Cardioprotection is a generic term given to any stimulus that affords a protection to cardiac muscle to reduce cellular damage during prolonged ischaemia. The most widely studied of these cardioprotective processes, Ischaemic preconditioning (IPC), is activated in vivo by several brief periods of hypoxic insult prior to the prolonged ischaemia. We previously hypothesised that early opening of sarcolemmal KATP channels (SarcoKATP) was part of the protective cascade by shortening action potential duration, sparing intracellular ATP and so reducing cytoplasmic calcium loading, however, our data demonstrated that the ‘traditional’ SarcoKATP, Kir6.2, activity is actually delayed after IPC (1). We hypothesise that the enhanced activity of Kir6.1-containing channels may be a key component in cardioprotection.Cardiomyocytes were isolated by enzymatic digestion from adult male Wistar rats. IPC cells were prepared by adding 3 cycles of 5 mins of halted perfusion with 5 mins of reperfusion prior to cell isolation. Cell-attached patching was used to measure the SarcoKATP currents. Cardiomyocytes were perfused with normal Tyrode (NT) at 30±2°C at a holding potential of -110mV. In IPC cardiomyocytes, the opening probability of Kir6.1 was significantly higher, 0.64±0.11 (n=9, P<0.001) compared with control group 0.12±0.04 (n=11). The contractile function of cardiomyocytes was tested using a metabolic inhibition and simulated reperfusion (MI/R) protocol, as previously described (1), to evaluate cardioprotection. Cardiomyocytes were perfused with NT and contractions triggered by 1Hz electric field stimulation. Cardiomyocytes were treated with metabolic inhibition (2 mM cyanide and 1 mM iodoacetic acid) in substrate free Tyrode’s solution for 7 mins followed by simulated reperfusion with NT for 10 mins. Contractile recovery after MI/R was used to determine cardioprotection. Consistent with our previous studies, contractile recovery was increased from 32±2 % in control to 83±3 % in IPC cells (P<0.0001). Treating IPC cells with 3μM PNU37883A (2), a specific Kir6.1 blocker reduced the contractile recovery to 42±2% (P<0.001). In control cardiomyocytes, contractile recovery was reduced to 24±4% with PNU37883A (P<0.05). All contractile function data from ≥6 experiments, ≥86 cells, ≥3 animals. Data compared using students T-test).Our current study demonstrates that expression of the putative vascular pore-forming subunit, Kir6.1, is evident at the cell surface in the cardiomyocyte, but also that it’s activity is increased after cardioprotective stimuli. These findings suggest that Kir6.1-containing SarcoKATP channels are more active after cardioprotective stimuli. We hypothesise that this opening causes a shortening of the cardiac action potential and hyperpolarisation to help preserve cardiomyocyte function during ischaemia so imparting cardioprotection.