BIOLOGY LAB REPORT

TITLE : HABITUATION

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Abstract

In this experiment, habituation in snail is being investigated. Each student is needed to find one snail

and 10 stimulations are being given to each snail. The time taken for the snail to retreat and re-emerge

is recorded and the graph is plotted. From this experiment, I found out that generally the time taken for

the snail is decreasing with increasing no. of stimulation. After a few stimuli had been given, the snail

became habituated. Besides, I also found that different snails (from other student) need different no. of

stimulation to become habituated.

Introduction

1. Snail(1)

Snails can be found in gardens, in ponds and even in the sea. They belong to a group of animals with a

soft body called molluscs which are related to oysters, clams, and other shellfish. Characteristically

they have soft, unsegmented bodies. Normally, their soft bodies are protected by a hard shell. The

scientific name for the garden snail is Helix aspersa. The body of the snail is long, moist and slimy. It

has a shell to protect its soft body. When the snail is disturbed, it simply withdraws or pulls itself back

into its shell. The snail also retreats into its shell and seals the entrance in dry weather to protect its

body from drying up. A snail is most active at night and on cloudy days. It does not like the sunshine

very much. During very cold weather or winter, it hibernates in the ground.

Garden snails are best for this activity. If possible, snails should be collected locally, during warm, wet

conditions. Students can find for snails early in the morning or evening hours, and check on leafy

plants and under overturned pots.

Snails can be housed in any closed container that has air holes. For bedding, moist paper towels work

well. The activities in this lesson will not harm snails. Snails eat various leafy foods; young green

leaves works well if the snails will be kept up to several weeks. Snails go dormant if they have not

been fed for several days, so if it will be a few weeks before they are studied, food should be withheld

and their enclosure kept drier; they can survive for several months in a dormant state. The snails can

be awakened usually in minutes by being rinsed and fed.

Figure 1: Snail and its life cycle(2)

2. Learning

Learning ability is one of many factors which will determine the survival of an animal. Learning is the

modification of behavior resulting from specific experiences. There are two main types of behavior:

innate and learned. In this biology practical, one form of learned behavior which is habituation is

explained with the help of garden snails.

i. Habituation(3)(4)

Habituation is learning process exhibited by several different species. Although it’s quite simple, it’s

been suggested that habituation may be the basis of all other forms of learning. The mechanisms of

habituation are usually closely regarded as akin to the mechanisms of learning.

An example of habituation is wearing a watch, if you were to put your watch on your other arm it

would feel strange and out of place, as it would if you took it off the arm it’s on now. This is because

your body has got used to it being on one arm and not the other. If you were to leave the watch on the

other arm for a few days then you would forget about it.

This process is known as Habituation and can be summed up as when stimuli are neither harmful nor

helpful to any creature it will eventually be ignored.

A quick way of testing this is to gently touch a snail’s tentacle, when touched it will withdraw sharply.

If it is touched repeatedly eventually the snail will see that this is doing it no harm and not bother

withdrawing its tentacle as much. Habituation is a phenomenon that is widespread across the animal

kingdom; it helps them survive by balancing responses to either harmful stimuli or stimuli with

uncertain significance. An effect of habituation is usually that a response that a habituated response

will soon reappear if stimuli are withheld for a long time, the guppy will start retreating again if no

shadows are shown for about a day.

However, habituation to the shadow the next day will occur quicker because the animal has previously

been habituated. This process is known as dishabitation. No animal can afford to ignore potentially

dangerous stimuli; the process of habituation is a compromise between saving energy and the animal

saving itself. In relation, the snail will not be afraid after a while of no harmful stimuli but when the

stimulus is first presented the snail should cower in an attempt to preserve its own life.

Humphrey (1933) studied the process of habituation, and he was one of the first. He placed a number

of snails on a glass plate; he would then give the plate a sharp jerk that would cause the snails to react

and to withdraw reflexively into their shells. After each jerk, the number of snails that had withdrawn

was counted, it was found that after several trials the number showing withdrawal reflexes decreased

and less and less would withdraw into their shells.

Since Humphrey’s study a variety of techniques have been used to study habituation. They also found

repeated stimulation of the aplysia’s siphon causes a reduction in the magnitude of gill withdrawal .

Figure 2: Habituation in Aplysia (5)

One of the famous experiments investigating habituation is the experiment involving the giant sea slug

Aplysia. The giant sea slug called Aplysia californica is often chosen for the studying of memory. Its

brain has about 20000 neurons, some of which are large enough to be visible to the naked

eye. Aplysia can learn and most importantly it is found that the mechanisms and principles involved in

its formation of short- and long-term memories are conserved throughout the animal kingdom,

including in humans. Aplysia exhibits a behavior of protective reflex in which the sea slug withdraws

its gill into the safety of the mantel cavity in response to a mild touch stimulus to another part of the

body called the siphon. If the stimulus is repeated a number of times, the gill withdrawal reflex

becomes weaker until finally the animal ignores the touch stimulus. The waning of sensitivity to

repeated stimulation is known as habituation and is a very simple form of learning found in all

animals, including humans.

At the macromolecule level, it is known that the neurotransmitter involved in the processes is the

serotonin. A puff of serotonin alone can substitute for the siphon shock. It is shown further that the

serotonin triggers the release of the second chemical messenger called cyclic-AMP. It activates an

important type of enzyme called a kinase, which modifies the properties of particular target proteins

by adding a phosphate molecule to them; the term for this is protein phosphorylation. The target for

this modification in the sensory neuron is a potassium channel protein which is important in the

downward phase of the action potential. The net result of phosphorylation is a prolongation of the

action potential in the sensory neuron and so more neurotransmitter is released by the sensory neuron.

