Brain and Behavior Student Manual 2022

College Core Curriculum
FOUNDATIONS OF SCIENTIFIC INQUIRY

LIFE SCIENCE: CORE-UA 306

BRAIN AND BEHAVIOR

FALL 2022
Professor André Fenton

LABORATORY MANUAL

Copyright © 2022 by the College Core Curriculum, Foundations of Scientific Inquiry.
All rights reserved. No part of this publication may be reproduced, stored in a retrieval system,
or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, or
otherwise, without the prior written permission of the publisher.

TABLE OF CONTENTS

Laboratory Development and Acknowledgments . . . . ii

Laboratory Policies and Practice iii
iv
Laboratory Policies . . . . . . . . xi

Laboratory Safety Rules . . . . . . . . 1
17
Guidelines for Good Laboratory Practices . . . . . 43
61
Laboratory Experiments and Exercises 91
115
1. The Scientific Method ....... 135

2. Electrical Potentials in Neurons I: A Model Dendrite ... 161
185
3. Electrical Potentials in Neurons II: Action Potentials in an Axon . .

4. Recording Action Potentials from the Model Organism Periplanteta americana (L) .

5. Organization of the Brain I: Microscopy – Neuron to Brain . . .

6. Organization of the Brain II: Dissection of the Sheep Brain . . .

7. Vision . . . . . . . . . .

8. Observing Behavior: . .
Learning and Memory with the Model Organism Caenorhabditis elegans .

9. Genetically Manipulating Behavior of C. elegans . . . .

Appendix

Statistical Appendix . . . . . . . . . 203

i

Laboratory Development and Acknowledgments

This laboratory manual has been compiled from experiments and exercises developed and
written by the following professors and laboratory staff at New York University:

Dr. Elizabeth Bauer
Dr. Aisling Dunne
Dr. André Fenton
Dr. Paul Glimcher
Dr. Mick Hawken
Dr. Trace Jordan
Dr. Lynne Kiorpes
Dr. Monica Lewin
Dr. Wendy Suzuki

John Augello
Emily Cowan
Ashley Kopec
Kathleen Jabara
Ali Jahromi
Susan Price
Robyn Schare
Kelli Kapri-Tate

Special thanks to the other laboratory instructors of this course for their insightful
suggestions.

ii

Laboratory Policies

Each laboratory project will be worth 50 points with the following allocations:

Attendance 10 points
Quiz 10 points
Lab Project 30 points

Attendance

You are expected to arrive punctually for the laboratory session. Arriving more than 10 minutes late

will result in a loss of attendance points for the laboratory session, and you will miss the quiz.
There are NO makeups.

Quiz

Students will take a short quiz of 4 to 6 questions in Brightspace, available to them a few days before
the lab session. Quiz questions are general based upon the introduction section of the lab.

Lab Assignments

The laboratory assignments must be completed and submitted at the end of the laboratory session.
Laboratory homework assignments must be handed in the following week.

Absence Policy

Excused absences will only be given in the case of illness (with a doctor’s note) or observation of a
religious holiday. All other absences will be considered unexcused and will result in a lab grade of
zero. More than three absences will result in an automatic failure of the laboratory portion of
this course.

Late Assignments

Late assignments will be penalized five points for each day late (excluding weekends). You must make
prior arrangements with you laboratory instructor to submit late work.

iii

Laboratory Safety Agreement
Brain and Behavior

Core Curriculum Labs

Lab class and section: _______________________________________________

Instructor: ________________________________________________

All students must be aware that potential hazards exist in a biology lab. To ensure that your experience
here is safe and productive, the following rules need to be followed. Keep in mind that some labs may
require lab specific safety practices.

General Laboratory Safety Guidelines

• You may not enter the laboratory until you instructor is present and instructs you to enter the
room. Please remain in the main hallway outside the double doors until you are given
permission to enter.

• NO FOOD OR DRINK (INCLUDING BOTTLED WATER) IS ALLOWED IN THE
LAB. CHEWING GUM IS PROHIBITED. THERE ARE NO EXCEPTIONS TO THIS
RULE.

• Do not place coats or bags on the laboratory benches or on the floor as they serve as a tripping
hazard. Please store out of the way or under the lab bench.

• Keep hands away from face, eyes, and mouth while in lab. Wash your hands with soap and
water after performing all experiments and before leaving the laboratory.

• Keep your work area clean. Inform your instructor immediately if you have any spills.

• Horseplay and other acts of mischief in the laboratory is unacceptable behavior and is cause for
immediate removal from the laboratory.

• Always follow the procedures as outlined in your lab manual. Your lab instructor will
communicate any modifications that need to be made. Unauthorized experiments or procedures
must not be attempted.

• Do not alter the laboratory set-up. Ask your instructor if you think something is missing or out
of place.

• Proper handling of equipment, chemicals, animal tissue, and animals in an experiment is your
responsibility. WHEN IN DOUBT, ASK YOUR INSTRUCTOR.

• Please turn your cell phone ringer off before entering the lab. You should not be using your cell
phone during the laboratory session, and it should not be kept on the laboratory bench. Since
hearing is important for learning and to avoid accidents, radios, mp3 players and headphones of
any kind may not be used in the lab.

• All electrical equipment must be turned off and unplugged before leaving the laboratory.

• YOUR WORK AREA MUST BE CLEAN BEFORE YOU LEAVE! POINTS WILL BE
DEDUCTED FROM YOUR LABORATORY IF YOU DON’T FOLLOW THE CLEAN
UP INSTRUCTIONS.

iv

Clothing

• Dress appropriately for the lab. Shorts (regardless of length), short skirts, sleeveless shirts,
tank tops, halter tops, open-backed shirts, torso, and midriff exposing tops are not
considered safe attire and are not permitted. Instead, long sleeved tops and long pants or
long skirts should be worn to protect your skin from spills.

