Muscle architecture and physiology

4.1.1. Organization of a muscle: from whole muscle to sarcomeres

A muscle is composed of several muscle fibers each of them being innervated by a motoneuron whose cell body is located is the anterior horn of the spinal cord. One single motorneuron innervates several muscle fibers constituting a motor unit, whose fibers contract simultaneously. The terminal end of the motorneuron axon and the muscle fiber forms a peculiar type of synapse called neuromuscular junction (figure III.11A).

Each muscle fiber is a unique multinucleate cell with a typical architecture dedicated to the conversion of the nervous stimulus, coming from the motorneuron, into a movement, the contraction. To this end, the fiber possesses a contractile apparatus that comprises several type of myofilaments regularly arranged to form repetitive contractile units, called sarcomeres, all along the fiber. There are mainly two types of myofilaments: the thinner one composed of filamentous actin and the thicker one that are myosin filaments. The actin filaments are anchored on the Z-line which itself is linked to the sarcolemma via the costameres. This cytoskeleton complex provides the physical association between sarcomeres and sarcolemma necessary for lateral force transmission during muscle contraction. Myosin filaments are in between the actin filaments. Those myofibers are surrounded by the sarcoplasmic reticulum (SR) that is largely developed in muscle cell to store sufficient amount of Ca²⁺ necessary for contraction. The SR contains Ca²⁺ buffering proteins, among them calsequestrin (CSQ) is the most important to maintain high Ca²⁺ level available for Ca²⁺ release during the contraction (Beard, Wei et al. 2009). The SR can be divided in two parts: the junctional SR (j-SR) formed by terminal cisternae in close apposition (10-15 nm) to the t(transverse)-tubule, that are deep invaginations of the plasma membrane, and the longitudinal SR (l-SR). One t-tubule juxtaposed to 2 terminal cisternae form a typical structure named triad (Sorrentino 2004) (figure III.11B).

Figure III.11: Muscle organization.

Adapted from Marieb, E. and Hoehn, K. (2015). Human Anatomy & Physiology. 9th ed. Pearson Education

(A) Scheme showing a muscle as a bundle of muscle fibers innervated by a motor neuron. (B) Details of a muscle fiber structure depicting the dense network of sarcoplasmic reticulum surrounded actin and myosin filaments organized as sarcomere. To note the deep invagination of the sarcolemma, t-tubule, that formed the triad with 2 adjacent SR terminal cisternae.

4.1.2. Sequential events from the nervous stimulus to the contraction:

The Excitation Contraction Coupling (ECC)

When the action potential (AP) conducted by the axon of the motorneuron arrives at the synapse, the neurotransmitter acetylcholine (ACh) sequestered in vesicles at the axon terminal, is released via exocytosis in the synaptic cleft. ACh binds to its receptor on the sarcolemma that are non-selective cation channels called nicotinic receptors. The binding of ACh to the nicotinic receptor depolarizes the membrane which leads to the opening of voltage gated Na⁺ channels. This triggers the generation of an AP that propagates along the sarcolemma and within the t-tubules (figure III.12) where is localized a voltage sensor, the dihydropyridine receptor (DHPR). The DHPR is a sarcolemma voltage dependent L-type Ca²⁺

channel which undergoes a conformational change upon depolarization. This conformational change is transmitted to the ryanodine receptor type 1 (RyR1, a SR Ca²⁺ channel), specific of skeletal muscle and localized in the terminal cisternae of the SR (Ebashi 1991, Meissner and Lu 1995). When RyR1 opens, Ca²⁺ goes out of the SR and unbinds from CSQ which modifies its aggregation state and allows even more Ca²⁺ unbinding for release within the cytosol

(Ikemoto, Antoniu et al. 1991). To note that the DHPR itself does not participate, at least significantly, in the rise of cytoplasmic Ca²⁺ elevation (less than 5%) as it kinetics of activation is too slow compared to the duration of the AP (2 to 5 ms) (Dulhunty and Gage 1988). Once in the cytosol, Ca²⁺ interacts with Ca²⁺ binding proteins such as Troponin C. Troponin C is part of the actin filament protein complex and masks the myosin binding site on the actin filament.

