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Outline of Research Proposal

Introduction:

What topic will we do? Food and its effects on learning ability

Purpose: To justify whether broccoli will affect learning ability or not so that the result of the

experiment could be implemented in human later on

How? Two male hamsters will be used within this project. One will get normal food which is

sunflower seeds and grains (control group). While another will also have the same food with

broccoli would be added into its diet (experimental group). Both of the hamsters would go

through the same classic maze and would have their ability to learn determined.

Central question: Does consuming broccoli contribute to improvement in brain functioning,

learning, and memory?

Why is this experiment/research important?

Brain foods or foods that allegedly improves the ability to function of the brain are

becoming more and more popular alongside the health-conscious trend. Many companies are

incorporating extractions from those allegedly brain food into their products, especially

supplements. Broccoli is one of the brain food that, reportedly, will enhance brain power and

improve cognitive function due to the presence of vitamin K . Despite the popularity of brain

foods, there is no strong evidence that the alleged foods have any effects on brain functioning.

Hence, this experiment would be done in order to justify whetfher the hype about brain food is

true or not.

Topic of study related to the question?

Nutrition, brain, and learning

Nutrients found in broccoli and its possible effects on learning and memory
Hypothesis: If the hamster had been fed with broccoli, then it would make less mistakes, use

less time to learn how to run the maze correctly, and would use less time to reach the finishing

position in most of the trials.

Variables:

Independent variable: diet of the hamster

Dependent variable: the learning ability of each of the hamsters

Controlled variable:

Literature Review :

1) Significant nutrients on brain function omega-3 fatty acids and folic acid

Experiment = a drink rich of omega-3 fatty acid and other nutrients given to Australian

students

Result all students given a drink get high score in measuring verbal intelligence and

learning and memory test

2) Vitamin K and brain

Experiment = 954 participants, who have age averaged is 81, from the Memory and

Aging project reported their daily food and beverage intake by answering a detailed 144-item

questionnaire at the beginning of the study. Then, total daily nutrient is calculated. They

followed participants for 2 to 10 years assessing cognition annually with a comprehensive 19

tests and adjusted for age, sex, education, smoking, genetic risk for Alzheimers disease and

participation in physical activities when estimating the effects of diet on cognitive decline

Result = eating green leafy vegetables and other food rich in vitamin K, lutein and beta-

carotene can help to keep the brain healthy to preserve functioning.


5C :

1. Cite = focus on brain food --- What effect can each nutrients give?

2. Compare = they both did an experiment with human --- human and rodents are almost

the same in genetics plus a lot of the structure and connectivity that exists in human

brains also exists in rodents

3. Contrast = one proves for omega-3 fatty acid, another proves for vitamin K

4. Critique = the experiment with vitamin K is more persuasive because the experiment

was done by comparing people who consume vitamin K from food not directly which

is different from the experiment with omega-3 using a drink with pure omega-3 and not

directly get from food. In reality, people will consume nutrients from food not from

purifying it so the experiment with vitamin K shows a real result of normal lifestyle.

5. Connect = This experiment will prove whether vegetable which is full of vitamin K

will be effective to brain or not by using specific kind of vegetable which is broccoli.

Comparing to the previous experiment that gained result without having a specific kind

of food to prove.

Knowledge Gap:

Question not answered in experiments mentioned in the literature reviews

What kind of vegetable is both rich in vitamin K and omega-3 fatty acid?

Can vitamin K affect learning and memory?

Is broccoli a good brain food?

Our experiment will answer these questions by using broccoli as an independent variable

because it is rich in vitamin K. However, it also has some omega-3 fatty acids. It would be

tested by using hamsters as subjects and observe whether consumption of the brain food

broccoli affects hamsters learning ability or not.


Overview of Experiment Details:

Consuming good food will affect your brain in a good way due to the good bacteria in

the food, which will make up the intestinal microbiome. The intestinal microbiome protects

the lining of the intestines and provides a barrier against bad bacteria. This will help to improve

ability of nutrient absorption and activate neural pathways that travel directly between the gut

and the brain. In other word, it affects your mood and energy level.