• Shoes that completely cover the foot must be worn during the entire time that you are in the
lab to protect you from splashes of chemicals, broken glass, and other hazards that may
occur. The shoes must cover your feet without gaps. Flip-flops, ballerina pumps, perforated
shoes and sandals (even if socks are worn) are never permitted in the lab.

• Long hair, dangling jewelry, and loose or baggy clothing are a hazard in the laboratory. Long
hair must be tied back, and dangling jewelry and baggy clothing must be secured. Dangling
jewelry can become entangled in equipment and can conduct electricity.

• You must wear goggles and gloves when instructed.
• Do not handle contact lenses in the lab. Application of cosmetics including lip balm is not

permitted in the lab.

v

Broken Glass

• If something is falling, let it drop! Catching it may cause the glassware to break in your hand.
• Any broken glass must be cleaned up immediately. The instructor will direct the clean-up of

broken glass. Everyone in the area must follow the instructor’s directions.
⋅ Do not touch broken glass and do not attempt to clean broken glassware yourself
unless instructed to do so.
⋅ Clean up broken glass using a dustpan and brush.
⋅ Small shards can be picked up using a wet paper towel or absorbent pad.
⋅ Broken glass must be placed in the box labeled “Broken Glass”.

• DO NOT overload glassware disposal containers.
• ONLY UNCONTAMINATED broken glass may be disposed of in a broken glass box.

Heating Chemicals

• Never leave a hot plate unattended. Check for the cord, it should NOT touch the hot plate
surface. Chemicals spilled on hot plates can result in fire.

• Never look down into a container being heated.
• Never heat substances in sealed containers.
• Allow hot glassware to cool before handling. If you need to handle hot glassware-be cautious,

use tongs and direct the opening of the containers away from yourself and others.
• Do not immerse hot glassware in cold water. The glassware may shatter.

Handling Chemicals

• Treat all chemicals in the lab as toxic substances.
• Always check the labels of all the chemicals you use. Know the name of, and hazards

presented by, any chemical involved in an experiment BEFORE you use it.
• Do not use reagent containers without labels or with improper labels. Report these situations

to your instructor.
• NEVER TOUCH, TASTE OR SMELL CHEMICALS.
• Never leave excess or spilled chemicals on equipment (in particular, the handling surface of

glassware); wipe clean with a damp towel immediately and dry immediately with another
towel.
• Do not use your fingers as a stopper for a tube or a bottle when shaking it to mix the contents.
• Hold stoppers and droppers while using a container. Do not put stoppers on bench or plastic
surfaces.
• Replace stoppers, lids, and droppers immediately after use to prevent contamination of
reagents.
• Dispose of chemical and biohazard waste as instructed.

vi

GHS Pictograms and Hazard Classes

• Oxidizers • Flammables • Explosives
• Self Reactives • Self Reactives
• Pyrophorics • Organic Peroxides
• Self-Heating
• Emits Flammable Gas
• Organic Peroxides

• Acute toxicity (severe) • Corrosives • Gases Under Pressure

• Carcinogen • Environmental • Irritant
• Respiratory Sensitizer Toxicity • Dermal Sensitizer
• Reproductive Toxicity • Acute toxicity (harmful)
• Target Organ Toxicity • Narcotic Effects
• Mutagenicity • Respiratory Tract
• Aspiration Toxicity
Irritation

vii

Electrical Safety

• Please exercise caution when dealing with electrical devices.
• If equipment is not working, turn off the power to equipment and inform the instructor.
• Maintain a workspace clear of extra material such as books, papers, and clothes.
• Avoid contacting electrical equipment with wet hands or wet materials.

Dissection Safety

• Pointed dissection probes, scalpels, razor blades, and scissors must be used with great care,
and placed in a safe position when not in use.

• Containers designated for the disposal of sharps (scalpel blades, razor blades, dissection pins,
etc.) are present in each laboratory. Never dispose of any sharp object in the regular trash
containers.

• When cutting with a scalpel or other sharp instrument, forceps may be used to help hold the
specimen. Never use fingers to hold a part of the specimen while cutting.

• Scalpels and other sharp instruments are only to be used to make cuts in the specimen, never
as a probe or a pointer.

• Always cut away from the body and away from other people. Reposition the specimen if
necessary.

In Case of Emergency

1. Identify the location of all exits from the laboratory and from the building.
2. Be familiar with the location and proper use of fire extinguishers, fire blankets, first aid kits,

spill response kits, and eye wash stations in each laboratory.
3. If a chemical, or any substance, should splash in your eye(s) or on your skin, immediately

flush with running water for at least 20 minutes.
4. If you or your lab partner is hurt, immediately report it to the instructor.

In the Event of a Fire or Fire Drill

1. Shut off all equipment (hot plates, microscopes etc.).
2. Take all your belongings and proceed to the nearest exit.
3. Do not use the elevators.

viii

PLEASE SIGN THIS FORM AND RETURN IT TO YOUR INSTRUCTOR.

Failure to observe laboratory safety rules and procedures may result in injury to you or to fellow
students. If you are found to be in violation of these rules, you will be asked to leave the laboratory
and lose the grade for the lab, at the discretion of the instructor.

By signing this I, , understand and agree to follow all the

(Students Name)

safety procedures described above as well as any other written or verbal instruction provided by my

instructor.

Signature Date

Course: Brain and Behavior
Day:

 Thursday
 Friday
Time:
 9:30 AM– 10:45 AM
 11:00 AM – 12:15 PM
 12:30 PM – 1:45 PM
 2:00 PM – 3:15 PM
 3:30 PM – 4:45 PM
 4:55 PM – 6:10 PM

ix

x

Guidelines for Good Laboratory Practices

Good practices and procedures in the laboratory are essential for the success of any experiment.
These include familiarizing yourself with laboratory equipment and chemicals, performing
experiments correctly in order to minimize errors, following safety procedures, and ensuring correct
disposal of all materials. Read the following guidelines carefully and refer to them throughout the
duration of the semester. Always read instructions carefully and thoroughly. You can ask your
laboratory instructor for assistance if you are still in doubt about a certain procedure.

