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SUMMARY

Zebrafish and medaka are powerful cardiovascular models, yet cellular and molecular investigations of adult heart cells have been constrained by suboptimal dissociation and characterization methods. To overcome these barriers, we developed a physiological-temperature workflow that generates high-yield, viable single-cell suspensions for FACS, imaging, and low-input molecular profiling. Using transgenic fluorescent reporters, we consistently isolate ∼6,000 cardiomyocytes per adult zebrafish ventricle and ∼12,000 from medaka, preserving cellular, structural, and molecular integrity. Single-cell morphometrics revealed cardiomyocyte heterogeneity and demonstrated that swimming exercise induces both hypertrophy and hyperplasia, while ventricular injury triggers expansion of regenerative gata4 ⁺ cardiomyocytes. We integrated proteomics and RNA-seq from FACS-purified cells to construct cell-type-specific proteo-transcriptomic atlases. Functional enrichment, transcription factor, and network analyses identified protein hubs and regulatory circuits defining cardiomyocyte and endothelial cell identity. Our platform delivers cell-type-resolved molecular datasets of adult teleost cardiac cells, establishing a systems-level resource for heart regeneration research, cardiovascular disease modeling, and drug discovery.

Teaser

A robust workflow for isolation, FACS, imaging, and multi-omics profiling of cardiomyocytes and cardiac endothelial cells in fish.

HIGHLIGHTS

• Temperature-optimized dissociation and standardized FACS enable high-yield isolation of viable cardiac cells.

• Single-cell imaging uncovers morphological heterogeneity in adult ventricular cardiomyocytes.

• Sustained exercise induces both hypertrophy and hyperplasia of zebrafish cardiomyocytes.

• Proteo-transcriptomics defines core molecular programs and interaction networks in cardiomyocytes and endothelial cells.