Biology 450 - Animal Physiology Lab Fall 2006

Lab 5 – Measuring Muscle Activity Patterns in Vivo

Muscle groups measured in vivo can display many of the same properties as individual muscle preparations examined in vitro.  In this lab you will measure activity patterns generated by muscles in human limbs to examine various aspects of muscle function.

The goals of these exercises are:

Background

Overview of muscle contraction

Contraction of muscle is triggered by a membrane depolarization.  This occurs when an action potentialfrom a motor neuron reaches the motor endplate and triggers the release of a neurotransmitter (acetylcholine), which in turn induces an action potential in the surface membrane of the muscle fiber.  The action potential propagates away from the endplate in both directions, exciting the entire muscle fiber membrane.  This excitation sets into motion the entire sequence of events leading to muscle contraction.  Because excitation represents a voltage change across the surface of the muscle cells, it can be measured directly and used to determine when muscles are active, a technique known as electromyography (EMG).

The following review describes the electrical sequence leading to membrane depolarization and muscle contraction:

  1. The surface membrane is depolarized by an action potential.
  2. The action potential is conducted deep into the muscle fiber via the T tubules.
  3. The electrical signal carries from the T tubules to the sarcoplasmic reticulum where Ca2+ is released.
  4. The free Ca2+ concentration in the myoplasm increases to the necessary concentration so that it binds to troponin.  As a result, troponin changes its configuration and allows cross-bridge formation.
  5. Force is generated as myosin heads form cross-bridges with actin, undergo a conformational change, detach from actin, and change conformation back to its original state.  This cycle of events continues, fueled by ATP, as long as Ca2+ concentrations remain sufficiently high.
  6. Ca2+ is taken up again by the SR, cross-bridge formation is inhibited, and the muscle relaxes until the next depolarization.

Electromyography

As with action potentials in nerves, properly placed electrodes can detect muscle membrane depolarization.  The most effective way to measure this potential (that is, to record an EMG) is to insert the electrodes directly into the muscle.  This is typically done using fine wire with a hooked end as the electrode.  The wires are run through a hypodermic needle, the needle is inserted into the muscle, and then the needle is pulled off the wires at their free ends.  Because only the very tip of the electrode wire is free of insulation, recordings from these electrodes typically detect the membrane potential of just a few muscle fibers.

Although the implantation method is the best way of detecting muscle activity, surface electrodes will also detect an action potential.  As was the case with measuring action potentials in nerves using external electrodes (Lab 3), the measured signals are not as strong or "focused" as with internal electrodes.  Instead, surface electrodes detect activity of whole muscles, with relative signal strength being roughly proportional to the number of active muscle fibers. In this lab, you will use surface electrodes to measure the electrical activity of your muscles as part of contraction activation.

Excitation-contraction coupling

An important point to note about EMG's is that they measure the excitation phase of the excitation-contraction process in muscles.  Contraction occurs as a result of excitation, but only after a delay of a few milliseconds.  This delay can be determined by measuring the time lag between the onset of EMG activity for a muscle (excitation) and a signal generated by the resulting muscular contraction – for example, pressing a button to close a circuit.

Force-velocity relationship

The velocity at which a single muscle fiber of a given length (number of sarcomeres in series) contracts is determined by two factors: 1) fiber type (fast or slow) and 2) load on the fiber.  Similar factors affect the velocity of contraction in a whole muscle: 1) the fiber types in the motor units in the muscle and 2) the load on the whole muscle.  However, in a whole muscle the number of motor units recruited also affects the speed of a contraction; the nervous system can continue to recruit motor units within the muscle in order to compensate for an increased load.  Therefore if you increase the load on a muscle in vivo, one of two things should occur: either the velocity of the contraction will decrease, or the EMG of the electrical activity of the muscle will become larger (see below) as more motor units are recruited.

