The ability to move is one of the basic characteristics of living things. To make this point, let us consider a number of different sorts of movement. Most of us are intimately familiar with more or less large scale movements, such as walking or running or watching birds and insects fly. We are also aware of a good number of more subtle movements, such as our heart-beats and movements of our viscera, actions that occur within organisms. Movements also occur at the cellular level. Unicellular organisms are able to move themselves around, by amoeboid movement or the actions of flagella and cilia. Flagella and cilia are also found in certain cells, sperm for example, within metazoan animals. Movement takes place within cells, as well. Chromosomes are moved within cells by contractile elements of a spindle apparatus. Nutriments and other materials are transported within cells, down the length of a nerve axon, for example.

All movements are driven by variations on two general systems of proteins. One is the tubulin-dynein microtubular system, mostly seen within cells, and the other is the actin-myosin filament system of muscles. From the point of view of a comparative physiologist, muscle, and its actin-myosin contractile elements is a special tissue because no other tissue so clearly illustrates the idea of a common mechanism with so very many variations adapted to specific functions.

Turning our attention to insects, one of the most obvious features of insects is that most of them are able to fly. The common names of insects reflect flight again and again, for examples, dragonflies, damselflies, houseflies, mayflies, stoneflies, fruitflies, hoverflies, butterflies. I am sure you can think of many more examples. All bespeak the point that most insects are fliers. We can fairly say that most of the world’s animals fly.

Flight bears on nearly every aspect of insect biology. Foraging for food, searching for mates, escape from predators or fly swatters, avoiding harsh or unfavorable conditions, and locating egg-laying sites are just a few points of insect biology intimately connected to flight behavior. It follows that understanding various behavioral, aerodynamic, physiological, biochemical and ecological aspects of insect flight is valuable in understanding the nature of insects.

We first consider structures of insect flight muscles. Flight muscles (take a look) are arranged into three broad categories based on structural grounds. In tubular muscles, fibrils are arranged radially about a column of sarcoplasm that contains several nuclei. Mitochondria are interspersed in the radial area of the muscle. Dragonflies feature this type of flight muscle structure. Tubular muscles occur in the legs of most insect species. Close-packed muscles are found in Lepidoptera and Orthoptera; people who get their security from scientific idiom call these microfibrillar muscles. In close-packed muscles large mitochondria are dispersed between the cylindrical fibrils, which are about 1.5 to 2.0 µm in diameter. Fibrillar muscles are characteristic of Hymenoptera, Diptera, Coleoptera and true bugs. These muscles have large fibriles, up to 5 µm in diameter, with huge mitochondria and nuclei scattered about.

Even though different phylogenic groups have developed various structural types of flight muscles, there are major similarities among them. All the muscle types have fibers as their physiological units because the fibers are bounded by a continuous sarcolemma or muscle cell membrane with the basement membrane attached. There are descriptions of fibers composed of several cells, however, a fiber is more correctly a single multinucleated cell, or syncitium. Also, the contractile proteins within the fibrils appear to be in similar arrangement.

Now, before we go forward, let us take a minute to clarify some language. People began looking closely at muscles during the 19th century, before articulation of a general cell theory. A consequence of this is a history of set of parallel nomenclatures, one for muscles and another for most other cell types. The terms for muscle usually include the prefix sarc- or my-, from Greek for flesh and muscle. So what we usually call the cell membrane or plasmalemma is called the sarcolemma in muscle. Similarly, mitochondria are called sarcosomes, the endoplasmic reticulum is called sarcoplasmic reticulum and cytosol is referred to as sarcoplasm. Still on nomenclature, muscles are comprised of fibers, which are made up of fibrils within which are the contractile filaments.

The ultrastructural arrangement of muscle is as follows: filaments are made up of the contractile proteins within muscle fibrils. Muscle fibers contain groups of fibrils; muscles are made of groups of fibers.

The most widely accepted mechanism of movement is known as the sliding filament theory of muscle contraction, first proposed in 1954. Each contractile unit is called a sarcomere, bounded on either end by Z-lines or discs. The area between Z-discs is sometimes called a Z-band. Other landmarks are the A-bands, the H-zone and the I-band (take a look). These bands are visible in microscopic preparations and are associated with the presence or absence of the major contractile proteins, actin and myosin. These terms date back to very early microscopic observation of muscles. The A of A-band stands for ANISOTROPIC, and the I of I-band stands for ISOTROPIC. The isotropic bands have uniform indices of refraction in microscope preparations. Consider a cross-sectional representation of muscle fibrils where thin actin filaments are arrayed around thicker myosin filaments. Try to get into the habit of saying thick myosin and thin actin. It may be easier to keep the names straight that way. In flight muscles, the actin filaments are very often found in a hexogonal array around the myosin filaments while in slow twitch visceral muscles the actin is in a circular arrangement. A last point here is that all insect flight muscles are striated. Variations in the register of the Z-discs within a muscle make this feature more or less apparent in muscles from insects of different orders.

