Strain softening is commonly reported during mechanical testing of passive whole hearts. It is typically manifested as a stiffer force-extension relationship in the first deformation cycle relative to subsequent cycles and is distinguished from viscoelasticity by a lack of recovery of stiffness, even after several hours of rest. The cause of this behaviour is presently unknown. In order to investigate its origins, we have subjected trabeculae to physiologically realistic extensions (5-15% of muscle length at 26 degrees C and 0.5 mm Ca(2+)), while measuring passive force and dynamic stiffness. While we did not observe strain softening in viable trabeculae, we found that it was readily apparent in nonviable (electrically inexcitable) trabeculae undergoing the same extensions. This result was obtained in both the presence and absence of 2,3-butanedione monoxime (BDM). Furthermore, BDM had no effect on the passive compliance of viable specimens, while its presence partly inhibited, but could not prevent, stiffening of nonviable specimens. Loss of viability was accompanied by a uniform increase of dynamic stiffness over all frequencies examined (0.2-100 Hz). The presence of strain softening during length extensions of nonviable tissue resulted in a comparable uniform decrease of dynamic stiffness. It is therefore concluded that strain softening is neither intrinsic to viable rat right ventricular trabeculae nor influenced by BDM but, rather, reflects irreversible damage of tissue in partial, or full, rigor.