P21-acitaved kinase 1 (Pak1) is a serine/threonine protein kinase implicated in cardioprotection (1). Mice lacking Pak1 in cardiac tissue (Pak1cko) respond to chronic isoprenaline (Iso) stress with a more pronounced hypertrophy and higher propensity to arrhythmias than wild type (WT) counterparts (1). The role of Pak1 during acute beta-adrenergic stress is under-studied in these mice. We used a high-speed (1kHz) optical mapping system to assess simultaneously calcium and voltage responses of WT and Pak1cko tissue to acute Iso exposure (10nM). Hearts were loaded with calcium (rhod-2-AM) and voltage (rh237) dyes via Langendorff perfusion. Iso caused a significant reduction in refractory period of WT left atrium (LA) (15±6% reduction, n=8) whereas Pak1cko LA exhibited no change (2±3% increase, n=10). Pak1cko LA also exhibited significantly shorter action potential durations (APD) but longer calcium transients (CaTD) than WT. At 135ms pacing interval during control CaTD was 31.73±2.5 ms in WT but 36.01±1.6 ms in Pak1cko (n=15 regions from 8 atria, both genotypes) whilst APD was 25.47±3.3 ms in WT and 20.22±2.3 ms in KO (n=16 regions from 8 atria and n=18 regions from 9 atria respectively). Pak1cko APDs were also significantly shorter in right ventricular preparations (p<0.05, n=17 regions from 9 RV for WT and n=14 regions from 7 RV for Pak1cko). Our data are consistent with a role of Pak1 in both calcium handling and electrophysiology. Lack of APD response to isoprenaline in Pak1cko LA suggests this protein is important in beta-adrenergic signalling. Further work will be required to pinpoint underlying mechanism(s).