Muscle Physiology

Bill Sellers

Email: wis@mac.com

This lecture can be found at:

http://mac-huwis.lut.ac.uk/~wis/lectures/

In order to understand how skeletal muscles work to produce to movement in the body – socalled macro-properties – it is necessary to understand what happens at a microscopic level.In this lecture I will describe the microscopic anatomy of muscle and start to explain what wethink is going on at a cellular level to bring about muscle contraction.

Figure 1 Macro Muscle Structure [McGinnis 1999]

The basic structure of muscle is shown in figure 1. The muscle belly is a bundle of bundles:the bigger bundles are called fascicles and each fascicle is a bundle of muscle fibres. Thecoverings of each bundle also have names with the epimysium covering the muscle itself, theperimysium covering the fascicle and the endomysium covering the fibres. Unlike mosttissues in the human body the muscle fibre does not consist of a large number of cells.Instead the cell membranes have fused together to produce one large ‘supercell’ orsyncytium containing multiple nuclei. The remaining outer cell membrane is called thesarcolemma and the contents are called sarcoplasm. Within the fibre individual contractileunits are called myofibrils .

Figure 2 Light Microscopy [Alexander 1992]

When viewed using light microscopy muscle fibres appear striated as in figure 2. Thissuggests some underlying structure at the sub-cellular level. The striations can be seen moreclearly if viewed under polarised light but to get a really good look you need to use electronmicroscopy.

Figure 3 EM Medium Power [Keynes & Aidley 2001]

In figure 3, a medium power electron micrograph, we can see these details. The differentsections are identified with letters. The dark and light bands that make up the striations areknow as I (isotropic) and A (anisotropic) bands. Within the A band there is a mid-tone H(heller) zone and dark lines can be seen in the middle on the bands: the Z (Zwischenscheiben)and M (Mittelscheibe) lines. The L zone can sometimes be seen as a slightly lighter areaaround the M line. This structure repeats along the length of the fibre and the distancebetween repeats (actually the distance between neighboring Z lines) is called a sarcomere(usually a few µm). This arrangement has been interpreted as a set of interdigitating filamentsas shown in the diagram below the micrograph.

Figure 4 EM High Power [Keynes & Aidley 2001]

When viewed at higher power (figure 4) there is more support for this interpretation. Inaddition cross-linking can be seen running between the filaments. These are know as cross-bridges.

Figure 5 Filament molecules [Keynes & Aidley 2001]

Furthermore it is possible to identify the proteins responsible for this structure with actinfilaments in the I band (light) and myosin filaments in the A band (dark). These two proteinsseems to be primarily responsible for producing contractions. Nebulin and titin seem to bethere to help maintain the arrangement.

Figure 6 Structure Overview [McGinnis 1999]

Figure 6 shows an overview of the probably intracellular organisation or a muscle fibre basedon our current state of knowledge.

What we want to know is how this complex arrangement of molecules actually brings aboutmusclular contraction. The standard theory nowadays is called the sliding filament theorywhich suggests that the actin and myosin filaments themselves do not change length but slideover each other, increasing the amount of overlap during contraction and reducing theamount of overlap during relaxation.

Figure 7 Tension & Sarcomere spacing [Keynes & Aidley 2001]

Figure 7 shows some experimental evidence to support this theory. It shows the maximumforce that a muscle can produce (in this case a frog gastrocnemius muscle that has beendissected out and put in a test rig) when it is stretched to different lengths. As the length ofthe muscle is altered so the sarcomere length changes and this can be measured by observingthe distance between striations. As you can see from the graph the tension falls to zero forvery short and very long muscle and is highest in the middle.

Figure 8 Myofilament Arrangements [Keynes & Aidley 2001]

Figure 8 shows how this sarcomere length translates to degree of overlap of the filaments. Atmaximum stretch there is no overlap and no tension. At minimum length there is overlap butthe myosin filaments are pushed up against the Z lines so the muscle cannot shorten any

further and no external force is generated. At mid-lengths the force generated varies due tothe amount of overlap changing and to interference between the actin strands.

