Bio 102 – City College of New York – Biology Dept.
Experiment 2: Conduction Velocity – How fast is a neuron?
Purpose:
We will study Conduction velocity—measuring the speed at which an action potential propagates down the medial neuron.
Objectives
After this exercise you should be able to:
1. learn how to measure the conduction velocity of neural action potentials using an earthworm preparation.
2. get an introduction to a basic electrophysiology laboratory setup using simple amplifiers and laptop computers to amplify and record neural data.
3. gain general process knowledge such as how to observe, collect, and analyze physiology data.
4. learn statistical hypothesis testing between two populations of data, and novel experiment design
Materials:
· Lumbricus terristrius
· 10% ethanol solution
· 2-channel SpikerBox
· Faraday Cage
· a piece of balsawood or thick cork
· USB cable
· Laptop with Spike Recorder software
· Ruler
· Tape
· Marker
Background
You probably think the nervous system is pretty fast. You seem to hear the spikes immediately when you touch the leg of the cockroach or blow on the cerci of the crickets. But is it instantaneous? Of course not! Not even light, the fastest signal in the universe, travels instantaneously. But how fast is a nervous system? Is it faster than a car, faster than a jetplane, or faster than a cell phone? And how can we measure it?
To measure speed (velocity) though, you need to measure both time (when a spike occurred) and distance (how far a spike has travelled down a nerve).
Take the analogy of a car on a highway. If you were looking out of a small observation hut by the road, you could tell what whether you saw a car, what kind of car it was, and the time that you saw it.
Similarly with your SpikerBox, you can currently tell if you saw a spike, perhaps what kind of neuron generated that spike (we will discuss this in a later experiment), and the time you saw the spike, but you can’t tell how fast the spike was travelling down the nerve.
Let’s go back to the car on the highway. Suppose you had a friend ½ mile down the road in a similar hut:
Later, you two could compare notes to determine the speed of the car. 1 minute = 0.016 hours. Dividing ½ mile by 0.016 hours = 31.25 mph.
[illustration]
So, why don’t we take our two-channel SpikerBox with our two electrodes and ground, put it in the cockroach, and measure the spike output of the two channels? You will notice immediately that there are a lot of spikes happening, in fact, way too many to keep track of it all.
Let’s go back to our analog of the road. Imagine a very busy, fast moving street with many similar looking cars, say, Lakeshore Drive in Chicago, and you and your friend can only set up stations very close to each other.
Ideally, given our limited tools, we’d want to measure spikes on a longer nerve, a nerve with only 1-3 large axons in it, and axons that do not fire many spikes.
Is there any creature in the animal king ...
Bio 102 – City College of New York – Biology Dept.Experiment 2
1. Bio 102 – City College of New York – Biology Dept.
Experiment 2: Conduction Velocity – How fast is a neuron?
Purpose:
We will study Conduction velocity—measuring the speed at
which an action potential propagates down the medial neuron.
Objectives
After this exercise you should be able to:
1. learn how to measure the conduction velocity of neural action
potentials using an earthworm preparation.
2. get an introduction to a basic electrophysiology laboratory
setup using simple amplifiers and laptop computers to amplify
and record neural data.
3. gain general process knowledge such as how to observe,
collect, and analyze physiology data.
4. learn statistical hypothesis testing between two populations
of data, and novel experiment design
Materials:
· Lumbricus terristrius
· 10% ethanol solution
· 2-channel SpikerBox
· Faraday Cage
· a piece of balsawood or thick cork
· USB cable
· Laptop with Spike Recorder software
· Ruler
· Tape
· Marker
2. Background
You probably think the nervous system is pretty fast. You seem
to hear the spikes immediately when you touch the leg of the
cockroach or blow on the cerci of the crickets. But is it
instantaneous? Of course not! Not even light, the fastest signal
in the universe, travels instantaneously. But how fast is a
nervous system? Is it faster than a car, faster than a jetplane, or
faster than a cell phone? And how can we measure it?
To measure speed (velocity) though, you need to measure both
time (when a spike occurred) and distance (how far a spike has
travelled down a nerve).
Take the analogy of a car on a highway. If you were looking out
of a small observation hut by the road, you could tell what
whether you saw a car, what kind of car it was, and the time
that you saw it.
Similarly with your SpikerBox, you can currently tell if you saw
a spike, perhaps what kind of neuron generated that spike (we
will discuss this in a later experiment), and the time you saw
the spike, but you can’t tell how fast the spike was travelling
down the nerve.
Let’s go back to the car on the highway. Suppose you had a
friend ½ mile down the road in a similar hut:
Later, you two could compare notes to determine the speed of
the car. 1 minute = 0.016 hours. Dividing ½ mile by 0.016 hours
= 31.25 mph.
[illustration]
So, why don’t we take our two-channel SpikerBox with our two
3. electrodes and ground, put it in the cockroach, and measure the
spike output of the two channels? You will notice immediately
that there are a lot of spikes happening, in fact, way too many to
keep track of it all.