GENERAL

• Each experiment has a unique set of directions! Always double check that you are following the
proper procedure.

• Use the metric system for all measurements, unless otherwise stated.

• To ensure precision in your measurements, you should repeat them whenever time allows.

SAFETY

• Wear goggles and gloves whenever necessary.

• If you should get a chemical in your eye or on your skin, rinse the affected area immediately. For
basic (alkali) or acidic substances, rinse for at least 15 minutes. Inform your lab instructor
immediately.

• Be familiar with the safety procedures associated with each experiment.

• Immediately replace bottle caps and covers to containers.

• Attend to all spills immediately.

CHEMICAL USE AND DISPOSAL

• Carefully read all labels and use only the reagents designated for a particular procedure.

• LABEL EVERYTHING! Many chemical solutions look like water and you cannot differentiate them
unless you label test beakers and/or tubes.

• NEVER RETURN EXCESS REAGENTS TO THE SUPPLY BOTTLES. Do not take more of a chemical than
is necessary.

• In order to avoid cross contamination of chemicals, make sure that any pipette you use is new or
has been used for the same chemical only. The plastic pipettes are disposable, and if you are in
doubt, throw away the pipette.

xi

• Always place chemical waste into the appropriate receptacle. Do not dispose of any chemical
used in the laboratory without receiving and understanding the directions given by your
instructor.

GLASSWARE

• Before using glassware, ensure its cleanliness.
• During the semester, you will be using a variety of glassware that may be unfamiliar to you.

Please refer to the pictures below if you are unsure about the glassware you are using.

• Erlenmeyer flasks and beakers should only be used for measuring approximate volume. The
graduations on these are only guides to their capacity.

• Special glassware and pipettes are available for measuring exact volume. A graduated cylinder
should be used volume measurements. Finnpipettes should be used to dispense small volumes.

xii

• The volume of a liquid is measured by reading the level at the bottom of the meniscus. The
meniscus is the curved surface formed by the liquid adhering to the walls of the glass. Reading
the meniscus is easier and more accurate if viewed at eye level as in the illustration below.

xiii

xiv

THE SCIENTIFIC METHOD

LABORATORY 1

THE SCIENTIFIC METHOD

SUMMARY
A central theme in this course will be the assessment of scientific information. To make informed
judgments about the validity of scientific studies, it is necessary to understand the process of
scientific investigation. It is likely that you have encountered some relevant terms before -
hypothesis, control group, etc. - but during this course, you will deepen your understanding of
how these general principles are applied to specific studies of the brain and behavior. In this first
laboratory session, you will engage in a simple experiment on working memory to illustrate the
scientific method.

OBJECTIVES

• To define the bolded key terms in this chapter.
• To understand how to use the scientific process to ask questions.
• To gain an appreciation for experimental design, interpreting data, and drawing

conclusions from your experiment.

 The Scientific Method

The ability to remember names, objects, telephone numbers, events, etc. seems to vary widely
among individuals. Some people have difficultly remembering the name of just one person they
meet, whereas other people seem to effortlessly recall the names of a whole group of new
acquaintances. Memories can be classified broadly as short-term memories, which last minutes
to hours, and long-term memories, which can last days, months, even a lifetime. Memory is still
a mystery in many ways and has inspired scientists for many years to apply the scientific method
to their questions to better understand how memories are formed and maintained in the brain.
We will be returning to the topic of memory later in the course.
The scientific method can be described in a series of steps:

1. General Observation
2. Develop a Hypothesis
3. Test the Hypothesis with an Experiment
4. Interpret the Results and form Conclusions
5. Revisit and Revise the Hypothesis

1

THE SCIENTIFIC METHOD

After you have revisited and revised your hypothesis based on the results of your experiment,
you can design a new experiment to test your new hypothesis, and so on. The exercises and
questions in today’s lab aim to illustrate this process.

 General Observation

The scientific method begins when you make a general observation that leads to questions.
Today in lab, we will use ourselves as subjects to perform an experiment to understand more
about our working memory. Working memory is usually defined as a type of short-term memory,
specifically for information that can be utilized immediately. For example, when you try to
remember a friend’s telephone number, we can rely on working memory to help us dial the
number, but the memory does not seem to persist beyond this task. We want to understand
more about the parameters of our working memory’s ability to store and accurately recall
information. Do we have an infinite ability to remember and immediately recall information?
Could we design an experiment to tell us more about the capacity of our working memory?

 Hypothesis

The word hypothesis derives from the root words “hypo” (under, beneath) and “thesis” (an
arranging). A hypothesis is an educated guess used to provide a tentative explanation of an
observation or phenomenon. We then design an experiment, which should include a set of steps
(i.e., the scientific method) that tests our hypothesis. A hypothesis is only useful if it can be tested
in a scientific manner, and pre-existing hypotheses are constantly being revised as experiments
reveal new information.

By gathering data with experimental tests, we can challenge our hypothesis, either by lending
support to or refuting our tentative explanation of the observation. It is rare, however, that a
hypothesis is accepted or rejected based on a single, so-called “crucial experiment.” It usually
takes many experiments and much discussion among scientists for a hypothesis to gain gradual
support.

In today’s experiment, you will be exploring the capacity of working memory. During the
laboratory experiment you will be shown an 8-digit number followed immediately by a set of 20
images. Your job is to remember the number and as many images as you can. You must first and
foremost accurately remember the number. If you are unable to correctly recall the number,
your data will not be included in the group data analysis. After a recall period, you will be shown
a 2-digit number followed immediately by a second set of 20 different images. Again, your job is
to correctly recall the number and as many images as you can.

