Journal article
Illuminating cell signaling with genetically encoded FRET biosensors in adult mouse cardiomyocytes
The Journal of general physiology, Vol.150(11), pp.1567-1582
11/01/2018
DOI: 10.1085/jgp.201812119
PMCID: PMC6219686
PMID: 30242036
Abstract
FRET-based biosensor experiments in adult cardiomyocytes are a powerful way of dissecting the spatiotemporal dynamics of the complicated signaling networks that regulate cardiac health and disease. However, although much information has been gleaned from FRET studies on cardiomyocytes from larger species, experiments on adult cardiomyocytes from mice have been difficult at best. Thus the large variety of genetic mouse models cannot be easily used for this type of study. Here we develop cell culture conditions for adult mouse cardiomyocytes that permit robust expression of adenoviral FRET biosensors and reproducible FRET experimentation. We find that addition of 6.25 mu M blebbistatin or 20 mu M (S)-nitro-blebbistatin to a minimal essential medium containing 10 mM HEPES and 0.2% BSA maintains morphology of cardiomyocytes from physiological, pathological, and transgenic mouse models for up to 50 h after adenoviral infection. This provides a 10-15-h time window to perform reproducible FRET readings using a variety of CFP/YFP sensors between 30 and 50 h postinfection. The culture is applicable to cardiomyocytes isolated from transgenic mouse models as well as models with cardiac diseases. Therefore, this study helps scientists to disentangle complicated signaling networks important in health and disease of cardiomyocytes.
Details
- Title: Subtitle
- Illuminating cell signaling with genetically encoded FRET biosensors in adult mouse cardiomyocytes
- Creators
- Gopireddy Raghavender Reddy - University of California, DavisToni M. West - University of California, DavisZhong Jian - University of California, DavisMark Jaradeh - University of California, DavisQian Shi - University of California, DavisYing Wang - University of California, DavisYe Chen-Izu - University of California, DavisYang K. Xiang - University of California, Davis
- Resource Type
- Journal article
- Publication Details
- The Journal of general physiology, Vol.150(11), pp.1567-1582
- DOI
- 10.1085/jgp.201812119
- PMID
- 30242036
- PMCID
- PMC6219686
- NLM abbreviation
- J Gen Physiol
- ISSN
- 0022-1295
- eISSN
- 1540-7748
- Publisher
- Rockefeller Univ Press
- Number of pages
- 16
- Grant note
- R01HL127764 / NATIONAL HEART, LUNG, AND BLOOD INSTITUTE; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA; NIH National Heart Lung & Blood Institute (NHLBI) 81729004 / National Natural Science Foundation of China; National Natural Science Foundation of China (NSFC) I01BX002900 / Veterans Affairs; US Department of Veterans Affairs 01BX002900 / Department of Veterans Affairs Merit grant; US Department of Veterans Affairs R01-HL127764; R01-HL112413 / National Institutes of Health; United States Department of Health & Human Services; National Institutes of Health (NIH) - USA American Heart Association predoctoral fellowship; American Heart Association
- Language
- English
- Date published
- 11/01/2018
- Academic Unit
- Endocrinology and Metabolism; Internal Medicine
- Record Identifier
- 9984359689202771
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