Let’s go back to our analog of the road. Imagine a very busy,
fast moving street with many similar looking cars, say,
Lakeshore Drive in Chicago, and you and your friend can only
set up stations very close to each other.
Ideally, given our limited tools, we’d want to measure spikes on
a longer nerve, a nerve with only 1-3 large axons in it, and
axons that do not fire many spikes.
Is there any creature in the animal kingdom that meets these
qualifications? Yes! and it is probably right now under your feet
and in your backyard.
In our previous lab we have been introduced to a new class of
invertebrate: annelids! Or more commonly, worms! Enter our
newest preparation: the common earthworm, Lumbricus
terristrius. The earthworm is ideal for studying action potential
conduction velocity in a classroom setting, as the simple linear
anatomy allows easy axon length measurements. The earthworm
contains three large axons that run its length, the “medial giant”
and the two “lateral giant” fibers. The medial giant fiber
transmits information about the front of the worm (the part
closest to the clitellum), and the lateral giant fiber transmits
information from the skin cells of the posterior end of the
worm[footnoteRef:1]. [1: Kladt N, Hanslik U, Heinzel HG.
Teaching basic neurophysiology using intact earthworms. J
Undergrad Neurosci Educ. 2010 Fall;9(1):A20-35. Epub 2010
Oct 15.
]
4. In addition to the earthworm’s long length, which allows us to
place our recording electrodes far apart, the earthworm also
exhibits what is known as sparse coding.
What is spare coding? In a sparse coding scheme, upon
stimulation, the nerves would only fire 1-2 times when you tap
it with a probe, and 1-2 times more when you remove it. This
sparse coding scheme is what we will see in the earthworm
experiment below, and we will exploit to measure the
conduction velocity (or speed) or the spikes.
Procedure:
1. Go to your nearest pet store, sporting goods store, or gas
station and purchase a box of earthworms (they are typically
used to feed lizards, turtles, and fish. Fishermen use them as
bait). They should be around $3-$4 for 12 worms. The
Earthworm box should stay in the refrigerator (not the freezer)
when not being used. The worms can last approximately 1-2
months.
2. Prepare a 10% ethanol solution. The easiest way to do this is
to use vodka (which is normally 80 proof, or 40% ethanol).
Since vodka is not much more than watered down pure ethanol,
dilute it further to 1 part vodka, 3 parts water. For example, we
mix 10 milliliters of alcohol with 30 milliliters of tap water.
Ask your teacher to prepare this for you. Note: You can also use
sugar-free carbonated water (also called club soda or sparkling
water) as an anesthetic if ethanol is not available. The CO2 in
the water serves as an anesthetic agent.
5. 3. Place a healthy earthworm in the alcohol mixture and wait
seven minutes. Do not wait too long; as with human anesthesia,
the delicate balance between too little anesthesia and too much
is tricky. To little anesthesia and the earthworm will move
around during the experiment, the resulting muscle electrical
activity (electromyogram) will drown out the small neural
electrical signals you are interested in. Too much anesthesia and
the nerves will not fire. We’ve found 7-10 minutes is a good
range.
4. Place the Earthworm on a piece of balsawood or thick cork,
and put your three electrodes of your two-channel SpikerBox in
the posterior end of the worm, like this:
5) Plug the electrodes into your Neuron SpikerBox Pro and the
USB cable into your PC. Place the entire set up into a Faraday
cage. Why are we using a Faraday Cage for this experiment?
6) Open our Spike Recorder software, and click on the USB
symbol to pair with the Neuron SpikerBox Pro.
7)Press the record button on your Spike Recorder software, and,
using a plastic or glass probe, tap the posterior end of the worm.
You should hear the evoked spikes caused by the tap.
Interestingly, the neurons in the earthworm are myelinated
(covered in insulating fat), and you will notice the spikes are
much quieter than you are used to.[footnoteRef:2] Many nerve
diseases, such as Multiple sclerosis, are caused by a
degeneration of this fatty covering. [2: Hartline DK, Colman
DR. Rapid conduction and the evolution of giant axons and
myelinated fibers. Curr Biol. 2007 Jan 9;17(1):R29-35.
6. ]
8) Make 3-4 taps, separated by about 3-4 seconds each.
9) Measure the time delay between the two channels by clicking
at the beginning of one spike, then dragging to the start of the
next. In the corner of the screen you should see the amount of
time you have selected.
10) Using a ruler with divisions in the mm range, measure the
distance between the electrode one and electrode two.
11) Divide the distance by the time. Viola! You have just
measured conduction velocity. Repeat 4-5 times on other spikes,
using the table on the following page.
12) Remove the electrodes from the worm, dip the worm briefly
in water to remoisturize it, and return the worm to its Styrofoam
container. It can tolerate the needle placement and be used for
another experiment another day, or you can return it to the
environment where you found if you live in wet climates where
Earthworms can be harvested in your backyard.
