Aim The isolated, retrogradely perfused heart (modified Langendorff model) is a widely used method in experimental heart research. The presence of an intraventricular balloon is necessary to get functional measurements. We have previously shown that the balloon induces phosphorylation of some suggested cardioprotective mitogen‐activated protein kinases (MAPK): P38‐MAPK, ERK 1/2 and JNK. We hypothesized that the balloon could influence cardioprotection, protect against ischaemia reperfusion injury and interfere with coronary flow. Methods and Results Isolated mouse hearts were perfused for 5, 10, 20, 40 and 60 min with a balloon in the left ventricle. We found a wavelike phosphorylation of all MAPK while AKT displayed a gradual dephosphorylation when compared to non‐perfused hearts. Hearts were subjected to 20 min of stabilization with or without the balloon, followed by 35 min of ischaemia and 120 min of reperfusion. Although the MAPK were phosphorylated, the infarcts were larger in the balloon group. When the balloon was present during the entire protocol, compared to removal at the end of ischaemia, the infarct size was also larger, especially in the endocardial layer. The balloon reduced post‐ischaemic endocardial coronary flow, despite a higher average flow, indicating a hyperperfused epicard. Blocking the balloon‐induced ERK 1/2 phosphorylation during stabilization did not affect infarct size. The effect of post‐conditioning was influenced by the balloon, showing reduced infarct size when the balloon was present. Conclusion The balloon used for pressure measurements may contributes to cell death possibly by reducing endocardial coronary flow.