In studies of gene and cell transfer for the treatment of heart disease, direct intramyocardial injection and antegrade intracoronary injection are common methods of delivering biomaterials to the heart. The authors, who carried out these surgical procedures in 377 rats, describe their methodology in detail and discuss surgical refinements that substantially reduced rat mortality. These refinements include a rigorous fluid replacement regimen, use of inhalational anesthesia instead of injectable agents, exposure of the heart without direct contact and use of a chest drainage cannula to remove air from the pleural cavity and prevent lung collapse.