Illuminating cell signaling with genetically encoded FRET biosensors in adult mouse cardiomyocytes

Gopireddy Raghavender Reddy, Toni M. West, Zhong Jian, Mark Jaradeh, Qian Shi, Ying Wang, Ye Chen-Izu, Yang Kevin Xiang

Research output: Contribution to journalArticle

3 Citations (Scopus)

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 μM blebbistatin or 20 μM (S)-nitroblebbistatin to a minimal essential medium containing 10 mM HEP ES 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.

Original languageEnglish (US)
Pages (from-to)1567-1582
Number of pages16
JournalJournal of General Physiology
Volume150
Issue number11
DOIs
StatePublished - Nov 1 2018

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Biosensing Techniques
Cardiac Myocytes
Transgenic Mice
Heart Diseases
Genetic Models
Health
Reading
Cell Culture Techniques
Infection

ASJC Scopus subject areas

  • Physiology

Cite this

Illuminating cell signaling with genetically encoded FRET biosensors in adult mouse cardiomyocytes. / Reddy, Gopireddy Raghavender; West, Toni M.; Jian, Zhong; Jaradeh, Mark; Shi, Qian; Wang, Ying; Chen-Izu, Ye; Xiang, Yang Kevin.

In: Journal of General Physiology, Vol. 150, No. 11, 01.11.2018, p. 1567-1582.

Research output: Contribution to journalArticle

Reddy, Gopireddy Raghavender ; West, Toni M. ; Jian, Zhong ; Jaradeh, Mark ; Shi, Qian ; Wang, Ying ; Chen-Izu, Ye ; Xiang, Yang Kevin. / Illuminating cell signaling with genetically encoded FRET biosensors in adult mouse cardiomyocytes. In: Journal of General Physiology. 2018 ; Vol. 150, No. 11. pp. 1567-1582.
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