Data Analysis
Now start exploring:
· Does this measurement change from spike to spike?
· Does it change from earthworm to earthworm?
· Are smaller earthworms faster or slower then large
earthworms?
· Is this speed sensitive to duration of anesthesia?
To help you learn how to identify Earthworm Spikes, here is an
Earthworm Recoding sample audio file.[footnoteRef:3] [3:
http://www.backyardbrains.com/experiments/files/recordings/Ea
rthworm_Double_Recording.mp3]
7. Next Steps
Have you ever wondered, when you stub your toe, how you
manage to feel the impact almost instantaneously, but the
throbbing pain takes about 1-2 seconds to reach your
consciousness? This is because these two signals (touch vs.
damage/pain) travel via two different fiber systems that have
very different speeds. Below is a teaser as to why - want to dive
deeper? We will investigate this in the next experiment:
Comparing Speeds of Two Different Fibers.
Trouble-shooting:
If your earthworm is not healthy (not moving around the soil
and not resisting/squirming when you try to pick it up), you will
not get good recordings.
Discussion Questions:
1) Why are we using alcohol to anesthetize the earthworm
instead of ice water?
2) What happens if you reverse the ground and recording
electrode 1?
3) What happens if you touch the anterior part of the worm (the
mouth).
4) Define Spare Coding
5) What is the purpose of the Faraday cage?
/
Topic 5
WEEK 5
8. Drug and Alcohol Use In Pregnancy, Fetal Alcohol Syndrome
Disorders, OB & Pediatric Complications
Homework
Case Study #2 (Originality scores greater than 30 will not be
accepted)
Complete Journal Entry #4 for the Abstinence Project
Topic 6
WEEK 6
Understanding Co-Occurring Diagnosis in Substance Use
Disorders.
Homework
Abstinence Final Project Paper due (Originality Scores greater
than 30 will not be accepted.)
(No Additional Assignments or Forums due this week)
6. What types of treatment plans and nursing interventions
would work best for this adolescent patient? Please
provide suggestions based on your literature review.
Abstinence Project Journal Entry #3
National Institute on Drug Abuse Video: Babies Born to Women
Addicted
WHO: Guidelines for the identi�cation and management of
substance use and substance use disorders in
pregnancy
ACOG: Opioid Use Disorder in Pregnancy
9. AAP Policy Statement: A Public Health Response to Opioid Use
in Pregnancy
AAP: Neonatal Drug Withdrawal
Neonatal Abstinence Syndrome (NAS) Treatment Guidelines &
Scoring
Abstinence Project Journal Entry #4
Case Study #2 Working with the Vulnerable Population
SAMHSA's Article on Co-occurring disorders
Final Abstinence Project Analysis Paper
Please submit your APA formatted, double spaced
conclusion paper about how this experience will assist you in
understanding or working with chemically dependent people.
Refer to the syllabus supplement for further
explanation and rubric guidelines. Originality scores greater
than 30 will not be accepted.
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Topic 7
WEEK 7
Medication-Assisted Treatment based on
symptoms. Motivational Interviewing, Medications, Treatment
modalities.
Observation of an OPEN AA/NA Meeting
Homework
Discussion Forum #5
Observation of a Virtual OPEN AA/NA Meeting
Use the following link to �nd a virtual meeting that �ts your
schedule. Remember to only attend an OPEN
meeting that is not for members only.
https://aa-intergroup.org/oiaa/meetings/?types=Open
Motivational Interviewing
APA Practice Guideline for the Pharm Tx of Patients with
Alcohol Use Disorder
TIP: Medications for Opioid Use Disorder
Alcoholics Anonymous: This is the Fourth Edition of the Big
Book, the basic text for Alcoholics Anonymous.
Prior to attending the open AA Meeting, please read Chapter 5:
How it Works in The Big Book. The Big Book is
12. the Basic Text for Alcoholics Anonymous. The entire book is
available by clicking on the above link.
NA Recovery Literature, Booklets, and Pamphlet resources
Attendance to an OPEN AA/NA Meeting
Please submit the date, time, location and type of meeting that
you attended. Attendance contributes to 5% of
your grade.
Forum #5: Understanding Co-Occurring Disorders and Self-
Re�ection of a 12 Step Program
Forum Directions: You should make a minimum of 2 postings in
total: one new thread and one thoughtful
responses to a classmate. (Please refer to the course syllabus
supplement for forum rubric information.
Your participation will be graded on a ten point . A researched
reference is NOT needed for this self
re�ection forum.
Participation in 12-step groups has been associated with
positive outcome among substance users.
Nurses can play an important role in fostering 12-step
participation, and the insights which they develop
in their practice can greatly contribute to a patient's willingness
to participate in a 12-step program.
Please answer the below questions as honestly as possibl e. /This
forum is based on your opinion and
observations. A researched reference is NOT needed. This
forum focuses on self-re�ection of your 12-step
meeting observations.
