Direct inhibition of the actomyosin motility by local anesthetics in vitro

Abstract

Using a recently developed in vitro motility assay, we have demonstrated that local anesthetics directly inhibit myosin-based movement of single actin filaments in a reversible dose-dependent manner. This is the first reported account of the actions of local anesthetics on purified proteins at the molecular level. In this study, two tertiary amine local anesthetics, lidocaine and tetracaine, were used. The inhibitory action of the local anesthetics on actomyosin sliding movement was pH dependent; the anesthetics were more potent at higher pH values, and this reaction was accompanied by an increased proportion of the uncharged form of the anesthetics. QX-314, a permanently charged derivative of lidocaine, had no effect on actomyosin sliding movement. These results indicate that the uncharged form of local anesthetics is predominantly responsible for the inhibition of actomyosin sliding movement. The local anesthetics inhibited sliding movement but hardly interfered with the binding of actin filaments to myosin on the surface or with actomyosin ATPase activity at low ionic strength. To characterize the actomyosin interaction in the presence of anesthetics, we measured the binding and breaking force of the actomyosin complex. The binding of actin filaments to myosin on the surface was not affected by lidocaine at low ionic strength. The breaking force, measured using optical tweezers, was approximately 1.5 pN per micron of an actin filament, which was much smaller than in rigor and isometric force. The binding and breaking force greatly decreased with increasing ionic strength, indicating that the remaining interaction is ionic in nature. The result suggests that the binding and ATPase of actomyosin are governed predominantly by ionic interaction, which is hardly affected by anesthetics; whereas the force generation requires hydrophobic interaction, which plays a major part of the strong binding and is blocked by anesthetics, in addition to the ionic interaction.