Abstract
Vertebrate skeletal muscle contraction and relaxation is a complex process that depends on Ca2+ ions to promote the interaction of actin and myosin. This process can be modulated by nitric oxide (NO), a gas molecule synthesized endogenously by (nitric oxide synthase) NOS isoforms. At nanomolar concentrations NO activates soluble guanylate cyclase (sGC), which in turn activates protein kinase G via conversion of GTP into cyclic GMP. Alternatively, NO post-translationally modifies proteins via S-nitrosylation of the thiol group of cysteine. However, the mechanisms of action of NO on Ca2+ homeostasis during muscle contraction are not fully understood and we hypothesize that NO exerts its effects on Ca2+ homeostasis in skeletal muscles mainly through negative modulation of Ca2+ release and Ca2+ uptake via the NO-sGC-PKG pathway. To address this, we used 5–7 days-post fecundation-larvae of zebrafish, a well-established animal model for physiological and pathophysiological muscle activity. We evaluated the response of muscle contraction and Ca2+ transients in presence of SNAP, a NO-donor, or L-NAME, an unspecific NOS blocker in combination with specific blockers of key proteins of Ca2+ homeostasis. We also evaluate the expression of NOS in combination with dihydropteridine receptor, ryanodine receptor and sarco/endoplasmic reticulum Ca2+ ATPase. We concluded that endogenous NO reduced force production through negative modulation of Ca2+ transients via the NO-sGC pathway. This effect could be reversed using an unspecific NOS blocker or sGC blocker.
Highlights
In skeletal muscle, Ca2+ release from the sarcoplasmic reticulum (SR) is triggered when dihydropyridine sensitive voltage dependent L-type Ca2+ channels (DHPR) mechanically activate the ryanodine receptor (RyR) type 1 (Stephenson et al, 1998)
In order to understand the role of nitric oxide (NO) on the basic response of muscle contraction of zebrafish, the force-frequency relationship was obtained from electrically stimulated decapitated fish using a voltage-time protocol (18 V, 2 ms during 200 ms every 2 min) in control solution only or with addition of an unspecific NO synthase (NOS) blocker (L-NAME) or the NO donor (SNAP) compared to control solution (Figure 1A)
Different to Andrade et al (1998), we (1) used a detailed method to describe different aspects of force and Ca2+ transients we collectively termed biophysical parameters, (2) we analyzed our data based on single twitch stimulation and not from submaximal tetanic stimulation, (3) we used L-NAME to evaluate the endogenous effect of NO on muscle contraction and on Ca2+ transients, and (4) we used zebrafish larvae as an animal model
Summary
Ca2+ release from the sarcoplasmic reticulum (SR) is triggered when dihydropyridine sensitive voltage dependent L-type Ca2+ channels (DHPR) mechanically activate the ryanodine receptor (RyR) type 1 (Stephenson et al, 1998). After Ca2+ release, the contraction process is terminated as a result of Ca2+ uptake mechanisms, removing Ca2+ from the cytosol primarily back into the SR mediated by SR Calcium ATPase (SERCA) and secondarily extruded into the extracellular space by a NO Modulates Calcium and Force sodium-calcium exchanger (NCX). This dynamic and fast Ca2+ movement from the SR into the myoplasm and back into the SR is denominated Ca2+ transient. The time course and the amplitude of the Ca2+ transients determine force production (Stephenson et al, 1998; Baylor and Hollingworth, 2012; Hollingworth et al, 2012)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