Questions:
13. 1. De�ne co-occurring disorders related to psychiatric/mental
illness and substance use disorders. What is the
prevalence of this disorder? (include literature research in your
response.
2. Explain what is meant by the term "substance induced mood
disorder". State the risk factors involved for this
disorder.
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15. you feel that clients should be mandated to attend self-help
meetings? Why or why not?
5. Before attending the 12 step meeting, what were your
attitudes, beliefs, or previous knowledge about these
programs? Your personal reactions as an individual attending
for the �rst time.
6. What did you observe / learn about yourself by attending?
Were you nervous? Why or why not? Did you have
any stereotypes that were con�rmed or shattered? What
emotions did you experience?
7. Did this opinion change after attending the meeting? Please
be speci�c and state how it did or how it did
not. Please describe one thing that surprised you or something
that you new that you learned while attending
the meeting.
8. What social, psychological and spiritual principles of human
behavior are the 12 step programs based on?
How do they contribute to its success? What potential barriers
do you see for patients participating in a 12 step
program. What suggestion do you have to alleviate this barrier?
Video: Substance Use Disorder (SUD) in the Nursing Profession
Virginia Health Practitioners' Monitoring Program
Maryland Board of Nursing: Rehabilitation Program
A Nurse’s Guide to SUD in Nursing
A Nurse Manager's Guide to SUD in Nursing
Screening for Substance Abuse Video
16. VA Depart of Health: Enforcement Disciplinary, Nursing Case
Decisions
Forum 6: Impaired Nursing Forum
Forum Directions: You should make a minimum of 2 postings in
total: one new thread and one thoughtful
response to a classmate. (Please refer to the course syllabus
supplement for forum rubric information.
Your participation will be graded on a ten point . A researched
reference is needed for this forum.
Review the below Case Study for a better understanding of
potential examples of impaired nursing in the
clinical environment. Answer the following questions.
CASE REPORT: The Supernurse
Monthly reports from the controlled substances vault showed
that a registered nurse on one of the patient care
units had a 3+ SD use of controlled substances compared with
fellow nurses in the same time frame. Further
investigation revealed that this nurse was the only one w ho gave
several patients acetaminophen with
oxycodone after charting their pain scales. Review of the charts
showed that the highest doses were always
removed, and no doses were wasted. The unit’s nurse manager
vouched for the nurse in question, saying she
was one of the most well-respected and reputable nurses in the
department. She insisted that this nurse was
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Bio 102 – City College of New York – Biology Dept.
Experiment 3: Comparing Speeds of Two Nerve Fiber
SizesObjectivesBy the end of the lab, students will
· learn how the earthworm's lateral and medial giant nerve
fibers transmit different sensory information from different
parts of the worm allowing the escape withdrawal reflex in
awake, behaving worms. · learn about Cable Theory and how
neuroscientists have been applying it to explain the physical
properties of axons.
· learn statistical hypothesis testing between two populations of
data, and novel experiment designMaterialsThe materials
required for this lab are exactly the same as the previous
experiment: Introduction to Conduction Velocity (Neural
Speed)Background
Previously, you learned how to measure conduction velocity
from the earthworm's nerve fiber system. You remember that the
worm has three large neurons that run the length of its body, the
medial giant nerve (MGN) and the two fused lateral giant nerve
(LGN).
Let's take a closer look at the ventral, or "bottom," nerve cord
containing these medial and lateral giant nerves. One of the
differences between invertebrates (insects, worms, and so on)
and vertebrates (dogs, lizards, us) is that invertebrates have a
ventral nerve cord (running along their "belly") whereas we
have a dorsal nerve cord (our spinal cord runs along our
backside).
Both the MGN and LGN play an important role in ensuring the
worm's senses communicate with its muscles (Drewes et al.
1978).[footnoteRef:1] The MGN transmits sensory information
about the anterior or front of the worm (the end closest to the
clitellum). In contrast, the LGN transmits sensory information
about the posterior, or back of the worm (the end farthest from
19. the clitellum). There is a physical size difference between these
two systems as well. The medial giant nerve, at 0.07 mm in
diameter, is slightly wider than the lateral giant nerve (0.05 mm
in diameter) (Kladt et. al 2010).[footnoteRef:2] [1: Drewes CD,
Landa KB, McFall JL. Giant nerve fibre activity in intact, freely
moving earthworms. J Exp Biol. 1978 Feb;72:217-27. ] [2:
Kladt N, Hanslik U, Heinzel HG. Teaching basic
neurophysiology using intact earthworms. J Undergrad Neurosci
Educ. 2010 Fall;9(1):A20-35.
]
In the previous earthworm experiment, you recorded from the
rear, or posterior end of the worm, and determined the
conduction velocity for the LGN. For this experiment, you will
record from both the posterior (LGN) and anterior ends of the
worm (MGN). We want to find out if there is any difference in
conduction velocity between the two nerves. Do you think there
will be any difference? Let's consider some.....