Load and motor unit recruitment

In general, as you increase the load on a whole muscle, it will keep activating motor units until all are in use.  If you continue to increase the load from this point, you should perceive a slowing in the contraction velocity.  As more motor units are recruited, the total force produced by the muscle increases.  Therefore the increased magnitude of the EMG signal is an indicator of the increased muscle recruitment, and an indicator of increased force.  In this lab we will measure the maximum amplitude of the EMG signal (distance from the origin to the top of the signal) as an estimate of the muscle's force.

Antagonistic pairing of muscles

The action of a muscle in the body typically has a counteraction provided by some other muscle or muscles.  For example, the biceps muscle in humans flexes (bends) the arm at the elbow, while the triceps extends (straightens) the arm at the elbow. Thus most muscles in the body are part of antagonistic pairings of muscles – pairs of muscles that act to opposite effect.  When one muscle in an antagonistic pair is contracting, the other muscle is usually inactive, and vice-versa.

Lab Procedures

Note – This lab requires applying surface electrodes to various parts of your arms.  The exercises will be much easier to perform if you wear or change into a short-sleeved or sleeveless shirt.  If you want to examine the activities of leg muscles, you will need to wear shorts under a pair of long pants.

Initial Setup

In these experiments, you will be applying surface electrodes (coated with a gummy conducting substance on one side) to your skin over major muscles.  The electrodes will be wired to our differential amplifiers, which will be connected to the PowerLab units.  You will use Chart, recording at a relatively fast rate, to record EMG's and other signals.

Exercise 1 – Muscle function & antagonistic pairing

In this exercise, you will verify your ability to collect useful EMG signals, and will examine the activity patterns of antagonistic muscle groups.  As with the measurements of action potentials in nerves (in Lab 3), you will use the differential amp to amplify the faint potentials generated at the skin by underlying muscles.

Procedure:

  1. For each muscle you want to record EMG's from, you will need to hook up a pair of EMG cables to one of the amp channels.  One cable goes to the (+), the other to the (-).  Each amp channel you are using must then be connected to a PowerLab channel.  In addition, you will need one ground cable (marked with green tape); this should be hooked to an unused channel directly on the PowerLab.
  2. On the amp, set the mode to "AC", the low pass filter to 1 KHz, the high pass filter to 10 Hz, and the gain to 1000.  Make sure both the (+) and (-) inputs are on for each channel in use.
  3. Use Chart to collect your data.  You will need one channel for each muscle EMG, and for each of these channels the "AC" mode should be selected from the "Input Amplifier" dialog.  For some exercises, you will also need another channel to record a timing signal – "AC" should be deselected for this data.  For initial measurements, record at 1000 samples/sec.  The voltage range will probably have to be adjusted to suit the strength of the EMG signal.  Try 500 mV as a starting point.
  4. When the equipment is ready, test the setup by attempting to obtain a biceps EMG.  Apply two surface electrodes about 5-7 cm apart on the belly of the muscle (i.e., the thickest part).  Attach EMG cables to the electrodes and use medical tape to secure the cables so they do not tear loose.
  5. You will also need to apply a third electrode some distance away to act as the ground.  This electrode should be places to avoid muscular activity.  The inner side of the wrist is one possible location.  Note that only one ground electrode is needed regardless of the number of muscles being recorded.
  6. When the cables are connected and the amp is on, make sure your biceps is relaxed and click "Record". You should expect a little noise in the trace, but if the signal is highly erratic or wandering there is likely a problem with your setup.  Otherwise, try contracting the biceps – you should see increased signal amplitude (in both positive and negative directions) as the muscle is activated.
  7. You can examine an isometric contraction by attempting to lift up the lab bench.  An isotonic contraction can be achieved by lifting an object with a more moderate weight, such as a book or backpack, with your elbow at your side.
  8. Use another set of EMG cables attached to amplifier channel 2 and place the surface electrodes on the biceps antagonist, the triceps, and do the following:
  9. Pick at least two other limb muscles.  Generate a hypothesis of the function of these muscles.  Test these hypotheses using the EMG equipment.
  10. You can also examine the function of some or all of these muscles during simple tasks (e.g., walking slowly, stepping up, doing a push-up).