Our general observation is that information in our working memory tends to not be stored for
long periods of time. In our experiment to explore the parameters of working memory, we will
ask the specific question: Do we have an infinite ability to remember and immediately recall
information?

2

THE SCIENTIFIC METHOD

Brain & Behavior: CORE-UA 306

LABORATORY COVER SHEET
Name: _____________________________ Lab Partner’s Name: _________________________
Date of Lab: ___________________________ Date Due: _______________________________
Lab Instructor: _____________________________ Lab Section Number: __________________

3

THE SCIENTIFIC METHOD

Procedures and Exercises

Now that we have some background information on the nature of working memory, let’s focus
on our specific experiment. We will be recording the number of pictures in the “8” condition
versus the number of pictures recalled in the “2” condition as our data.
QUESTION 1:
There are three possible outcomes of this experiment related to the number of pictures recalled
in each condition.
A. In your groups, write down the three possible outcomes.
B. Which outcome do you predict will occur?
C. Why?
A.

B.

C.

With your group, discuss your own experiences to come up with a general hypothesis for working
memory as it relates to this experiment. Remember that a hypothesis is an educated guess; it is
okay if you are not correct in your predictions.

4

THE SCIENTIFIC METHOD

 Testing the Hypothesis

Now that we have a hypothesis regarding the capacity of working memory, we must design an
experiment to test the hypothesis. Experimental design is an art and can take a lot of time and
planning, so we have designed an experiment in advance, and you have already identified the
potential outcomes of the experiment. Here are 2 key points to remember about experimental
design:

1. In many experiments, you want to have an experimental group that receives some sort of
manipulation, and a control group that is treated the same except for that one manipulation.
For instance, if you are giving someone a drug and then asking if that drug enhances memory,
that subject is in the experimental condition because he/she is receiving a manipulation (the
drug). The control subject would then be given a placebo, e.g., a sugar pill that has no effect,
but he/she would be given the same memory tests. Everything is the same, except one person
gets active drug and one person gets ‘pretend’ drug, thus allowing us to determine the effect
of this specific manipulation on memory.

2. Randomization of subjects into either the experimental group or the control group is very
important. If, as we introduced at the beginning of the lab, a person’s innate ability to
remember items is above average or below average, randomly assigning people into each
group will help to create a ‘balance’ of baseline abilities. Randomization will also prevent the
introduction of any additional variables that could contribute to differences in the conditions,
for example, if some subjects are always run first in the morning but others at night,
randomizing group assignment would ensure these variables would be equally spread across
the two conditions.

 Data Collection

Now, we are ready to perform our experiment!
You must record your data on the next page titled DATA SHEET.

SPECIAL NOTES:
 Your data will only be counted and included in the group analysis IF you can correctly
recall every digit of each number flashed. This is to ensure that we are truly seeing the
effects of memory for the number on working memory capacity for the subsequent
images.
 Your instructor will collect your data sheet and compile the class’ data for group analysis.

5

THE SCIENTIFIC METHOD

…

6

THE SCIENTIFIC METHOD

DATA SHEET

Condition 1: 8-digit number + image set NUMBER: ___________________

Condition 2: 2-digit number + image set NUMBER: ___________________

7

THE SCIENTIFIC METHOD

Record and report the following data to your instructor.

• Condition 1: 8-digit number + picture set

Was your 8-digit number correct? Circle your answer: Y / N

Number of images correctly recalled: __________

• Condition 2: 2-digit number + picture set

Was your 2-digit number correct? Circle your answer: Y / N

Number of images correctly recalled: __________

You will tear out this DATA SHEET and hand it in to your laboratory instructor.
While the group data is being analyzed and figures are being created for discussion, please
review the information in the next section.

8

THE SCIENTIFIC METHOD

 Analyzing the Results

QUESTION 2:
Your instructor has analyzed the data of the class using the three different methods.
Reproduce the figures on the following graphs provided (they do not have to be perfect):

A. Frequency Distribution: Number of Images Recalled
B. Averages Histogram: Number of Images Recalled
C. Frequency Distribution: Difference Scores

A. Frequency Distribution:
Number of Images Recalled

Number of Students 20 "2-digit" Students
18 "8-digit" Students
16
14
12
10

8
6
4
2
0

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21

N = Number Images Recalled

B. Averages Histogram:
Number of Images Recalled
COLOR KEY:
Number of Images Recalled 20 Fill in the below circles with 2
19 different colors to represent
18 the 2 different groups of
17 students.
16
15 = ‘2-digit’ students
14 = ‘8-digit’ students
13
12 9
11
10

9
8
7
6
5
4
3
2
1
0

"2" "8"

Condition

Frequency THE SCIENTIFIC METHOD

Frequency Distribution:
C. Difference Scores

-10 -9 -8 -7 -6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6 7 8 9 10
Difference Score

Now that we have performed the experiment it is time to analyze the data. In our case, we are
going to examine the number of images recalled in the eight-digit condition and the two-digit
condition. We need to understand what the data is telling us so we can make conclusions and re-
evaluate our hypothesis. Most scientists will first calculate a measure of central tendency, like an
average, and a measure of variance, such as a standard deviation.
Many of you are probably familiar with the average or mean of the scores. To calculate the mean,
you add up all the data and divide by the total number of data points. Often in a data set you will
encounter a result that is well outside the expected range. This result is known as an outlier.
Because experiments often encounter unexpected results, it is important to not rely only on the
mean but also to describe the variance or standard deviation of the scores. These measurements
are based on the difference between each score in the data set and the mean value. Variance
and standard deviation tell us how spread out the data points are from the mean. You can find
the equation for calculating the standard deviation in the Statistical Appendix at the back of your
lab manual. For the sake of brevity, variance is simply the square of the standard deviation.
Figure 1 shows the comparison of two theoretical distributions with the same number of
measurements (n) and the same mean (x)̄ , but with different values of standard deviation (s). In
one case the standard deviation is small, so the distribution appears as a tall and narrow peak.
This reflects the fact that almost all the data values are very close to the mean and there is little
variability in the sample. In the other case, however, the distribution is short and spread out. This
tells us that only a small number of the data points are close to the mean, which produces a large
standard deviation and a lot of variability.

