In vitro muscle testing
In vitro muscle testing is a method used to characterize properties of living muscle 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 with oxygen bubbling through in order to keep the tissue alive and metabolically active. Muscles are stimulated to contract by applying electric 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. Stimulation level is often set to the level which ensures maximal motor unit 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 loops. When pennate muscles are used, sonomicrometry 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 muscles (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, 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 loops, 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.