List of Materials:

1. 2 male hamsters of the same species

2. Broccoli (preferably organic)

3. Baking soda (to clean the broccoli)

4. Hamster food

5. Cardboard

6. Timer

7. Glue (could be either liquid latex, elmers glue, or other non-toxic glue)

8. Cage

9. Bedding (such as wood sheddings)

10. Water bottle

11. Water

12. Ruler

13. Cutter

14. Pencil
Methodology:

Part I: Prepare the hamster

1. Prepare the cage by putting the bedding onto the ground of the cage, set up the food

bowl, water dispenser, and toys for the hamsters

2. Obtain the hamster. Let the hamster adapt to its new environment for 1-3 days

3. For the experimental hamster, slowly incorporate broccoli into its diet

a. As time passes, adjust the proportion of the diet by adding more broccoli and

lessen the normal food; add 1g of broccoli each 2 days until broccoli compose

about 50% of the experimental hamsters diet

15. Change the food and water of the hamster daily, so that the hamster would have a fresh

food every day

16. If the bedding is soiled, replace the bedding with the new one

17. Train the hamster so it would not be afraid or alarmed by the presence of human by

using the counterconditioning technique

a. Kneel down, let the hamster be at your eye level and put your hands into the

cage

b. At first, the hamster probably will show signs of fear due to the unfamiliar

presence of human

1. The sign could include being startled, showing incisors,

chattering of teeth, squeaking, biting, hiding and freezes in place

(playing dead)

c. Within 3 seconds of having presence of human around hamster, give the treat

to the hamster from your hand


d. Repeat step c in a random manner until the hamster shows no sign of fear when

human is present

Part 2: Prepare the maze

1. Design a classic maze based on information obtained from researches

2. Prepare cardboard and latex glue for building the maze

3. Let a big piece of cardboard be the floor of the maze. Cut rectangles of cardboard to be

the walls of the maze.

4. Build the maze following the design provided in Methodology: Maze Design and

Justification. Make the maze at least 16cm tall so that the hamster cannot jump out from

the maze.

5. Leave the completed maze for 2 days to dry. The smell of glue would be gone after it

is dried, and the hamster wont be distracted by the smell of glue.

Part 3: Experiment, data collection and data analysis

1. Place the food at the finish position

2. Let the hamsters explore the maze for the first time, timing the amount of time that the

hamster used to reach the finish position and count the amount of mistakes that the

hamster did. Record the information

3. Wait for about 30 minute before starting another trial in order to let the hamster rest

4. Repeat the process again. Record the information and record the amount of trials needed

for the hamster to run the maze without mistakes.

5. Use the information obtained to plot a number of mistakes versus time graph of each

of the hamsters. Compare the graph, time that each of the subjects used to run the maze

and the time that subjects used to learn how to run the maze.
Maze Design and Justification:

Why classic maze?

o Easy to build

o Can be used to measure learning of the rodent

o Complicate enough to figure out the results

Design

o Same ground levels not dangerous to ensure that hamster will not be

harmed

o 7.5 cm wide (pathways) not too tight or compress when hamsters run or turn

around itself inside the maze

o At least 16cm tall hamsters cannot jump out of the maze


o Have many branches show how do hamsters choose direction and determine

whether if the hamster could memorize the path or not ensure that differences

between two hamsters will be seen

o Have pathways that are blocked to see errors of hamsters

o There will be no anything else inside the maze except hamster and the food at

the finishing position avoid external stimuli that could not be controlled

because it might harm hamsters or cause the result being not accurate

o Start and finish lines are very far from each other to let hamster run around

the whole maze which will let us can observe their behavior easily

o Long enough for hamster to take time to complete the maze can record time

of each trials and see some differences between two hamsters

Materials

o Use cardboard for building the maze does not harm hamsters even they eat

it but also strong enough and not be broken when the hamster push

Methodology Literature Review:

Information about how to educate rodent based on experiment of other scientists

Introduction of experiment

o By Rachel Dehart, Amanda Gibson, Liz Miller

o Teaching rodent by using different stimuli

- What is the experiment about

o Purpose of the experiment = To solve how different stimuli affect learning ability

by teaching rodent using different stimuli

o Related to our experiment good stimuli will be used in our experiment in training

part need stimuli to make hamsters follow the right route


- Summary procedure of the experiment

o Type of rodents = Mice

o Materials used in their experiment food (crackers, cheese) and sound(bell)

o Method = Using different stimuli with different groups of rodents

o How the method solves their question = Collecting the data and observe their

behavior during running in the maze

- Result

o How the result relates or support our experiment what type of stimuli should be

used in the experiment (shape, sound, food,)

o How can we learn from this experiment = Learning from their good technique and

their errors

Timeline:

Table 1: Table showing the objectives, things to do, and the time frame

The table below shows the goal needed to be completed in order to finish the experiment

successfully and things to do in order to complete each of the objectives set with a time frame

that each of the objective are expected to be completed within.