10

THE SCIENTIFIC METHOD

Figure 1: Comparison of two distributions with the same mean (x)̄ , but different values of standard deviation.
Experimental data is often summarized in the form: mean ± standard deviation.

Another way to depict data is by calculating difference scores. A difference score is the difference
between a subject’s performance before a manipulation (the baseline) and the performance
after a manipulation. In our case, we can calculate the difference between your two-digit
condition score and your eight-digit condition score. This is an important way to describe data
because it considers that there will be individual variability in baseline performance. Some people
might remember 10 pictures accurately at baseline, while others will remember 5 at baseline.
One last method of describing data that we will look at in this lab is as a frequency distribution.
A frequency distribution shows how many subjects had a certain score. For example, if 3 students
in the class had a score of 5 pictures in the two-digit condition, then under the score value of 5,
the corresponding height of the bar would be 3 to represent the 3 students who obtained that
score.

It is important to point out that not all methods of depicting data are going to be useful for all
data sets. You must determine the most meaningful and clear way of depicting your data.

 t-Tests and p-Values
When we have data from more than one condition, like a control group and an experimental
group, or in our case, the eight-digit condition and the two-digit condition, there is a
mathematical way to determine if there are meaningful differences between the groups. To do
this, we employ statistical tests. A very common statistical test for two groups is called a t-test.
In short, the t-test tells us the degree of likelihood that the difference we observe between two
groups is a real phenomenon and is not caused purely by chance.

By convention, we consider groups that demonstrate a statistical significance, as expressed by
the p-value (or ‘probability value’) less than 0.05 (written as p < 0.05) to be significantly different
from each other. This would mean that there is a difference between the groups that is not likely
due to chance, and potentially due to your experimental manipulation. The number 0.05
indicates that there is a 5% (or less) chance that the differences we see between two groups are
due to chance, if our hypothesis is true.

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THE SCIENTIFIC METHOD

For more information on p-value, t-test, and other statistics terminology, please consult the
Statistical Appendix at the end of this manual to assist you in answering the remainder of
exercise questions.
QUESTION 3:
Take a closer look at each graph. Briefly describe what each graph is depicting and discuss
whether the depiction is useful with your class data set.

QUESTION 4:
If you determined a p-value, what did it indicate about your data?

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THE SCIENTIFIC METHOD

 Interpretations, Conclusions and Future Directions

Once the data have been collected it is necessary to interpret the results of the experiment,
which can be more difficult than it first appears. We must assess whether differences can be
attributed to specific variables that we are testing, or whether our interpretation is complicated
by confounding variables that we have not controlled.
A confounding variable is an aspect of the experiment that isn’t controlled for or changed by the
experimenter that could by itself cause variations between experimental conditions. For
example, one confounding variable in our experiment is the amount of sleep each subject (each
student) got last night. That’s not something we controlled for (we would have had to monitor
each of you to make sure you all got precisely the same amount of sleep) but being very tired
could certainly affect your performance on this task. Experimenters try to keep the confounding
variables to a minimum, but no one can control everything!
Once you have results you can formulate basic conclusions about your experiment. It is important
to be conservative with your conclusions. Remember that it is rare to accept or reject a
hypothesis based on a single experiment. Typically, you will need to perform additional
experiments to analyze the repeatability and reproducibility of the experiment.
Repeatability refers to the variations in data taken by a single person using the same
experimental parameters and conditions.
Reproducibility refers to the ability to reproduce an entire experiment, regardless of the person.
After performing the memory experiment and developing conclusions, you should be able to
interpret your results and answer questions to guide future experiments. For instance, an
experimenter might consider what additional experiments could be performed to further
investigate the newly revised hypothesis. He/she would also consider if in future experiments
there should be any methodological changes or confounding variables that could be controlled.

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THE SCIENTIFIC METHOD

CONCLUSION QUESTIONS

QUESTION 5:
A. Are you able to make any basic conclusions based off your results and data analysis?
B. How do these conclusions relate to your original hypothesis?

QUESTION 6:
A. Can we say that the data and conclusions from our experiment are indisputable facts?
B. Why or why not?

QUESTION 7:
Remember, no experiment is perfect.
A. Can you identify any confounding variables (other than sleep, the example given to you)
present in the current version of the experiment?
B. How would you change the experiment if you had to do it again?

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THE SCIENTIFIC METHOD

CLEAN-UP PROCEDURES

When you are done with your work, you must clean up. You will not be permitted to leave the
lab until the lab instructor checks your lab bench. Failure to properly clean-up will result in a 5-point
deduction from your laboratory assignment.

 Return all colored pencils to their container.
 Please leave your lab station as clean and orderly as when you arrived.
 Before leaving the lab, double-check your area for your personal belongings.

THE LAB STAFF IS NOT RESPONSIBLE FOR ANY LOST OR STOLEN ITEMS.
NOTE:
Pages 3 through 14 are to be turned in as a full assignment (without DATA SHEET, pp. 7-8).
If more writing space is needed, use pages 15 and 16 to complete your responses.

