These experiments investigated the effects of physiological concentrations of L(+)-lactate on the contractility of chemically skinned rabbit fast-twitch psoas, slow-twitch soleus, and cardiac muscles at pH 7.L(+)-Lactate depressed maximal calcium-activated force (Fmax) of all muscles studied within the range of 5-20 (slow-twitch muscle) or 5-25 mM (fast-twitch and cardiac muscles). Fmax of fast-twitch fibers was inhibited to the greatest degree (9% in K2 creatine phosphate solutions). In all of these muscle types, Fmax returned to control levels as L(+)-lactate was increased to 30-50 mM. Substitution of neither D-lactate nor propionate for L(+)-lactate significantly altered Fmax. In addition, with the exception of fast-twitch muscle (where the Hill coefficient decreased), L(+)-lactate concentrations, which maximally inhibited Fmax, did not affect the force vs. pCa relationship of muscles tested. These results demonstrate that L(+)-lactate significantly contributes to the depression of muscle function noted during lactic acidosis, directly inhibiting Fmax of the contractile apparatus. This contribution is maximal in fast-twitch muscle where L(+)-lactate is responsible for as much as one-third of the depressant effect on Fmax of the contractile apparatus noted during lactic acidosis.