timofeyev and James Overton and Thai, {Phung N.} and Yamoah, {Ebenezer N} and Nipavan Chiamvimonvat and Xiaodong Zhang",
note = "Funding Information: We thank our laboratory members for their constructive comments. We thank Dr. Lu Ren for her kind help in making the graphics. This study was supported by NIH R56 HL138392 to X.D.Z.; NIH R01 HL085727, HL085844, HL137228; VA Merit Review grants I01 BX000576 and CX001490 (N.C.); and American Heart Association Postdoctoral Fellowship Award, no. 906319 (Y.L.). Additional support was provided by NIH R01 DC015135, AG051443, DC015252, DC016099, AG060504 (E.N.Y.). N.C. is the holder of the Roger Tatarian Endowed Professorship in Cardiovascular Medicine and a part-time staff physician at VA Northern California Health Care System, Mather, CA. Institutional permissions: All animal care and procedures were performed in accordance with the protocols approved by the Institutional Animal Care and Use Committee of the University of California, Davis, and by National Institutes of Health guidelines. The experiments described in the protocol were conducted in a blinded fashion. Ventricular and atrial myocytes are isolated from wild-type (WT) male and female mice (10?16 weeks old). Except specifically indicated, all chemicals are purchased from Sigma-Aldrich (St. Louis, MO). X.D.Z. proposed and developed the concept, designed the study, established the method, initially tested the concept by experiments, analyzed the data, made figures, and wrote the manuscript. N.C. helped on experiment design, data analysis and interpretation, and revised the manuscript; Y.L. performed the intracellular pH and contraction measurement, data analysis and organization, and revised the manuscript; V.T. isolated the mouse cardiomyocytes, and contributed to the cardiomyocyte isolation protocol writing; J.O. helped on cardiomyocyte isolations; P.N.T. helped on cardiomyocyte isolations; E.N.Y. revised the manuscript and helped on the data acquisition and analysis. The authors declare no competing interests. Funding Information: We thank our laboratory members for their constructive comments. We thank Dr. Lu Ren for her kind help in making the graphics. This study was supported by NIH R56 HL138392 to X.D.Z.; NIH R01 HL085727 , HL085844 , HL137228; VA Merit Review grants I01 BX000576 and CX001490 (N.C.); and American Heart Association Postdoctoral Fellowship Award, no. 906319 (Y.L.). Additional support was provided by NIH R01 DC015135 , AG051443 , DC015252 , DC016099 , AG060504 (E.N.Y.). N.C. is the holder of the Roger Tatarian Endowed Professorship in Cardiovascular Medicine and a part-time staff physician at VA Northern California Health Care System, Mather, CA. Institutional permissions: All animal care and procedures were performed in accordance with the protocols approved by the Institutional Animal Care and Use Committee of the University of California, Davis, and by National Institutes of Health guidelines. The experiments described in the protocol were conducted in a blinded fashion. Ventricular and atrial myocytes are isolated from wild-type (WT) male and female mice (10–16 weeks old). Except specifically indicated, all chemicals are purchased from Sigma-Aldrich (St. Louis, MO). Publisher Copyright: {\textcopyright} 2022 The Author(s)",
year = "2022",
month = jun,
day = "17",
doi = "10.1016/j.xpro.2022.101301",
language = "English (US)",
volume = "3",
journal = "STAR Protocols",
issn = "2666-1667",
publisher = "Cell Press",
number = "2",
}