15

THE SCIENTIFIC METHOD

…

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

LABORATORY 2

ELECTRICAL POTENTIALS IN NEURONS I:
A Model Dendrite

SUMMARY
All behaviors and cognitive functions are processed as signals between neurons.
Neurons communicate through both chemical signals via neurotransmitters, and electrical
signals as electrical currents resulting from the movement of charged molecules called ions.
In today’s lab, we will take a closer look at the properties of electrical currents. We will build a
model dendrite to examine some of the passive electrical properties of neurons. We will first
study how the electrical current created by a synaptic input will experience losses in strength
with distance traveled down the dendrite. We will calculate the length constant, which is the
distance at which the current has lost two-thirds of its original strength. We will explore how the
length constant is dependent on both the diameter of the neuron’s dendrite and the resistance
of its membrane. In the second part of the experiment we will simulate the effects of synaptic
summation. Lastly, we will explore the idea of a ‘threshold’ gating the conversion of passive
conduction into active conduction.
OBJECTIVES

• Understand the differences between passive conduction of local potentials and active
conduction of action potentials.

• Understand what a length constant is, how it is calculated, and how properties of a
neuron can modulate the length constant.

• Understand how spatial summation relates to action potential thresholds.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

INTRODUCTION

 Active vs. Passive Conduction in Neurons

The resting membrane potential (Vm, or voltage of the membrane) of a neuron is defined as the
difference in voltage of the inside of the cell relative to the outside of the cell when there is no
net flow of ions. No net flow of ions means that for any ion that moves in, one moves out. The
membrane is at ‘rest.’ In the case of neurons, the inside of the cell is more negative relative to
the outside, which is why Vm is a negative number. However, if a stimulus causes ion channels on
the membrane to open, ions are then able to rush into or out of the cell, resulting in a current
that can make the membrane potential more positive or negative. This conduction can be active
or passive.

• Active conduction refers to a change in the membrane potential that is actively regenerated,
and therefore can be maintained over time and space. A good example of active conduction
is the action potential. The action potential is a dramatic change in the voltage across the
membrane (the membrane potential) that follows a stereotyped pattern as it travels down
the axon.

• Passive conduction refers to a change in the membrane potential that is localized to the
stimulus source and then dissipates over time and space. The resulting change in the
membrane potential is temporary and dependent upon the strength of the initial stimulus.
Local, graded potentials are a good example of passive conduction, and are the focus of
today’s lab.

 Passive Conduction: Local, Graded Potentials

A local, graded potential is the result of a synaptic input which occurs when one neuron uses
neurotransmitters to communicate with another neuron. For instance, if the synaptic input
(neurotransmitters released, which bind to receptors) causes Na+ channels to open on the
dendrite of the post-synaptic cell, Na+ would rush into the cell. Because Na+ is highly concentrated
outside the cell, it will want to move down its concentration gradient.

This flow of ions across the membrane creates a ‘positive’ or depolarizing current, which would
cause the membrane potential to become more positive. This positive change in voltage (of the
membrane potential) is called an excitatory post-synaptic potential, or EPSP. It is considered
excitatory because the depolarization of the membrane potential would bring the neuron closer
to its threshold for discharging an action potential.

The threshold for an action potential is the membrane potential (voltage) at which voltage-gated
Na+ channels will open, thus initiating an action potential due to the rapid influx of Na+. Each type
of neuron may have a different value of the threshold membrane potential, but will be more
positive than its resting potential.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

However, if the synaptic input causes K+ channels to open on the post-synaptic cell, K+ would
leave the cell (because K+ is highly concentrated inside the cell and will want to move down its
concentration gradient) creating a ‘negative’ or hyperpolarizing current that would cause the
membrane potential to become more negative. This is called an inhibitory post-synaptic
potential, or IPSP. It is considered inhibitory because the hyperpolarization of the membrane
potential would bring the neuron farther from its threshold for an action potential.

 Local, Graded Potentials “Dissipate” over Time and Space

The current and change in membrane potential resulting from some such synaptic input would
be localized to the area where the synaptic input had caused channels to open. Local, graded
potentials have two important properties: they are both temporally and spatially restricted. This
means that over time the changes in membrane potential will become smaller and smaller, and
eventually the membrane will return to its resting state. This property can be described
mathematically by calculating the time constant, or τ (tau). Furthermore, the change in
membrane potential (the change in voltage) caused by a synaptic input will be progressively
smaller as you measure it farther and farther away from where the stimulus originated. This
property can be described mathematically by calculating the length constant, which we will do
today in lab.

 Calculating the Length Constant, λ

The length constant, λ (lambda), is defined as the distance from the source of the stimulus (e.g.
a synaptic input) at which that signal falls to 37% of its initial value. Thirty-seven percent is
derived from the reciprocal value of the mathematical constant called Euler’s constant or 1/e.
The length constant is calculated with the following equation:

λ = Rm
Ri + Ro

It’s important not to just plug in numbers, but to understand what this equation means in terms
of neurobiology. Each R variable stands for the resistance of different parts of the neuron (see
Figure 1), and it is measured in a unit called Ohms (Ω). For our purposes, the resistance of
something means the difficulty with which current (via ion flow) can move. A high resistance
means current flow across the subject is very difficult, and a low resistance means current flow
across the subject is relatively easy.

Rm refers to the resistance of the membrane. A cell’s membrane is a lipid bilayer that repels
charged and polar molecules due to its hydrophobic core. Unless there is a channel open, ions
won’t freely move through it (because they are charged). As such, the resistance across the
membrane is typically very high (i.e., it is hard for ions to flow through the membrane), though it
does vary from neuron to neuron.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

To change Rm, the membrane’s permeability to ions would need to be changed. For example,
opening many ion channels would decrease the Rm because there would be more holes, making
it easier for ions to move across the membrane. On the other hand, the addition of a layer of
insulation like myelin, will increase Rm by adding an extra layer of fortification to the membrane
which further restricts ion ‘leakage’ into or out of the neuron.
Ri refers to the internal resistance of the neuron, which can also be thought of as the
cytoplasmic resistance. The cytoplasm (‘stuff’ inside the neuron) can vary in the kinds and
amount of proteins it contains, depending on the type and location of the neuron. Therefore, Ri
is a measure of how hard it is for current to flow within the cytoplasm itself (think of it as trying
to move along the crowded internal material of the neuron). One determinant of Ri is the
diameter of the dendrite. If the dendrite is very narrow, current flow will be more difficult
through the cytoplasm. If the dendrite is very wide, current flow will be easy through the
cytoplasm.
Ro refers to the external resistance (the resistance outside the neuron). Compared to the
membrane and the cytoplasm, there is a very low resistance in the extracellular space (because
it’s so vast in comparison), so Ro is typically very small.