When thinking about how an action potential travels down a
neuron's axon, it is useful to think about an analogy of a
television's volume. Think about turning on your TV and then
slowly walking away from it. As you walk further and further
away what happens?
The sound coming from the speaker gets quieter and quieter the
farther you are away from the source. This example is
analogous to a voltage change (basis of an action potential)
flowing down a neuron's axon. In a hypothetical neuron with the
active ion channels removed, let's change the voltage in the cell
body and take three measurements along the axon. What do you
think the measurements will look like?
20. Notice that the signal decays. The strength of this decay is
determined by two things, the time constant and length constant.
Time for some math and electronics, our favorite subjects
(besides neurons of course).
What do the r's and c's mean? r is "Resistance" to current flow,
and c is "Capacitance," a measure of the storage of charge
across an insulating barrier.
First, let's talk about the length constant (this is sometimes also
called the "space constant"). The length constant (λ, or lambda)
is a measure of how far the voltage travels down the axon
before it decays to zero. If you have a length constant of 1 mm,
that means at 1 mm away from the cell body in an axon, 37% of
the voltage magnitude remains. At 2 mm away from the cell
body in an axon, 14% of the magnitude remains, and at 3 mm
away, 5% remains. This is representative of an "exponential
decay" function.
The length constant is calculated from rm and ri. rm is the
electrical resistance of the neuron's membrane, or how
"electrically leaky" it is. The larger rm ("less leaky") is, the
larger the length constant will be. ri is the resistance of the
intracellular fluid (called axoplasm) inside the axon.
Conversely, the lower ri is, the larger the length constant will
be.
The time constant (Τ, or tau) is similar to the length constant,
but applies to time. If a voltage change is applied i nside a
neuron, it takes time for the neuron to fully "charge" to a stable
voltage. In the time constant equation, cm is the capacitance of
the neural membrane, which is a measure of the membrane's
ability to store charge. The higher the capacitance, the more
time it takes for the capacitor to fully charge (or discharge),
acting as a "buffer" to any sudden voltage change.
21. Thus, the smaller both rm and cm become, the smaller the time
constant is and the less amount of time is needed to change an
axon's voltage.
An "ideal neuron" would have an infinitely high length constant
and an infinitely low time constant. Thus, any voltage change
anywhere in the neuron would instantly change the voltage
everywhere else in the neuron.
Both the time constant and length constant are "passive"
properties of the neurons. So, how do your neurons stop
electrical signals from decaying to zero? By becoming "active"
and using Ion Channels! Your neurons use sodium and
potassium channels to regenerate the action potential flowing
down the axon to "fight the decay" that occurs due to the length
and time constants. As an action potential fires down your axon,
sodium and potassium channels continually open and close to
recharge the action potential and "propagate it" down the axon.
As you know from the previous earthworm experiment, this
action potential propagation down a neuron has a finite speed.
Each time an ion channel needs to open to recharge the action
potential, this delays the propagation of the action potential by
~1 ms. And the smaller your length constant is, the more you
have to regenerate the action potential by having ion channels
open along the length of the axon. How can we increase the
length constant? We can do this by increasing rm. Is there a way
we can do this?
Yes! We can increase rm by wrapping the neuron in....
Myelin is a fatty covering produced by special cells called
Schwann Cells and Oligodendrocytes. This covering is what
makes the axons look similar to hot dog rolls, and why the brain
22. is sometimes called a "lump of fat." This fatty covering makes
the neural membrane less leaky and increases rm substantially.
But what do you think would happen if you covered the whole
axon in myelin? Unfortunately, the length constant is not
increased enough for you to get away with this. The action
potential still needs to be regenerated along the axon, though
not as many times as an unmyelinated axon.
This is why the myelin sheath is discontinuous, with periodic
exposed bits of neural membrane called "Nodes of Ranvier." In
these nodes no myelin covers the membrane, and plenty of
active ion channels reside there. The discrete regeneration of
action potentials between lengths of myelin at the Nodes of
Ranvier is called "saltatory conduction."[footnoteRef:3] [3:
Related Fact: Saltar is Spanish for "to jump." A grasshopper is
called "Saltamontes" or "hill jumper."]
But wait! Covering the neurons with myelin makes the inside
and outside of the neural membrane farther apart from each
other. As capacitance is affected by the distance of separation
between the charged bodies (see your Haliday and Resnick),
myelin will decrease cm. Does this cause a decrease in time
constant as well then? Well, perhaps not, since, as we said
earlier, the myelin also substantially increases rm.
The result of this simultaneous reduction in cm and increase in
rm is hypothesized to cause no net change in the time constant,
although direct experimental evidence in the literature is
lacking. If you have two equal diameter axons and one has a
myelin sheath of 1 mm thickness, and the other has a myelin
sheath of 2 mm thickness, how much faster will the second axon
be? Unfortunately, again, this answer appears to be
23. experimentally unknown, as neurons with increased myelin
thickness also simultaneousl y have increased axon diameter.