Figure 9 Myosin Molecule [Keynes & Aidley 2001]

It is a good theory but somehow we need to come up with a molecular method for producingthis sliding movement. The force generated seems to be a function of the cross-bridges whichseem to be primarily associated with the myosin molecules. Figure 9 shows a more detailedstructural analysis of the myosin molecule. It consists of 4 peptide subunits and formscomplex chains as indicated.

Figure 10 Actin Molecule [Keynes & Aidley 2001]

As we can see from figure 10, actin chains are much simpler. Actin is a globular moleculeand forms spiral filaments in association with two other proteins: troponin and tropomyosin.When actin and myosin are mixed they produce a viscose complex called actomysion andthis complex acts as an ATPase – that is it splits ATP (adenosine triphospate) into ADP(adensosine diphospate) and P (phosphate) with the normal concomitant release of energy.ATP → ADP + P is the standard chemical reaction that provides energy for cellular activityso this is a promising finding. This reaction seems to be controlled by calcium ionconcentration (with no Ca2+ actomyosin disassociates).

Figure 11 In vitro actin & myosin [Keynes & Aidley 2001]

This experiment can be extended by producing a lawn of myosin on a glass plate which canbe observed to move actin filaments under the influence of ATP (figure 11).

Figure 12 S1 lever arm [Keynes & Aidley 2001]

Detailed analysis of the myosin components has suggested that the S1 part swings when ATPbinds and that this movement acts like a ratchet pulling the actin filament along the myosinfilament. The calcium dependence of the chemical reaction appears to be controlled by thetropomyosin and troponin proteins since in their absence (if pure solutions of actin andmyosin are used) actomysin will split ATP molecules without calcium.

Thus the evidence for this mechanism for muscle contraction is very good. However it is stillonly half the story. We still need to be able to control the muscle contraction.

Figure13 Resting and Action Potentials [Keynes & Aidley 2001]

Muscle fibres are excitable cells just like nerve cells. This means that at rest the inside of thecell is 100mV negative compared to the outside of the cell as shown in figure 13. Whenstimulated, which can be chemical, electrical or even physical, there is a brief change in thelocal electrical potential: the cell membrane becomes depolarised and the inside becomes20mV positive with respect to the outside. This effect is local to the stimulus but the changein potential stimulates the surrounding membrane so that part of the cell becomesdepolarised. However immediately after depolarisation the membrane is briefly no longersensitive to further stimulus so secondarily stimulated areas cannot stimulate the area thatstimulated them. They do stimulate previously unstimulated neighboring areas and this way apulse of depolarisation propagates throughout the whole muscle fibre.

Figure 14 Extracellular recording [Keynes & Aidley 2001]

Figure 14 shows how this wave of depolarisation looks to external electrodes. This showshow the waveform looks rather different. This waveform is from a single muscle fibre andrecordings from a whole muscle is a summation of many individual fibre electrical signalsand so is a much more complex shape.

Figure 15 Ion concentrations [Keynes & Aidley 2001]

The electrical potential is generated by differences in ionic concentrations. These ionconcentrations are electrogenic when the membrane is selectively permeable. The restingpotential is due to the selective permeability of the membrane to potassium ions and can becalculated using the Nernst equation (at room temperature EK = 25loge[K] o/[K] i mV). Thevalues do not quite add up because the cell membrane is not perfectly selectively permeablebut experiments varying the external potassium concentration suggest that the principle iscorrect.

Figure 16 Ion channels [Keynes & Aidley 2001]

The difference in concentration needs to be produced in the first place (and maintainedbecause of leaks) and this is achieved by a sodium/potassium exchange pump (figure 16).This pump pumps out 3 Na+ ions for every 2 K+ ions it pumps in. Thus the pump is itselfelectrogenic. It is also active, requiring ATP to function. As well as this active pump themembrane has two electrically gated channels that are important for the action potential.