Exercise 2 – Load and motor unit recruitment

In this exercise, you will examine the relationship between the force production requirements of a whole muscle (i.e., the amount of weight it must lift) and the number of muscle fibers recruited during muscle activity.  This is possible because EMG amplitude is roughly proportional to the number of muscle fibers excited.

Procedure:

  1. Locate a dumbbell and accompanying weights.  Connect the EMG electrodes.
  2. You will be examining the recruitment of additional motor units as the biceps is required to generate more force to lift increasing amounts of weight on the dumbbell.  You should either save a new recording for each weight or add comments to your recording so you know which EMG goes with which weight.
  3. Start with a lightly loaded or unloaded dumbbell.  Slowly lift the dumbbell in an arm curl (bring your hand up to your shoulder).
  4. Either now or at the end of the exercise, estimate the amplitude of the EMG signal as the maximum voltage spikes (minus the baseline if it looks different from zero).
  5. Repeat this exercise with a number of different loads, up to about the maximum you can lift.  Be sure to perform the arm curl at approximately the same slow speed during each trial (although you may slow down a bit at your maximum weight).  Find the amplitude of the electrical signal your muscle is producing at each load.

Exercise 3 – Latent period between excitation and contraction

The delay between the spread of action potentials across a muscle’s surface and the beginning of cross-bridge cycling will be determined by recording EMG activity of a muscle on one A/D channel and a signal created by muscle contraction (in this case, a button being pushed) on another channel.

Procedure:

  1. Locate one of the push-button transducers and attach the cable to an open input channel.  Make sure that this channel is being recorded, and set the voltage range to 10 V.  Be sure that a voltage registers on your button channel when the button is pushed.
  2. For this exercise, you will want to increase the sampling speed to 2000 samples/sec or more to accurately measure the latent period.
  3. You will push the button using your thumb, holding the body of the transducer by wrapping your fingers around it.  This movement of the thumb is an example of flexion.  With your hand empty, locate the muscle or muscle group that will be responsible for the "button-pushing" action.  Do this by pressing your thumb hard against the index finger of a closed fist and determining where contraction is occuring.  Verify with one of the instructors that you have located the correct muscle group.
  4. Apply additional electrodes as needed to record the EMG from the button-pusher muscle and make sure you are getting a good signal.
  5. Start recording and press the button several times.  Allow a pause of a few seconds between pushes.  Try to push the button as rapidly as possible each time to minimize the delay between muscle activation and the appearance of the button’s signal.
  6. Find a few button-pushes that give clear EMG's, then find the delay between the start of each EMG and the voltage increase produced by the button’s action.

Exercise 4 – Force-velocity relationship

As the load on a muscle increases, its shortening velocity should decrease.  This exercise investigates that phenomenon by measuring the time required to perform a biceps contraction while lifting weights of different sizes.

Procedure:

  1. Locate one of the flexion transducers and attach the cable to the bridge amp, then ask an instructor to help you attach the transducer to your elbow.  You will also need to locate a dumbbell.
  2. Place your arm as though you were going to lift the dumbbell.  Place your arm as you would about 1/3 of the way through a lift (or "curl") and find the voltage, then do the same for a 2/3 position.  You will use these positions to determine relative velocity of contraction.
  3. With your arm still in position, have your lab partner hand you a dumbbell with weights attached to it.  Begin with only a moderate amount of weight.  (You might even use just the bar.)  Lift the dumbbell in a curl as quickly as you can (without smashing the weight into your face).  As with exercise 2, save files or insert comments to keep track of which data go with which weights.
  4. Measure the time it took to move your arm between your two reference points (i.e. between your two voltages) in seconds.  The inverse of this value is your relative velocity.
  5. Repeat this exercise a number of time, adding more weight to the dumbbell each time until you can just barely manage to bring the weight up to your shoulder.  Be sure to make the appropriate measurements of velocity for each trial.