In vitro muscle testing is a method used to characterize properties of living muscle

Muscle

Muscle is a contractile tissue of animals and is derived from the mesodermal layer of embryonic germ cells. Muscle cells contain contractile filaments that move past each other and change the size of the cell. They are classified as skeletal, cardiac, or smooth muscles. Their function is to...

 tissue after having removed the tissue from an organism. This allows more extensive and precise quantification of muscle properties than in vivo testing. In vitro muscle testing has provided the bulk of scientific knowledge on muscle structure and physiology, as well as how both relate to organismal performance.

Equipment

In vitro muscle testing typically requires a servomotor, which can both control and detect changes in force and length. One end of the sample tissue is anchored in place, while the other is attached to the motor, and the entire muscle is bathed in Ringer's solution

Ringer's solution

Ringer's solution is the name given to a solution of several salts dissolved in water for the purpose of creating an isotonic solution relative to the bodily fluids of an animal. Ringer's solution typically contains sodium chloride, potassium chloride, calcium chloride and sodium bicarbonate,...

 with oxygen bubbling through in order to keep the tissue alive and metabolically active. Muscles are stimulated to contract by applying electrical current to either the nerve which innervates the muscle or the entire muscle, and the servomotor detects changes in force and/or length due to muscle contraction

Muscle contraction

Muscle fiber generates tension through the action of actin and myosin cross-bridge cycling. While under tension, the muscle may lengthen, shorten, or remain the same...

. Stimulation level is often set to the level which ensures maximal motor unit

Motor unit

”A motor unit is a single α-motor neuron and all of the corresponding muscle fibers it innervates; all of these fibers will be of the same type . When a motor unit is activated, all of its fibers contract...

 recruitment. The servomotor can be programmed to maintain a given force while allowing the muscle to change length, vice versa, or the muscle may be subject to more complex testing, such as in work loop

Work Loop

The work loop technique is used in muscle physiology to evaluate the mechanical work and power output of skeletal or cardiac muscle contractions via in vitro muscle testing of whole muscles, fiber bundles or single muscle fibers...

s. When pennate muscles are used, sonomicrometry

Sonomicrometry

Sonomicrometry is a technique of measuring the distance between piezoelectric crystals based on the speed of acoustic signals through the medium they are embedded in. Typically, the crystals will be coated with an epoxy 'lens' and placed into the material facing each other...

 is often used to accurately determine fiber length during the test.

Scale

In vitro muscle testing can be done on any scale of muscle organization - entire groups of muscle

Muscle

Muscle is a contractile tissue of animals and is derived from the mesodermal layer of embryonic germ cells. Muscle cells contain contractile filaments that move past each other and change the size of the cell. They are classified as skeletal, cardiac, or smooth muscles. Their function is to...

s (provided they share a common insertion or origin, as in the human quadriceps), a single muscle, a "bundle" of muscle fibers, a single muscle fiber, a single myofibril, a single sarcomere

Sarcomere

A sarcomere is the basic unit of a muscle. Muscles are composed of tubular muscle cells . Muscle cells are composed of tubular myofibrils. Myofibrils are composed of repeating sections of sarcomeres, which appear under the microscope as dark and light bands...

, or even a half-sarcomere. Muscle fibers may be intact, or may be "skinned", a process which removes the cell membrane, sarcoplasmic reticulum, and cytoplasm, allowing greater access to the contractile components of the sarcomere.

Typical properties tested

Several properties are commonly tested, and a given experiment will often use a subset of these properties, including twitch times, tetanic force, force-length relationship, force velocity relationship, work loop

Work Loop

The work loop technique is used in muscle physiology to evaluate the mechanical work and power output of skeletal or cardiac muscle contractions via in vitro muscle testing of whole muscles, fiber bundles or single muscle fibers...

s, fatigue trials, fusion frequency, and energetic cost.

In situ

A hybrid approach between in vitro and in vivo has recently been used, called in situ, in which the organism is put under terminal anesthesia, and in vitro tests are performed with the muscle still attached to the organism. This ensures the muscle is kept at the right temperature and amply supplied with nutrients and oxygen by the blood, but the procedure is more difficult and some tests may not be possible.

Species

In vitro muscle testing is almost never used in humans, with the exception of small sections of muscle removed via biopsy or while undergoing surgery for other ailments. Testing is generally more difficult in mammals and birds because of the high temperature and oxygen requirements of the muscle, leading to rapid cell death once muscle tissue is removed from the organism. In ectotherms (reptiles, amphibians, fish, and invertebrates), the muscle tissue can survive outside of the organism for hours or even days, depending on the temperature and organism. Many experiments are conducted at or near 0°C to prolong the usable life of the muscle. Additionally, in fish and amphibians, it is possible to separate out a single muscle fiber while keeping it intact, but in other species this is usually not possible.