Major burn injury results in impairment of left ventricular (LV) contractile function. There is strong evidence to support the involvement of gut-derived factor(s) transported in mesenteric lymph in the development of burn-related contractile dysfunction; i.e., mesenteric lymph duct ligation (LDL) prevents burn-related contractile depression. However, the cellular mechanisms for altered myocardial contractility of post-burn hearts are largely unknown and the cellular basis for the salutary effects of LDL on cardiac function have not been investigated. We examined contractility, Ca²⁺ transients and L-type Ca²⁺ currents (I_Ca) in LV myocytes isolated from four groups of rats: 1) sham burn, 2) sham burn with LDL (sham+LDL), 3) burn (40% of total body surface area burn), and 4) burn with LDL (burn+LDL). Myocytes isolated from hearts at 24 hrs post-burn had a depressed contractility (20%) at baseline and blunted responsiveness to elevation of bath Ca²⁺. Myocyte contractility was comparable in sham+LDL and sham burn hearts. LDL completely prevented burn-related changes in myocyte contractility. Mechanistically, the decrease in contractility in myocytes from post-burn hearts occurred with a decrease in the amplitude of Ca²⁺ transients (20%) without changes in resting Ca²⁺ or Ca²⁺ content of the sarcoplasmic reticulum (SR). On the other hand, density was decreased (30%) in myocytes from post-burn hearts, with unaltered voltage-dependent properties. Thus, burn-related myocardial contractile dysfunction is linked with depressed myocyte contractility associated with a decrease in I_Ca density. These findings also provide strong evidence that mesenteric lymph is involved in the onset of burn-related cardiomyocyte dysfunction.