In addition to actin and myosin filaments, other proteins are present in relatively small quantities. These include troponin and tropomyosin, collectively called native tropomyosin. The native tropomyosin is involved in regulation of the muscle contraction.

We know, now, that the physiological unit of contraction, the muscle fiber, is invested by the sarcolemma, or functional cell membrane. At frequent intervals this membrane is invaginated to form a system of transverse T-tubules. The invaginated transverse T-tubules lie in close association with vesicles of sarcoplasmic reticulum. Additional invaginations of the sarcolemma allow tracheoles to be very close to the sarcosomes which consume amazing amounts of oxygen during in-flight energy production. Note also that still more invaginations allow motor axons to have multiple synapses with the muscle fiber. This is an important point because unlike vertebrate muscles, the muscle membranes of insects does not propagate action potentials very. There is a requirement, then, for synaptic junctions about every 100 µm or so.

Summarizing muscle structure, the three structural types of muscle, tubular, close-packed and fibrillar, have in common the basic cellular organization of contractile proteins that present a more or less visible striation. Various invaginations for neuromuscular junctions, transverse T-tubule systems and tracheole position appear in all muscle membranes. We can now move on to physiology of muscle contraction.

A motor axon impulse is transmitted across a neuromuscular synapse to the muscle cell membrane resulting in a depolarization which spreads to the interior of the muscle fiber by the transverse T-tubule system. This is very well developed in insect flight muscles. Depolarization of the transverse T-tubule system causes release of calcium from vesicles of the sarcoplasmic reticulum. The calcium activates myosin ATPase, resulting in the two type of filaments sliding past each other. What we want to see is that neither the thin actin filaments nor the thick myosin filaments shorten during contraction. Rather, the Z-bands become increasingly shorter as the two types of filaments overlap each other more and more.

There is a great deal of experimental evidence on muscle contraction. If calcium is removed from an in vitro preparation of locust leg muscle, for example by addition of EGTA which selectively removes divalent cations, the muscle can not be stimulated to contract. Adding calcium chloride back into the buffer restores the ability to contract. Furthermore, calcium is resequestered by vesicles of the sarcoplasmic reticulum in the presence of ATP. The requirement for ATP shows that uptake of calcium requires energy: that is, it takes energy to relax!

The role of calcium in the interaction between thin actin and thick myosin filaments that result in muscle contraction is modulated through native tropomyosin. Troponin inhibits the actin-myosin interactions. This inhibition is calcium dependent due to the presence of tropomyosin. Thus, the contractile process involves an impulse from a motor axon which depolarizes the post-synaptic muscle cell membrane; the depolarization sweeps inward by way of the transverse T-tubules, resulting in release of calcium from vesicles of sarcoplasmic reticulum. Interaction of calcium and tropomyosin, which has specific calcium receptors, temporarily breaks the inhibition of actin-myosin interaction by troponin so that the two main contractile proteins slide past each other with the concomitant expenditure of ATP. The contraction cycle is concluded with the repolarization of the muscle cell membrane, the resequesterization of calcium by sarcoplasmic reticulum and relaxation of the muscle. Please note that relaxation is not the same thing as stretching the muscle to its uncontracted length. Most skeletal muscle systems in vertebrates and invertebrates occur in antagonistic pairs: following relaxation, a flight muscle is restretched by its pair-mate.

Let’s go back to the idea that the two main contractile proteins slide past each other with the expenditure of ATP. We want to consider how the hydrolysis of ATP is translated into directional movement. We begin by appreciating a few points about myosin. Thick myosin filaments have a certain springiness or elasticity. Within a single filament, this allows some myosin heads can be attached to thin actin filaments while others are not attached. Each myosin filament has something in the range of 500 myosin heads, and each myosin head is able to bind and hydrolyze ATP. The process of actin and myosin filaments sliding past each other results from the combined actions of all the myosin heads.

