BACKGROUND: Exertional dyspnea is a significant clinical concern in individuals with overweight or obesity. The pathophysiology of dyspnea is multifactorial and complex. Previous data suggest that diaphragm and cardiac abnormalities should be considered as likely contributors to obesity-related exertional dyspnea. Additionally, oxidative stress is a causative factor in the general etiology of obesity as well as skeletal and cardiac muscle pathology. Thus, this preclinical study aimed to define diaphragm and cardiac morphological and functional alterations following an obesogenic diet in rats and the therapeutic potential of an antioxidant supplement, N-acetylcysteine (NAC). METHODS: Male Wistar rats (~7 weeks old) consumed ad libitum either lean (20% protein, 70% carbohydrate, 10% fat) or high-saturated fat, high-sucrose (HFHS, 20% protein, 35% carbohydrate, 45% fat) diets for ~22 weeks. Rats receiving HFHS diet were randomized to drink control water or water with NAC (2 mg/ml) for the last eight weeks of the dietary intervention: Lean, HFHS, and HFHS+NAC (n = 8 per group). We evaluated diaphragm bundles (in vitro function and histology) and hearts (weights and echocardiography) for all groups. RESULTS: Final body weights of HFHS rats, but not HFHS+NAC rats, were significantly higher than Lean controls. Neither HFHS diet nor NAC supplementation affected diaphragm specific force (N/cm2), peak power (W/kg), or morphology. In cardiac muscle, right and left ventricle weights (normalized to tibia length) of HFHS rats were greater than those of Lean controls and HFHS+NAC rats. Cardiac functional abnormalities were also present in HFHS rats, with left ventricular fractional shortening (%) and posterior wall maximal shortening velocity (cm/s) increasing compared to Lean controls, but HFHS+NAC rats did not demonstrate these markers of hypercontractility. HFHS rats showed an elevated deceleration rate of early transmitral diastolic velocity (E/DT) consistent with diastolic dysfunction, but NAC eliminated this effect. CONCLUSION: Our data suggest that an HFHS diet does not compromise diaphragm muscle morphology or in vitro function, suggesting other possible contributors to breathing abnormalities in obesity (e.g., neuromuscular transmission abnormalities). However, an HFHS diet resulted in cardiac hypertrophy, hypercontractility, and diastolic dysfunction. Supplementation with NAC did not affect diaphragm morphology or function but attenuated cardiac abnormalities in the HFHS diet. Our findings support future studies testing NAC supplementation in clinical trials of humans with obesity.