Effect of acetylcholine (ACH) on contraction of cardiac muscle. Calcium chloride was applied on the cardiac ring (uniform transverse section of the LV) for maximum pre-contraction followed by dose dependant treatment of ACH. A. The upper and lower panels represent the contraction pattern of WT and Ins2+/-/CBS+/-, respectively. B. The percentage relaxation (indicating the potential of contraction) was calculated with respect to the maximum contraction after treatment with 20 mM CaCl2 and compared among WT, CBS+/-, Ins2+/- and Ins2+/-/CBS+/-. The capability of contraction decreased in the increasing order of CBS +/-, Ins2+/- and Ins2+/-/CBS+/- when compared to WT. Each data point represents mean ± SD of at least six animals.