Aims: Bilirubin is associated with reduced risk of cardiovascular disease, as evidenced in conditions of mild hyperbilirubinaemia (Gilbert's Syndrome). Little is known regarding myocardial stress resistance in hyperbilirubinaemic conditions or whether life-long exposure modifies cardiac function, which might contribute to protection from cardiovascular disease. Methods: Hyperbilirubinaemic rats and littermate controls underwent echocardiography at 3, 6 and 12 months of age, with hearts subsequently assessed for resistance to 30 min of ischaemia. Heart tissue was then collected for assessment of bilirubin content. Results: No difference in baseline cardiac function was evident until 6 months onwards, where Gunn rats demonstrated aortic dilatation and reduced peak ejection velocities. Additionally, duration of ventricular ejection increased progressively, indicating a negative inotropic effect of bilirubin in vivo. Ex vivo analysis of baseline function revealed reduced left ventricular pressure development (LVDP) and contractility in hyperbilirubinaemic rats. Furthermore, stress resistance was improved in Gunn hearts: post-ischaemic recoveries of LVDP (76 ± 22% vs. 29 ± 17% Control, P < 0.01) and coronary flow (96 ± 9% vs. 86 ± 16% Control, P < 0.01) were improved in Gunn hearts, accompanied by reduced infarct area (21 ± 5% vs. 47 ± 15% Control, P < 0.01), and ventricular malondialdehyde and protein carbonyl content. Expression of myocardial nitric oxide-regulating genes including Nos1 and Noa1 were not significantly different. Conclusions: These data reveal life-long hyperbilirubinaemia induces age-dependent hypocontractility in male Gunn rats, and improved stress resistance. In addition, bilirubin exerts sex-independent effects on vascular structure, myocardial function and ischaemic tolerance, the latter likely mediated via bilirubin's antioxidant properties.