What has been generally borne out with computer simulations is
that a myelinated neuron twice as thick as another myelinated
neuron will have a conduction velocity twice as fast.
There is another way to increase the conduction velocity
without bothering with all these special cells that coat the
neurons with fat. This method is also what many invertebrates
use...
The larger the radius of the axon, the smaller both ri and
rm will be. Remember our length constant equation states that :
If both the top and bottom vary with the radius... it seems like
the size of the axon wouldn't make a difference at all! But let's
take a careful look at how these two values vary with the size of
the axon. The membrane resistance (rm) changes with the
circumference of the axon (where the membrane is) like so:
while the internal resistance changes with area of the axon.
Both Ri and Rm are constants that can be measured from the
neuron regardless of it's size (whereas ri and rm DO take size
into account), π is 3.14, and radius is the radius of the axon. So
now let's look at that equation again:
We are interested in seeing what changes when we change the
size of the axon (radius), so we want to remove things that are
constants and see what is left that changes. Both Rm and Ri are
constants, so are 2 and π, and one radius cancels out. We are
left with simply that:
Thus, the length constant, and conduction velocity, scales with
the square root of the radius.
24. Note that the benefits of myelin substantially outweigh the
benefits of axon diameter size. Tripling the myelin thickness
increases the conduction velocity 3x, whereas tripling the axon
diameter only increases the conduction velocity by the square
root of 3, or 1.7 times. There is a metabolic cost, however, to
making myelin (you need to keep the special cells alive that
coat the neurons in fat), so it's not the perfect solution for all
animals. But...even the largest axons without myelin in the
animal kingdom, such as the 1 mm diameter squid giant axon,
only has a conduction velocity of 20-25 m/s second! You have
myelinated axons in your body (the A alpha fibers) that are only
13-20 μm in diameter (1/100 of the size of the squid axon), yet
have conduction velocities that are 80-120 m/s! Myelin is a
wonderful biological invention, allowing neurons to get both
small and fast, but it is expensive.
Sound confusing? Don't worry, it was confusing for us too
during our education.
Welcome to "Cable Theory", which was originally developed in
the 1800's when engineers were trying to understand signal
transmission across long distance telegraph lines.
Neuroscientists then applied this theory to neurons in the early
20th century.
But what does all this cable theory mean with regard to the two
nerve types in the earthworm? Keep in mind that MGN is 1.4
times greater in size than the LGN. What do you expect in terms
of speed of conduction? We previously measured the LGN to be
~10-14 m/s, so what would you expect the MGN speed to be?
Let's try the experiment to see if our results match the
theory!State you hypothesis and justification
below:Hypothesis:Justification:Procedure
1. Anesthetize and take a recording of the posterior end of the
worm as you did in the previous experiment.
2. Once you get several spikes by tapping the posterior end with
25. a probe three times, rotate the worm 180 degrees and reposition
the electrodes. You will be measuring from the anterior end of
the worm this time.
3. Now record several spikes from the anterior end by touching
the head of the worm with a wooden probe three times. Once
you have several spikes, you can return the worm to its soil.
The earthworm is quite resilient and recovers well from this
experiment.
4. Now you are ready to look at your data. You should see a flat
line or excessive noise when you flipped the electrodes around.
This serves as your time marker of when you flipped the worm,
and now you know which spikes belong to the posterior end and
which spikes belongs to the anterior end. The figure below
shows a recording of electrode 1 on the bottom and electrode 2
on the top.
5. You can now zoom in on your spikes and measure the
conduction velocity. Take readings of 5-6 spikes.
6. Repeat the experiment several times with some worms. This
will give you a good data set to work with. Don't forget to clean
your electrodes with some alcohol or water and a paper towel
after each worm.
7. You now need to run a statistical test, namely the T-test, to
examine whether the conduction velocities are different for the
two nerves. If you do not yet know how to do this you can take
your data set and follow along in our statistics lesson plan
online. If you have done this lesson plan or have some
experience in statistics then you can go ahead and perform the
calculations below.
8. Take the average and standard deviation of your MGN and
LGN recordings.
9. Finally, let's calculate our t-statistic and p-value.
26. What did you find? Are the two conduction velocities different
from each other?Discussion
What have you found about the MGN (anterior end) conduction
velocity in relation to LGN? If it is faster, how many times
faster? Why is this? You may recall that the earthworm neurons
are actually myelinated! Some invertebrates, such as some
shrimp and some worms, actually do have myelin.
Typically, as axon increases its diameter, its myelin thickness
also increases. Perhaps the MGN has a thicker myelin sheath as
well. This would make for an excellent histology project to find
out. Let us know if you are up to the challenge, and let us know
what you find!