Once Ca²⁺ binds to troponin C modifying its conformation, myosin can interact with the actin filament. Myosin and actin interaction induces the sliding of the myosin filament and the shortening of the sarcomere: this is the contraction. During the relaxation phase, Ca²⁺ is pumped back into the SR by sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA) or to a lesser extent extruded outside the cell via Na⁺/Ca²⁺ exchanger (NCX) and the plasma membrane Ca²⁺ATPase (PMCA) (Hidalgo, Gonzalez et al. 1983, Hidalgo, Gonzalez et al. 1986, Hidalgo, Cifuentes et al. 1991, Rossi and Dirksen 2006). The combined action of Ca²⁺ pumps and exchangers allows the rapid decrease of cytoplasmic Ca²⁺ level and the unbinding of Ca²⁺ ions to troponin C which returns in place between myosin and actin. SR is refilled and the system is ready for another cycle of excitation-contraction (Gehlert, Bloch et al. 2015) (figure III.13).

Figure III.12: Simplified scheme showing sequential events leading to muscle contraction upon a nervous stimulus.

Transmission of the nervous stimulus from the motorneuron to the muscle fiber. (1) Arrival of the action potential (AP) at the terminal button of the motorneuron axon. (2) Exocytosis of the ACh containing vesicles into the synaptic cleft. (3) Fixation of ACh to its receptor. (4) Opening of the inotropic channel associated ACh receptor that induces Na⁺ entry into the muscle fiber. (5) Sarcolemma depolarization that triggers opening of Na⁺ voltage gated channel and generation of an AP along the sarcolemma (6).

Figure III.13: Simplified scheme showing sequential events leading to muscle contraction upon a nervous stimulus.

(1) AP arrival at the t-tubule. (2) Conformational change of the DHPR upon depolarization. (3) Opening of the RyR1 due to DHPR modified interaction and Ca²⁺release in the cytosol. (4) Binding of Ca²⁺ions to the myofilaments inducing their sliding and sarcomere shortening (5), this is the contraction. (6) Ca²⁺ is released from its target proteins and pumped back within the SR by SERCA pump. AP, Action potential; DHPR, dihydropyridine receptor; RyR1, Ryanodine Receptor 1; SERCA, sarco/endoplasmic reticulum Ca²⁺-ATPase; SR, sarco/endoplasmic reticulum.

4.1.3. Different types of muscle fibers

Muscle fibers that compose a muscle, belong to 3 main categories (fiber type I, IIA and IIX in human) according to their ability to contract more or less rapidly and to endure fatigue. This heterogeneity in fiber type composition allows a muscle to fulfill a wide variety of tasks requiring various level in force contraction and duration. Thus, muscles dedicated to posture maintenance differ in fiber typing from those dedicated to fast twitches. However, the fiber type composition is not static and can changes over time depending on several stimuli, mainly the activity of the motorneuron that innervates the fiber. The type of myosin heavy chain (MyHC) present in a certain fiber determines its belonging to a category but there are other numerous features specific of each fiber type. The most commonly used and those related to Ca²⁺ are detailed briefly in the following table (table III.2). To note, the presence of hybrid fibers in some muscle that possess a mixture of MyHC isoforms (Schiaffino and Reggiani 2011).

Type I Type II

IIA IIX

Contractile speed Slow Fast

MyHC isoform (corresponding gene)

MyHC-β/slow

(MYH7) MyHC-2A (MYH2) MyHC-2X (MYH1)

Primary source of ATP Oxidative

phosphorylation Both Glycolysis

Resistance to fatigue High Intermediate Low

Firing pattern from motoneuron:

Median frequency and longest train duration

18Hz-21Hz and 290s-548s

48Hz-83Hz and 59s-141s

69Hz-91Hz and 0.8s-3.9s

Cytosolic free Ca²⁺ concentration

at rest High Low (efficient cytosolic buffer like

Troponin C)

Ca²⁺ transients Low High (more developed SR, higher density

of RyR1 and SERCA) Table III.2: Comparative table of the different muscle fiber types.

This table pinpoints key differences between fiber type I, IIA and IIB in terms of contraction velocity, metabolic status, nervous stimuli, Ca²⁺ handling. MyHC, Myosin Heavy Chain; RyR1, Ryanodine Receptor 1; SERCA, sarco/endoplasmic reticulum Ca²⁺-ATPase; SR, sarcoplasmic reticulum.

4.2. Overview of skeletal muscle regeneration