Nitric oxide (NO) derivatives and reactive oxygen species (ROS) modulate contractile function of respiratory and limb skeletal muscle physiology contractile function. The intracellular processes regulated by NO and ROS remain enigmatic, however. Studies of reduced preparations have identified a number of regulatory proteins that exhibit altered function when exposed to exogenous NO or ROS donors ex vivo. The relative importance of these targets in the intact cell is not known and conflicting theories abound regarding the mechanism(s) whereby NO and ROS regulate contraction. To determine the processes regulated by NO and ROS in vivo, we will use intact fibers isolated from mouse skeletal muscle to address three specific aims:
Aim 1. To determine the regulatory mechanisms by which endogenous NO depresses force production. Pharmacologic probes of NO and cGMP signaling will be used to test three hypotheses: 1A.) endogenous NO acts via cGMP to inhibit calcium transients during tetanic contractions, 1B.) NO does not alter calcium transients in the absence of cGMP signaling, and 1C.) NO inhibits calcium sensitivity of the myofilaments via a cGMP-independent mechanism.
Aim 2. To establish the processes whereby endogenous ROS modulate contraction of unfatigued muscle.
This aim will evaluate intracellular events regulated by ROS and the antioxidant properties of endogenous NO by testing three hypotheses: 2A.) tetanic calcium transients are not regulated by endogenous ROS, 2B.) ROS enhance calcium sensitivity of the myofilaments, and 2C.) endogenous NO inhibits ROS effects on muscle contraction.
Aim 3. To determine the site(s) of action and reversibility of ROS effects in muscle fatigue. Endogenous ROS contribute to loss of force in acute fatigue; recovery of force is accelerated by treating fatigued muscle with reducing agents. We propose to assess the intracellular events that mediate these changes by testing three hypotheses: 3A.) ROS contribute to the early stages of fatigue by decreasing calcium sensitivity of the myofilaments; 3B.) ROS contribute to late-stage fatigue by disrupting calcium homeostasis, and 3C.) oxidant-induced losses in myofilament function and calcium regulation are acutely reversible by a reducing agent.
Showing the most recent 10 out of 36 publications