Figure 17 Action potential processes [Keynes & Aidley 2001]

Figure 17 illustrates the processes that occur during an action potential. The membranepotential switches rapidly from approximately the Nernst potential for potassium toapproximately the Nernst potential for sodium (depolarisation) then back again.Remembering that the potential across a membrane depends on the ions it is selectivelypermeable to, this is therefore associated with a brief increase in the sodium permeability dueto the activation of the voltage dependent sodium channel. This channel activates veryquickly but only stays open for a very short period causing the positive voltage pulse. Thepotassium channel opens and closes rather more slowly and causes a period ofhyperpolarisation (i.e. a more negative potential) just after the positive spike. Thispotassium effect allows the potential to return to normal more quickly but is not strictlynecessary. The local circuits show how the action potential in one area causes thedepolarisation of neighboring areas.

Figure 18 Calcium effects [Keynes & Aidley 2001]

The action potential still needs to cause muscle contraction. Experiments measuring theinternal calcium concentration of a muscle fibre using the jelly fish protein aequorin (figure18) indicate that depolarisation leads to an increase in calcium level and we already knowthat this triggers the actin and myosin binding. However the calcium ions do not appear tocome from outside the cell.

Figure 19 Muscle T system [Wheater et al. 1979]

Muscle fibres have a system of transverse tubules (figure 19) – invaginations of the surfacesarcolemma – that run into the fibre. These tubules are closely associated with thesarcoplasmic reticulum (a series of membranous structures visually similar to theendoplasmic reticulum in normal cells). This system would seem ideally suited to transmitthe action potential signal.

Figure 20 Ryanodine receptor [Keynes & Aidley 2001]

The T tubule membrane is closely associated to the sarcoplasmic reticulum by structuresknown as ‘feet’. These consist of two sets of receptors (DHP and ryanodine) and it isbelieved that the action potential causes a conformational change to the foot complex whichleads to the release of calcium from the sarcoplasmic reticulum cellular compartment into thesarcoplasm where it leads to contraction. This calcium is eventually transferred back into thesarcoplasmic reticulum by the active calcium pump as indicated in figure 20.

Figure 21 Motor end plate [Wheater et al. 1979]

The only area still to cover is the coupling of the nerve to the muscle. This occurs at themotor end plate (figure 21). This is effectively a synapse with the neurotransmitteracetylcholine being released by the nerve endings. Note how one nerve contacts severalmuscle fibres – this is a motor unit since these fibres will always contract together.

Figure 22 EM of frog neuromuscular junction [Keynes & Aidley 2001]

In figure 22 you can see the vesicles (V) containing acetylecholine ready for release into thesynaptic cleft (C). Note the prominent folds (F) in the sarcolemma and the numerousmitochondria (Mi).

Figure 23 End-plate potential [Keynes & Aidley 2001]

The acetylecholine activates specific channels on the sarcolemma which are believed to allowboth sodium and potassium ions through. This causes a small change in the resting potential

(end-plate potential: see figure 23) which is just sufficient to trigger the voltage gatedsodium channels and start the required action potential.

In summary:

• Force is generated by filaments of actin and myosin sliding over each other• This is triggered by calcium ions• The calcium is released from the sarcoplasmic reticulum by an action potential• Action potentials are due to the change in membrane permeability from potassium to

sodium and back again• This is initially triggered by acetylecholine receptors which change the resting

potential by increasing both the potassium and sodium permeability

Bibliography

Alexander RMcN. Exploring Biomechanics. 1992 New York: Scientific American Library.

Keynes RD, Aidley DJ. Nerve and Muscle. 2001 Cambridge: Cambridge University Press.

McGinnis PM. Biomechanics of Sport and Exercise. 1999 Chamapaign, IL: Human Kinetics.

Wheater PR, Burkitt HG, Daniels VG. Functional Histology. 1979 Edinburgh: ChurchillLivingstone