Figure 1:
Depiction of the three main forms of
resistance that act upon local, graded
potentials: outer, or extracellular resistance
(Ro), membrane resistance (Rm), and inner, or
intracellular resistance (Ri).

A useful way to think about these variables is to think of the neuron as a hose with water running
through it. If there are a lot of holes, the water will run out and may not make it to the end of the
hose. In this case, Rm - resistance across the membrane - would be very low, because water can
easily move through the many holes.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

But, if we add insulation (simulating myelin) and/or close the holes (i.e. channels), the water will
move quickly without many leaks, and Rm will be much higher. Using the same analogy, if the
hose is very narrow, it will be difficult for the water to move through it (a high Ri), but if the hose
has a wide diameter the water will flow easily (a low Ri). If we pick a location along the hose as
our source and insert a finite amount of water into the hose, Rm and Ri would determine how far
the water would travel. Similarly, with a single synaptic input causing an ionic current, Rm and Ri
will dictate how far down the dendrite the charge would travel.
Let’s look at the equation for the length constant again, and think about what the terms mean:

λ = Rm
Ri + Ro

The length constant will utilize a neuron’s measurements of resistance across the membrane
divided by the internal resistance (plus the usually negligible resistance outside the neuron) to
determine how far electrical potentials will travel.

 Summation of Local, Graded Potentials

Neurons can make hundreds, even thousands of connections with other neurons. As such, a
single neuron can receive many synaptic inputs at any given time. Together, these may provide
enough input to sufficiently change the membrane potential and reach threshold, leading to the
generation of an action potential. Neurons must therefore be able to account for all of the EPSPs
and IPSPs that it receives to determine if an action potential will be fired. You can think of this as
a summation of the synaptic inputs.

There are two different ways local, graded potentials can sum: spatially and temporally.

Spatial summation refers to the sum of local, graded potentials that arise from different physical
locations on the neuron. The synaptic inputs could be on different locations of the same dendrite,
on different dendrites, etc. Temporal summation refers to the sum of local, graded potentials
that arise from the same synapse in quick succession over time. It is important to note that when
we talk about “summing” up synaptic inputs, we must consider variables like the length constant.
In the case of spatial summation, a synaptic input close to where we are measuring the
membrane potential will cause a larger change than a synaptic input of equal strength that is
farther away. In lab, we will explore spatial summation in more detail.

 Simulating a Dendrite

Today we will simulate passive conduction on a dendrite using a breadboard that contains a
series of resistors. We will be injecting current into the breadboard using a battery. Resistors
resist current flow and voltage output from a stimulus source. The resistors on the breadboard
will simulate the three main conductive elements of the neuron; Ri, Rm, and Ro. Take a moment
to review Figures 2 and 3 to make sure you understand what our simulation will mean in
biological (neuronal) terms.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

22
Figure 2. We will be using 9-Volt batteries to simulate synaptic inputs and measuring the membrane potential (Vm) at different points along
the ‘model dendrite,’ or breadboard.

ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Voltage/Probe lead (Red) Common/Ground lead (Black)

9V Figure 3. Schematic diagram of breadboard.
BATTERY

I 100K O

Ri Rm Ro
Ro
Ri Rm Ro You will work with two different breadboards labeled as
Ro ‘100K’ and ‘30K’.
Ri Rm Ro
Below is summary of the different resistors present on
Ri Rm each breadboard.

• 100K Breadboard: resistor band colors
Ri = 100K = 100 x 103 ohms: brown, black, yellow, gold
Rm = 1M = 1 x 106 ohms: brown, black, green, gold
Ro = 10K = 10 x 103 ohms: brown, black orange, gold

• 30K Breadboard: resistor band colors
Ri = 30K = 30 x 103 ohms: orange, black, orange, gold
Rm = 1M = 1 x 106 ohms: brown, black, green, gold
Ro = 10K = 10 x 103 ohms: brown, black, orange, gold

Ri Rm

You will connect the 9V battery source to one end and measure the current after each resistor moving
away from the current source.

In terms of biology, the current from the 9V battery will be the change in membrane potential caused
by a single synaptic input. This current will flow down the breadboard (our “dendrite”) and as we
record the voltage at different locations farther away from the source, we are moving along the
dendrite, away from the synapse. While the amount of current injected will be the same from one
experiment to the next, as we change breadboards this current will encounter different degrees of
resistance.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Brain & Behavior: CORE-UA 306

LABORATORY COVER SHEET
Name: _____________________________ Lab Partner’s Name: _________________________
Date of Lab: ___________________________ Date Due: _______________________________
Lab Instructor: _____________________________ Lab Section Number: __________________

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

…

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Procedures and Exercises

General Notes

You will be working in pairs for this experiment. One student will be responsible for moving the
test leads to new recording positions and calling out the voltage values. The other student will
be responsible for accurately recording the data.

Practice Exercise

Prior to beginning the experiment, it is important that you understand how to use the digital
multimeter, or DMM. Complete this brief practice exercise before moving onto the main
experiments. Use the below diagram as a guide.