Objective Things to do Time

frame

To collect footages Every members within the group help each other 29/3/17 -

for the to collect footages of the learning process 19/5/17

documentation

To do research Every members within the group make researches 29/3/17 -

and fill out their individual learning record form 19/5/17


To do worklog Every members within the group make work on 29/3/17 -

their own individual work log every working 19/5/17

period

Objective Things to do Time

frame

To finish the Every member help each other to think of the 21/4/17 -

outline of the topic needed to make researches about for the 30/4/17

proposal background information

Every member help each other to make researches

and make literature reviews in order to include it

into the proposal

Every member within the group list out the

materials needed to acquire in order to make the

experiment and think of the method of data

collection and experiment

Each of the members help each other to edit out

the possible mistakes and help each other to

improve the proposal

To finish the actual Finishing the proposal 1/5/17 -

proposal Check that the proposal includes an ethical 7/5/17

method of experiment and the wellbeing of the

rodents used in the experiment is taken in account


To make the maze To build the maze by using cardboard as the 9/5/17 -

material 15/5/17

Objective Things to do Time

frame

To prepare the Prepare the hamster that will be used in the 9/5/17 -

hamster experiment by gradually adding the broccoli into 15/5/17

the diet of the hamster (experimental group

subject) in order to prevent stress from the drastic

change of diet

To teach the To gradually train the hamster to run the maze 15/5/17 -

hamster to run the successfully 19/5/17

maze

To collect data To record the number of trials needed to run the 15/5/17 -

maze correctly 22/5/17

To record the amount of time needed to run the

maze correctly

To record the amount of time that the hamster run

to the wrong direction

To collect footages To film the process of the experiment in order to 15/5/17 -

of the experiment include into the final product 22/5/17


Objective Things to do Time

frame

To finish the final To put the footages together as seamlessly as 22/5/17 -

product (video) possible 26/5/17

To put some captions or description in order to

make the footages shown in the video clear and

easy to be understood

To make individual Each of the members within the group reflect on 22/5/17 -

reflection the learning process used in order to finish the 26/5/17

self-directed project.

Ethical/ Humane Considerations:

To ensure that no animals were harmed in this experiment

No drastic change of diet

o Drastic change would contribute to stress of the rodent

Minimize possible factors that would contribute to stress in hamsters

o Design the the maze so that the path within the maze would not be too narrow

so that the rodent would not feel uncomfortable or stressed

Use toxic-free materials

o Cardboards are safe for hamsters


o Non-toxic Elmers glue or liquid latex is safe for hamster in case that they

accidentally consume it

o No dyed parts and the glued parts are hidden

No inedible food was incorporated into the experiment

o Broccoli is edible for hamster

o To minimize pesticide use organic veggies & use baking soda to remove

pesticides

Maze

o The width maze should not be too tight for hamsters

o No slope / incline plane which would cause an accident to hamsters

Surroundings

o Silent environments

o No disturbance that would distresses hamsters

Literature review to justify the maze design:

According to the literature review I & II that the food beverages affect to

human learning and memorizing system which has been approved that are similar to the

hamsters, due to, the Research of Learning During Stressful Times by Tracey J. Shors from

the department of Psychology, Center for Collaborative Neuroscience, Rutgers University,

New Jersey 08854 ,USA. As researched, hamsters are considered as one of the most famous

species of rodents that were used to experiments for medication usage. Therefore, in this

experiment, it was decided that the anxiety in hamsters would be controlled , since it might

provide an invalid result from our experiment, and also as an ethical / human consideration,

anxiety would be controlled in order to prevent mental harm to the subject.


References from the Learning During Stressful Times research that provides an

information about Stress and Operant Conditioning (also known as the Learning Helplessness

states) that when the creatures are exposed with inescapable stress, it would express in

various ways; such as sleep and eating disorders, stomach ulcers, and the reduction of

immune system. [The phenomenon was promoted as creatures modeled for the depression in

humans, Seligman 1975]. Therefore, this research provides a strong answer about anxiety that

whether in rodents or human, it would result a negative impact. According to the information

above, the researchers have been contributing an experiment that focuses on both stressed

and unstressed rats and their learning. In the experiment, they used the 8-arms radial maze to

test the hungriness of the 2 different groups. The result shows that in the first day, the

stressed group did better than the unstressed group, but in the next day the unstressed group

revealed a better learning ability than the stressed group. In the other word, learning while

stressed is not efficient, therefore, controlling the surroundings and environments and

designing the maze are being concerned as much as possible in our experiment. As the

researcher chose the 8-arms radial maze that was recorded as the best maze design for testing

the rats hungriness abilities, which is similar as the classical maze design that our group have

chosen.

