Abstract Body: Diastolic dysfunction is a prevalent and therapeutically intractable feature of heart failure which arises from increased ventricular compliance and impaired relaxation. Properly assessing diastolic performance is challenging because it requires simultaneous in-vivo measurement of ventricular pressure and volume. Furthermore, identifying diastolic dysfunction may require the simultaneous application of a stress such as physical exertion. Herein, we demonstrate a preparation that recapitulates the complexity of diastole in working myocardial slices: a force-feedback system providing precision control of pacing frequency, preload, afterload, and a tunable stretch at the end of diastole that mimics late filling driven by atrial systole. We assess the diastolic performance (isovolumic relaxation, early and late filling characteristics) in working myocardial slices from a known model of diastolic dysfunction, ZSF1 Obese rats and their Lean control. We find that parameters relating to myocardial performance such as slice work, stroke length, and twitch force are the same between Lean and Obese animals but parameters of diastolic performance are altered in the Obese rats. Specifically, we observe a decreased separation between peak early and late filling with a reduced diastolic filling interval at the same pacing frequency while we do not observe a difference in isovolumic relaxation time. Taken together, these results indicate that while the work output of myocardium is preserved in ZSF1 Obese rats, a reduction in available filling time and early filling efficiency may indicate a reduced capacity maintain output at higher heart rates.