Myosin heads move actin filaments in a cyclic process (take a look). A free myosin head can bind a molecule of ATP, the immediately hydrolyze the ATP into ADP and inorganic phosphate. Both products remain bound to the myosin head, and the energy released by hydrolyzing the ATP forces the myosin head into a highly strained conformation. The energy from the ATP is stored in the strained myosin head. The strained head can then weakly bind to a subunit of an actin filament. The weak interaction between the actin and strained myosin results in the release of the inorganic phosphate. The strained myosin, with ADP still bound, then binds very tightly to its actin subunit. Once tightly bound, the myosin head undergoes a conformational change that pulls the actin filament. This is the "power stroke", at the end of which ADP is released from the myosin head. The myosin head can then bind another molecule of ATP and release the actin. The free myosin can then hydrolyze the ATP, and the cycle continues. A myosin head may go through about 5 contraction cycles per second.

The key point here is that the hydrolysis of ATP is not directly coupled to the power stroke. Rather, the energy from ATP hydrolysis is stored briefly in a strained protein conformation and subsequently released. This is a theme running through many aspects of cell biology. Ion pumps and multi-enzyme macromolecule synthesizing systems similarly capture energy in strained protein conformations.

In addition to placing muscles into three groups based on structure, flight muscles are grouped into two sorts based upon a functional relationship between nerve firings and muscle contraction. In synchronous flight muscle systems, there is a one-to-one correspondence between each nerve impulse and subsequent muscle contraction. Synchronous flight systems are also called neurogenic systems and include Odonata, Orthoptera and Lepidoptera. In asynchronous flight systems, a single nerve impulse gives rise to several muscle contractions. Asynchronous systems are also known as myogenic systems because the stretch of one set of muscles by their antagonistic pair mates stimulates contraction: the contraction is caused by the muscle, myo, rather than nerve, neuro. Diptera and Hymenoptera are examples of myogenic fliers.

The synchronous flight muscle systems are associated with tubular and close-packed flight muscles, and are characterized by relatively slow wing-beat frequencies. Butterflies produce about 4 to 20 beats/second and locusts about 15 to 20. On the other side, the myogenic systems are associated with fibrillar muscles and quite high wing-beat frequencies. Bees and some flies, for examples, operate at about 190 beats/second, with extremes of up to about 1000 beats/second in some very small flies. The highest frequency recorded is 2218 Hz in the midge Forcips , tested at high temperature and with the wings clipped.

At first inspection, the contractile process as we understand it may not seem consistent with such high frequencies. Let us try to reconcile this. In both neurogenic and myogenic systems the muscles are attached such that a very short contraction permits the full excursion of wing movement. This is more extreme in myogenic systems. For example, in Sarcophaga flight muscles contract by only about 1% of their length during flight.

Myogenic muscles can be made to rhymetically contract if the muscle is stretched and set into oscillation by a mechanical device. Stretching the fibrile produces a slightly delayed contraction, and releasing the tension produces a slightly delayed relaxation. The muscle works by adding energy to maintain an already oscillating system. In the whole animal the thoracic box and wings together form a resonating system, and the timing and duration of muscle contraction is mechanically induced. The operation is started by a direct nervous command; after that first impulse, further nervous input regulates the availability of calcium which in turn regulates the power output of the mechanical system by way of the regulatory proteins we mentioned, that is, native tropomyosin. The fact that the muscle is stimulated to contract by stretching rather than by a nervous impulse gives rise to the term myogenic.

In synchronous fliers, clipping the wings has little effect on frequency because the basic frequency is programmed into the thoracic ganglia. In flies and other myogenic fliers, however, clipping the wings nearly doubles with wing beat frequency without a change in membrane potential spike frequency. These muscles can not be driven by electrical stimulation to go as fast as their normal frequency.

Again, this points to the mechanical nature of the asynchronous flight muscle system. In the whole animal the muscles are not attached to the wings, but to particular elements of the thorax such that the wings are part of a lever with two stable positions - up and down. When muscle contractions move one arm of the lever beyond the mid-point, the skeletal elements spring in, which shortens the muscle and releases tension. But at the same time this motion stretches the pair-mate, causing it to contract. The contraction moves the lever in the other direction, and tension is redeveloped as the lever reaches the mid-point. This oscillating cycle goes on as long as the muscle is maintained in an active state by a relatively small number of nerve impulses.

In the presence of calcium, which is released from the sarcoplasmic reticulum due to nerve impulses, myosin ATPase splits ATP during oscillatory work. Stretch itself increases the activity of calcium-activated myosin ATPase - here stretch acts like an increase in calcium. This is also consistent with differences between myogenic and neurogenic systems. In neurogenic systems, each wing beat may be associated with a contraction-relaxation cycle, with substantial movement of calcium out of and back into the sarcoplasmic reticulum. Asynchronous muscles are less sensitive to calcium flux, which is also correlated with reduced development of sarcoplasmic reticulum.