The effects of changing loading conditions and inotropic state on maximum isotonic lengthening rate (+ dL/dt, muscle lengths/sec) were examined in isolated rat myocardium. Physiologically sequenced contractions were studied in 18 left ventricular papillary muscle preparations (stimulation rate, 12/min). To study the effects of changing loading conditions, only one loading variable (preload, total load, or late load) was changed during each contraction, while the others were held constant. To study the effects of isoproterenol (10−6M) and temperature (28 vs. 33 ± C) on maximum isotonic lengthening rate, preload and late load were held constant and +dL/dt was examined at a common total load. When preload was increased from 0.7 ± 0.1 to 1.4 ± 0.1 g/mm2, muscle length increased from 0.98 ± 0.003 to 1.01 ± 0.002 muscle lengths, the extent of shortening increased from 0.05 ± 0.003 to 0.08 ± 0.003 muscle lengths, but minimum length (0.93 ± 0.01 muscle lengths) and + dL/dt (1.1 ± 0.1 muscle lengths/sec) were unchanged. When total load was increased from 1.5 to 4.5 g/mm2, minimum length increased from 0.91 ± 0.05 to 0.97 ± 0.05 muscle lengths and + dL/dt fell from 1.4 ± 0.1 to 0.5 ± 0.1 muscle lengths/sec. Late load (the load borne by or applied to the muscle during isotonic lengthening) was altered by changing its magnitude (g/mm2) or time (milliseconds after stimulation) of application. As late load was increased from 1.4 ± 0.02 to 2.1 ± 0.3 g/mm2, + dL/dt increased from 1.3 ± 0.2 to 2.1 ± 0.3 muscle lengths/sec. As the time of late load applications increased from 230 ± 9 to 289 ± 12 msec after stimulation, + dL/dt increased from 1.3 ± 0.2 to 2.5 ± 0.4 muscle lengths/sec. At constant preload and late load and for a given value of total load and minimum length, + dL/dt was faster at a higher temperature (28 vs. 33 ± C) and with the addition of isoproterenol, 10−6M. At a common total load (2.5 g/mm2), +dL/dt rose from 1.6 ± 0.1 muscle lengths/sec at 28 ± C to 2.4 ± 0.2 muscle lengths/sec at 33 ± C and 2.05 ± 0.1 muscle lengths/sec with isoproterenol. Thus, the load against which the muscle shortens (and/or the length to which it shortens), the magnitude of the late load, and the time at which the late load is applied are independent determinants of maximum isotonic lengthening rate. Under any constant set of loading conditions, both isoproterenol and increased temperature increase the maximum isotonic lengthening rate.