In your discussion, you should explain what causes this
unexpectedly large difference. Also, discuss why having a
longer time constant increases conduction velocity.
Discuss your class findings and compare what you found in
class to previously documented conduction rates of the nerves
of a wide variety of vertebrate and invertebrate animals shown
below. Are your data consistent with this broader data set?
Figure: Velocity of nerve impulse conduction as a function of
fiber diameter in a variety of animals. Modified from Bullock
and Horridge, 1965, Structure and function of the Nervous
System of Invertebrates. W. H. Freeman and Company.
Other Questions to Consider in the Discussion
1. Does the anesthetic have an effect on the conduction
velocities of the MGN and LGN?
2. Does a worm's general size have an effect on conduction
velocity?
3. You can also anesthetize the worm in a 40% - 60%
carbonated water solution for 5-9 minutes as an alternative
27. anesthetic. Will this change the conduction velocity
measurements?
4. The worm Lumbriculus variegatus (California Blackworm)
actually has a larger LGN than MGN, so we would expect our
results to be the opposite of what we observed here with
our Lumbricus terrestris nightcrawlers. Why?Troubleshooting
This can sometimes be a difficult experiment, because the worm
may not produce spikes depending on the amount and time of
anesthetic used as well as the general health of the worm. If you
stick to the 10% alcohol solution for about 3-6 minutes, the
worm should produce spikes most of the time as soon as you
start (don't forget to wash the worm in water after you
anesthetize it).
You may also want to try touching the worm with more or less
pressure. Sometimes a very small tap will work, other times a
stronger press might be needed. Some worms respond better to a
stimulus at the very end of their bodies, while other respond
better to a stimulus a few centimeters inwards.
Finally, sometimes you will cause an artifact when you touch
the worm. Looking closely on the artifact waveforms, the
artifacts will appear at exactly the same on both channels. This
is a fake spike and not physiological! Sometimes, drying your
probe periodically helps; also do not rehydrate the worm in
water too much (though also be careful not to dry the worm
out). It is a careful balance, and you will develop your own
style and technique as you gain experience.
You can also use an air stimulus from an air can in lieu of a
plastic, wooden, or glass tip if you are getting too many fake
spikes. You may also want to flip the worm over so the ventral
or bottom side is facing up. Doing this means when you touch
the worm with your probe the touch will be closer to the nerve.
Lab Report Assignment:
28. Use data from the whole class to compare the mean conduction
velocities of the two nerves examined today. For each worm,
take five measurements from different spikes in both the LGF
and MGF. Conduct a t-test comparing the speed of conduction
(m/s) for the combined earthworm data for the two nerves. Is
the difference significant at the 0.05 alpha value probability
level? On the physiological level, what is responsible for this
difference in conduction velocity between these two nerves?
Present your experiment in a lab report format.
Title
Abstract
Introduction and Background with hypothesis and justification
Materials and Methods
Results - Use data from the whole class to compare the mean
conduction velocities of the two nerves examined. Conduct a t-
test comparing the speed of conduction (m/s) of the two nerves
for the class data. Present your data as:
· Fig 1 shows sample recordings and traces from an individual
earthworm.
· Table 1 shows conduction velocity (m/s) values for individual
worms for MGF and LGF measurements, standard deviation
· Fig 2 shows the compiled data in the form of scatter plot for
Intra and Inter Worm Measurement of Lateral and Medial Giant
Fiber Conduction Velocity from all worms in your section.
· Fig 3 shows averaged spike data in the form of box-plot for all
warms for LGF and MGF, including standard deviation. Paired
t-test p value with an alpha level of 0.05 should be indicated in
the figure between the two groups.
· Table 2 shows t-test statistic and p-values between the two
groups.
Discussion
Bibliography
29. Long Lab Report Grading Checklist
This checklist may be used to double-check your lab report
before turning it in for grading. Point values for each item are
given. The instructor reserves the right to make minor
adjustments to the list before grading the report.
TITLE (10) – informative & concise
ABSTRACT (20)
____ The abstract contains a summary of the introduction. (6)
____ The abstract provides an overview of the method. (4)
____ The abstract includes the hypothesis and whether the
hypothesis was supported by the data. (6)
___ The abstract is no more than 200 words. (4)
INTRODUCTION (30)