DIGITAL MULTIMETER TEST LEADS 9-VOLT BATTERY

DC 09. 1 6 Red Black

VDC Black

Red

COM V

1. Insert the black banana plug into the COM terminal of the DMM (solid line) and the red
banana plug into the V terminal of the DMM (dotted line).

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Each test lead contains a tiny, metal hook called a minigrabber which is used to clamp onto a
battery terminal or resistor when the wired end is pushed inward (see image below).
If either test lead is dysfunctional (i.e. missing metal hook, loose wires, misshaped lead), inform
your lab instructor immediately.

2. The 9V battery has two terminals: the hexagonal-shaped terminal is negative and the
circular-shaped terminal is positive. Connect the Common (or ground) test lead (black)
from the DMM to the battery’s negative terminal and the Voltage (or probe) test lead
(red) from the DMM to the battery’s positive terminal.

3. Turn the dial on the DMM to read VDC. The displayed output will be in volts (V); if your

DMM is not reading a value close to 9V, inform your lab instructor immediately.
4. What do you predict would happen if you switched the leads on the terminals?

Give it a try. Was your prediction correct?
You are now ready to begin the experiment.

Simulate a Synaptic Input: Connecting the Battery to the Model Dendrite

The battery provides current to the breadboard, delivering approximately 9V of current to the
point on the breadboard where it is connected. Refer to the schematic diagram in Figure 3 for
visual details. Using the 100K breadboard:

1. Connect the negative lead (black) from the 9V battery to the outside of the dendrite,
opposite the positive input. This is marked with the letter O (for outside).

2. Connect the positive lead (red) from the 9V battery to the “cytoplasm of the dendrite” at
one end of the breadboard. This is marked with the letter I (for inside).

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Measure the Current: Connecting the Digital Multimeter

The digital multimeter will measure the current at the location it is connected.
1. Since the banana plugs are already plugged in to your DMM, you can now connect their
terminal ends to the breadboard:
a. Connect the Com lead from the DMM to an Outside resistor of the model dendrite,
at the same end as the battery connection.
b. Connect the V lead from the DMM to an Inside resistor of the model dendrite, at the
same end as the battery connection
2. Verify that your DMM is displaying a voltage of approximately 9V. Inform your instructor
if your setup is not working properly. Once your setup is working properly, you can begin
the length constant experiment.

PART I: LENGTH CONSTANTS

During this first exercise you will experimentally determine the length constants of our synaptic
inputs (9V batteries) on the 100K and 30K breadboards. Recall the formula that determines the
length constant:

λ = Rm
Ri + Ro

QUESTION 1:
A. In theory, what would happen to the length constant if the Ri were increased?
B. What is a biological mechanism that would increase Rm?
C. If the length constant is low, what does that tell you about how far down the dendrite the
synaptic input will cause a change in the membrane potential?

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Make sure your 100K breadboard is set up like Figure 3. You are now ready to collect data.
You will keep the synaptic input (9V battery) stationary while moving the recording probe down
the dendrite to record the change in membrane potential resulting from the synaptic input at
each location.
QUESTION 2:
Write a hypothesis to describe what will happen as you move the recording probe away from the
synaptic input. Remember, a hypothesis includes an educated guess to explain a phenomenon,
so be sure to explain why you expect to see the changes you predict.

Your probe should be positioned at distance = 0.
Record the voltage at distance = 0 in DATA TABLE 1: Ri = 100K.
Next, move the probe (red lead) and the ground (black lead) of the DMM, in tandem, one
segment (one resistor) away from the battery connections. This measurement is at distance = 1.
Record the voltage in DATA TABLE 1: Ri = 100K.

NOTE: PLEASE BE VERY CAREFUL WHEN MOVING THE PROBES FROM RESISTOR
TO RESISTOR. IF A RESISTOR IS ACCIDENTLY REMOVED FROM THE BREADBOARD
PLEASE NOTIFY YOUR LAB INSTRUCTOR.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Continue recording the voltage at each segment (or distance) away from the battery along the
length of the dendrite. If you see a sudden jump from a low number, like 1 to a high number, like
800, the DMM has automatically switched from volts to millivolts, i.e. 1V to 800 mV, (equivalent
to 0.800V).

DATA TABLE 1: Ri = 100K

Distance Voltage
(# of resistors away
from the battery)

1
2
3
4
6
8
10
12
14
16
18
20
24

QUESTION 3:
Using the data above, determine the length constant of the 100K dendrite you derived
experimentally. The length constant is the distance (the number of segments or steps down the
breadboard) at which the voltage has declined to 37% of the original value. This voltage can be
determined by calculating 37% of the original value (distance=0) that you recorded.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

QUESTION 4:
When you move from the 100K breadboard to the 30K breadboard, what are you simulating in
terms of biology?

QUESTION 5:
Write a hypothesis about how the length constant measured on the 100K breadboard will
compare to the length constant for the 30K board. Why? Be sure to support your hypothesis.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

Now you will repeat the experiment using the 30K breadboard. Follow the instructions for
connecting the battery and digital multimeter, and then follow the steps outlined above to
measure the current as it travels down the resistors.

Record your data below in DATA TABLE 2: Ri = 30K.

DATA TABLE 2: Ri = 30K

Distance Voltage
(# of resistors away
from the battery)

1
2
3
4
6
8
10
12
14
16
18
20
24

QUESTION 6:
Using the data above, determine the length constant of the 30K dendrite you derived
experimentally.

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ELECTRICAL POTENTIALS IN NEURONS I: A Model Dendrite

QUESTION 7:
A. Use the length constant formula, as well as the resistance values for each breadboard to
calculate the theoretical length constants for each breadboard. How do they compare to the
values you derived experimentally?
B. Are they identical?
C. Why or why not?

λ = Rm
Ri + Ro

A. 100k Breadboard 30k Breadboard
Experimental length constant

Theoretical length constant

B. Are they identical?
C.

34