In order to critique, among the learning during stressful times research, there

are some developments that could be based from the research information and implied to our

experiment; there are some factors that cause an error in their experiment, whether the

quantities of trials that used to test was insufficient to give out reliable result and the

conditions of the rats, for instance, the days that the rats stay at those place because their

emotional statement plays a crucial role of how they going to stimuli back to the stimulators ,

etc.
Data Collections and Limitations:

Create a number of mistakes done versus time graph

o The learning curve could be obtained from the graph

o When the number of mistakes is low or is zero and after several trials more is

consistent the rodent had learned how to run the maze

Figure 1: Sample of the #of mistakes versus time graph that would be used in order to obtain

the learning curve of each of the subjects and analyze the information obtained from the

experiment

Record the time used to train each of the subject

Record the time used to run the maze and find the best time (least time used) and worst

time (most time used)

Compare the time recorded from both subjects


Table 2: Sample of the data table that would be used in order to collect and compare the

information that would be obtained from the experiment.

Hamster that does not eat broccoli Hamster that eats broccoli

Attempt Time Time Amount of Time Time Amount of How


used to for mistakes used to for mistakes much
complete training done before complete training done before broccoli
the maze (round) completing the maze (round) completing it eats
(sec) the maze (sec) the maze before
(times) (times) running
the maze
(grams)

Trial 1

Trial 2

Trial3

Trial 4

Limitations

Since two different hamsters were used, there is a factor that couldnt be controlled

o The factor includes difference in morphology (shorter/longer limbs, better

sense of smelling, etc.), difference in initial/innate learning ability

o To solve this problem, the same rodent should be used in further studies

Since the diet of the hamster couldnt be changed into 100% broccoli due to the

possible lack of nutrients the hamster could get, the result might vary

o The result could be from other factors, not broccoli alone


o Might not result in a clear difference of information/time in both rodents

o To solve this problem, the diet of the rodent that would be the experimental

group might be planned by using other methods that would ensure sufficient

nutrient, yet undisturbing to the experimental result

Since this experiment was planned as a short term experiment, the long term result

cannot be tested

o If the brain food doesnt have a short term result, or the time is not sufficient

enough for the brain food (broccoli) to effect anything, it might result in no

difference of information from both subjects

o To solve this problem, the same experiment could be carried out in a long-

term period

Conclusion:

In order to justify the effects of broccoli, which is one of the brain foods, on learning,

experiment by using hamsters as subjects would be conducted. By using the control hamster,

which is the hamster that would have a normal diet, and the experimental hamster, which is the

hamster that would have broccoli incorporated into its diet, the difference in ability to learn

would be determined by using the amount of mistakes done, time used to learn how to run the

maze, and time used to complete the maze. The maze that would be used in the experiment

would be the same, classical modeled maze made with cardboard and other hamster-safe

materials in order to ensure the safety of the subjects and to ensure that there would be no

difference in the maze that would alter the result of the experiment.
References

Chiba, A. (2015, April 02). Why we use rodents to research the brain. Retrieved April 29, 2017,

from http://www.sandiegouniontribune.com/news/science/sdut-brain-logic-behind-using-

rodents-2015apr02-htmlstory.html

Dehart, R., Gibson, A., & Miller, L. (2014, May 7). Amazing Mice. Retrieved April 23, 2017, from

http://jrscience.wcp.muohio.edu/nsfall01/FinalArticles/AmazingMice.html

Federation of American Societies for Experimental Biology. (2017, April 25). Vitamin K keeps

brains sharp. Retrieved April 28, 2017, from http://www.newhope.com/nutrition/vitamin-k-

keeps-brains-sharp

H. (n.d.). Stress in a hamster. Retrieved March 30, 2017, from

http://www.hammysworld.com/index.php?p=stress

MD, E. S. (2015, November 17). Nutritional psychiatry: Your brain on food. Retrieved April 28,

2017, from http://www.health.harvard.edu/blog/nutritional-psychiatry-your-brain-on-food-

201511168626

Wolpert, S. (2008, July 09). Scientists learn how what you eat affects your brain - and those of your

kids. Retrieved April 28, 2017, from http://newsroom.ucla.edu/releases/scientists-learn-how-

food-affects-52668

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