MECHANISM OF FORCE INHIBITION BY 2,3-BUTANEDIONE MONOXIME IN RAT CARDIAC-MUSCLE - ROLES OF [CA2-BRIDGE KINETICS(](I) AND CROSS)

1. We investigated the mechanism of force inhibition by 2,3-butanedion e monoxime (BDM) on rat cardiac trabeculae. [Ca2+](i) was measured by iontophoretic injection of fura-2 salt. Isometric force was recorded a t an end-systolic sarcomere length of 2.1-2.2 mu m. 2. With an externa l [Ca2+] of 1 mM, peak twitch force was monotonically reduced with inc reasing [BDM]; at 5 and 20 mM [BDM], force was 35 and 1% of the contro l force. In contrast, the mean peak [Ca2+](i) during transients was on ly reduced at [BDM] greater than or equal to 10 mM. 3. The duration of the twitch was dramatically reduced by BDM in a dose-dependent fashio n with no significant change in the time course of the underlying Ca2 transients. The abbreviation of twitch force duration was much greate r than expected for the observed reduction in peak force by this agent . 4. The mechanism of the inhibition of force by BDM was explored by e xamining the relationship between twitch force and Ca2+ transients at various values of external [Ca2+]. In the presence of BDM, the steepne ss of the relationship between peak force and peak [Ca2+](i) was reduc ed compared to control conditions. As a result, significant elevation in the [Ca2+](i) transient was unable to reverse the reduction in forc e observed in the presence of BDM. 5. The direct inhibitory effects of BDM on the contractile system were examined using ryanodine tetani in intact trabeculae to measure the steady-state force-[Ca2+](i) relatio nship. In contrast to the effects on twitch force at 5 mM BDM, maximal force was only reduced to 71 % of control. Furthermore, the [Ca2+](i) required for half-maximal activation (Ca2+) was increased while the H ill coefficient was reduced slightly by BDM. 6. BDM dramatically slowe d the rate of rise of tetanic force. At maximal activation, the time r equired to reach 90 % maximal force was prolonged by a factor of 3-8 i n the presence of 5 mM BDM. This suggests that the observed reduction in twitch force and steady-state force may result from slowed kinetics of cross-bridge attachment, consistent with recent biochemical studie s. 7. The contribution of altered cross-bridge kinetics to the effects of BDM was investigated using a co-operative cross-bridge model of th e contractile system. Changing the rate constants for cross-bridge att achment in the model to mimic the reported biochemical effects of BDM reproduced the observed effects of BDM. In particular, changes in cros sbridge kinetics predicted the reductions in maximal force, twitch for ce, and the slope of the peak force-peak [Ca2+] relationship, the shif ts in the Ca-50 for the steady-state force-[Ca2+](i) relation and the abbreviation of twitch duration observed during the application of BDM . 8. Thus, slowing of cross-bridge cycling figures prominently in the negative inotropic effects of BDM. Alterations in Ca2+ transients only come into play at high BDM concentrations (greater than or equal to 1 0 mM).