_____ There is only one Introduction section in the report.
There are no other subtitles in this section. (6)
_____ The introduction provides the reader with enough
information to know what the experiment is about and why it is
being conducted. Sufficient information is provided that the
reader can understand the rest of the lab report. The writer also
explains why the hypotheses are being proposed, providing
justification for them. (14)
_____ The hypothesis is stated and is clearly identified as
the hypothesis that will be tested in the experiment. For
example, "the hypothesis tested is..." clearly identifies the
hypothesis. The hypothesis is appropriate for the observation
being studied. The hypothesis is stated as a sentence that is
either true or false. If the laboratory exercise has more than one
hypothesis, they should all be stated. (10)
METHODS (30)
_____ This section contains only methods. There are no
results or discussion in this section. (4)
_____ This section provides the reader with enough
information that the reader could repeat the experiment. The
writer uses his/her own words to write this section. (16)
_____ This section is written using whole sentences and
30. paragraphs. Outline format, numbering, “cookbook”
instructions, directions, or commands should not be used. (6)
_____ Avoid using the words I, he, she, or we. This section
should be written in third person and there should not be
references to other people. This section should be written in
past tense. Example: "After the milk was added, three drops of
rennin were added to tube #1." (4)
RESULTS (40)
_____ Only results are found in this section, nothing else.
(4)
_____ This section is written using sentences and
paragraphs. Any tables and figures in this section should be
summarized using sentences and paragraphs in a narrative. (4)
_____ All of the results are included in this section. The
results are written clearly and are easy to understand. (4)
_____ Descriptive statistics (e.g. mean, median, mode,
standard deviation) are provided where appropriate. (2)
_____ At least one table is included. The table should be
embedded within the Results section, not included on a separate
page. (4)
_____ Each table has a caption that briefly describes the
table. Example - Table 1. Reaction time 30 minutes after
swimming 1000 meters at 8:00 AM. Be sure to provide enough
information in the caption so that the reader can understand the
table without reading the paper. Tables are labeled Table 1,
Table 2, etc. At least one sentence in the Results section refers
to the table(s). For example, "The reaction time decreased after
swimming 1000 meters (Table 1)." (2)
____ There is a figure that plots the mean LGN and MGN
velocity for each worm. (4)
____ There is a figure that compares the overall LGN and MGN
velocities. (4)
_____ Figures have captions that briefly describe the
figures. Be sure to provide enough information in the caption so
that the reader can understand the figure without reading the
paper. Graphs and other figures are labeled Figure 1, Figure 2,
31. etc. At least one sentence in the Results section refers to the
figure(s). For example, "The mean reaction times of the two
groups differed (Figure 1)." (2)
_____ The axes of graphs are labeled correctly. The
independent variable should be on the x-axis and the dependent
variable is on the y-axis. Please see the notes on variables if
you have any questions. (4)
_____ Tables and figures are placed in the Results section
near where they are discussed, not at the end of the report. The
tables are not printed on a separate page. Instead, they are
inserted into the document. (2)
_____ The probability level (p-value) for statistical tests is
stated. If a t-test is used, the value of the t-statistic and degrees
of freedom (d.f.) are also stated. All of these statistics should be
stated in sentence form. (4)
DISCUSSION (40)
_____ This section states whether you accept or reject the
hypothesis or hypotheses. (6)
_____ If a hypothesis is rejected, an alternative hypothesis
is given. (4)
_____ Acceptance or rejection of the hypothesis is
supported by the data and statistical analysis. (4)
_____ Adequate interpretation of the data is provided.
Unexpected results are discussed adequately. (16)
_____ Limitations to the experiment & dataset and possible
future research are mentioned. (10)
REFERENCES (20)
_____ APA format is used for references. (6)
_____In-text citations are used. (4)
_____ Cited literature is appropriate. (10)
OTHER ITEMS (10)
_____ Point deduction if direct quotations were used and all
citations must be reworded in the author’s own words. (4)
_____ The writer understands that hypotheses are not
proven; they are accepted or rejected. (2)
32. _____ Complete sentences were used throughout. English is
used correctly; grammatical errors are minimal. A spelling
checker was used; there are no spelling errors or typographical
errors. (4)
Fall2020_CondVeloDataMGNLGNWorm
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1114.4110.1114.3310.414.5610.6MEANSD
![Background
You probably think the nervous system is pretty fast. You seem
to hear the spikes immediately when you touch the leg of the
cockroach or blow on the cerci of the crickets. But is it
instantaneous? Of course not! Not even light, the fastest signal
in the universe, travels instantaneously. But how fast is a
nervous system? Is it faster than a car, faster than a jetplane, or
faster than a cell phone? And how can we measure it?
To measure speed (velocity) though, you need to measure both
time (when a spike occurred) and distance (how far a spike has
travelled down a nerve).
Take the analogy of a car on a highway. If you were looking out
of a small observation hut by the road, you could tell what
whether you saw a car, what kind of car it was, and the time
that you saw it.
Similarly with your SpikerBox, you can currently tell if you saw
a spike, perhaps what kind of neuron generated that spike (we
will discuss this in a later experiment), and the time you saw
the spike, but you can’t tell how fast the spike was travelling
down the nerve.
Let’s go back to the car on the highway. Suppose you had a
friend ½ mile down the road in a similar hut:
Later, you two could compare notes to determine the speed of
the car. 1 minute = 0.016 hours. Dividing ½ mile by 0.016 hours
= 31.25 mph.
[illustration]
So, why don’t we take our two-channel SpikerBox with our two](data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7)