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Photolytic release of MgADP reduces rigor force in smooth muscle
A S Khromov1, A P Somlyo, A V Somlyo
1Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville 22906, USA.
Abstract:
Photolytic release of MgADP (25-300 microM) from caged ADP in permeabilized tonic (rabbit femoral artery-Rfa) and phasic (rabbit bladder-Rbl) smooth muscle in high-tension rigor state, in the absence of Ca(2+), caused an exponential decline (approximately 1.5% in Rfa and approximately 6% in Rbl) of rigor force, with the rate proportional to the liberated [MgADP]. The apparent second-order rate constant of MgADP binding was estimated as approximately 1.0 x 10(6) M(-1) s(-1) for both smooth muscles. In control experiments, designed to test the specificity of MgADP, photolysis of caged ADP in the absence of Mg(2+) did not decrease rigor force in either smooth muscle, but rigor force decreased after photolytic release of Mg(2+) in the presence of ADP. The effects of photolysis of caged ADP were similar in smooth muscles containing thiophosphorylated or non-phosphorylated regulatory myosin light chains. Stretching or releasing (within range of 0.1-1.2% of initial Ca(2+)-activated force) did not affect the rate or relative amplitude of the force decrease. The effect of additions of MgADP to rigor cross-bridges could result from rotation of the lever arm of smooth muscle myosin, but this need not imply that ADP-release is a significant force-producing step of the physiological cross-bridge cycle.
Insights
Magnesium adenosine diphosphate (MgADP) release from caged ADP in smooth muscle caused a force decline. The rate of force decline was proportional to MgADP concentration, indicating MgADP binding kinetics in muscle contraction.
Area of Science:
- Muscle Physiology
- Biochemistry
- Cellular Mechanics
Background:
- Smooth muscle contraction is regulated by complex molecular interactions.
- Understanding the role of adenosine diphosphate (ADP) and its magnesium complex (MgADP) is crucial for elucidating muscle function.
Purpose of the Study:
- To investigate the direct effect of MgADP on smooth muscle rigor force.
- To determine the kinetics of MgADP binding to cross-bridges in rabbit femoral artery (Rfa) and bladder (Rbl) smooth muscle.
Main Methods:
- Photolytic release of caged ADP from permeabilized Rfa and Rbl smooth muscle in a high-tension rigor state.
- Measurement of rigor force decline in response to liberated MgADP in the absence of Ca(2+).
- Control experiments using photolysis of caged ADP without Mg(2+) and release of Mg(2+) with ADP.
Main Results:
- Photolytic release of MgADP caused an exponential decline in rigor force, with rates proportional to MgADP concentration.
- The apparent second-order rate constant for MgADP binding was approximately 1.0 x 10(6) M(-1) s(-1) in both Rfa and Rbl.
- Control experiments confirmed the specificity of MgADP's effect, and force decline was independent of myosin light chain phosphorylation state or minor length changes.
Conclusions:
- MgADP binding to rigor cross-bridges influences smooth muscle force.
- The kinetics of MgADP binding are similar in tonic and phasic smooth muscle.
- While MgADP binding can induce myosin lever arm rotation, ADP release may not be a primary force-generating step in the cross-bridge cycle.