Thus the sensory neuron's synapse with the gill motor neuron is strengthened.

In short-term memory, special enzymes quickly remove phosphates from the proteins and return them

to their original state, restoring the synaptic strength to its lower pre-sensitized level. However,

following repeated serotonin delivery, the level of cAMP-activated kinase is much higher and this

allows the crucial step in the formation of long-term memory to occur. This crucial step is the

transport from the synapse to the cell body of kinase molecules that have been activated by c-AMP.

Once in the cell body the activated kinases enter the nucleus and start to regulate the expression of

particular genes. In Aplysia, proteins that result from this process of gene activation are transported

back to the synapse where they are used to maintain the strength of synapses already affected by local

effects of c-AMP and to grow new synaptic connections. So in Aplysia the conversion of a short-term

into a long-term memory involves the reinforcement of the short-term changes in synaptic strength and

the growth of new synapses, both of which require the synthesis of new proteins.

Objective

To investigate whether snails will become habituated due to the repeated stimulation.

Problem Statement

Can snail become habituated to a repeated stimulus?

Hypothesis

When the snail is being given stimulation (touch) repeatedly, it will eventually become habituated.

Thus, the snail would not retreat when another touch is given. As number of stimulations between the

eyes stalks of the snails increases, the time for the eye stalk withdrawal will decreases , thus showing

negative correlation.

Null hypothesis

There is no significant correlation between the number of pokes on the snails and the time for the

eyestalk to be withdrawn.

Experiment design

Independent measures will be used for this experiment, as they are separate, different snails being

used.

Apparatus

Stopwatch

Materials

Garden snails, dampened cotton wool bud, clean and firm surface for the snails and water

Variables :

Types of Variables Ways to control the variables

Manipulated Variable:

Number of pokes on snail’s eye stalk (no.)

Damp cotton bud used to poke eye stalk of snail

ranges from one to ten pokes.

Responding Variables:

Time for eyestalk withdrawal (s)

Using stopwatch, time taken for the eyestalk to re-

r-emerge calculated

Control Variables:

Same size and species snails

Same volume and force of stimulation

Assuming same age between samples. Different

species or age snails react differently.

Each snail’s trauma control to be minimal,

avoiding ethical problems.

Procedure :

1. A garden snail is being captured and placed on a clean, firm surface. The snail is being left for

a few minutes so that it has gotten used to the new surrounding and fully emerged from its

shell.

2. A cotton wool bud is being dampened with water.

3. The snail is firmly touched between the eye stalks by using the dampened cotton wool bud and

the stopwatch is immediately started.

4. The time taken for the snail to retreat and fully emerged is recorded.

5. Procedure in step 3 and step 4 are repeated for a total of 10 touches with the timing for the

snails to re-emerge each time.

6. The data is tabulated in a suitable table and other snail’s data were tabulated as well.

7. A graph of time against no. of stimulation is then plotted.

Risk Assessment

Safety precaution

In order to avoid any accident or injury during the experiment in laboratory, the precautionary steps

should be taken and applied. Wearing lab coat and a pair of suitable shoes are compulsory when

conducting an experiment in the lab at all times to protect the skin and clothing from spillage of any

chemical substance. For the snail, make sure to be careful, not to make it stressful that it already has

due to different surrounding. The surrounding needs to be constantly damp, mimicking the snail’s real

habitat.

Ethical issue

There are no ethical problems when dealing with invertebrates. The snails should suffer no

psychological harm and will be released straight back to the wild afterwards.

In this case the knowledge gained justifies the procedure, as the procedure should cause no harm

whatsoever. The experiment will not harm or stress the snails, they aren’t social animals so they

won’t mind being caged for a short period as long as the experiment takes and common garden

snails are not an endangered species, they are actually an incredibly common species. Snails are a

great model for this experiment because they can be easily handled and manipulated. Snails are

advantageous when it comes to experiments because they are easy to keep and maintain, also

abundant in surrounding, easy to study plus they are inexpensive. But, humans can’t be compared to

snail due to their difference in the genetic make-up. In this experiment, the snails are not tested until

they died. Only 10 stimulations are being given and after that, they are released back to the

environment. In addition, the use of cotton wool buds instead of sharp object can reduce the risk of

injury to the snails. Although the snails are simple organisms that may not suffer in the same way as

higher animal, they still deserve respect. These snails are released back to their original habitat and

reduce the disturbance on the food chain.

Bias

The only bias possible in an experiment testing reflexes is experimenter bias, there could be some

ambiguity as to whether or not a snail did actually reflexively withdraw or not.

Reliability

This experiment is relatively simple, and as a result there is no reason why it shouldn’t work. The

results should show correlation with what’s expected to happen and it will be repeated a few times to

make sure that it works properly.

Validity

This experiment has been done before by Humphrey (1933) so the procedure will just be following

his that was already proved valid, yet it was quite an old experiment so it can’t be trusted

completely. Also, the presence of a human at the time of the experiment may affect the snails and

may make them react more. However, eventually the experiment should still have the same effect.

Results

No. of

stimulation

Time taken / s

Snail 1 Snail 2 Snail 3

1 22.2 27.9 13.4

2 21.8 17.5 7.3

3 20.3 16.0 5.4

4 19.0 12.4 5.2

5 14.1 10.9 4.6

6 12.8 7.7 3.6

7 8.9 5.2 2.3

8 4.2 0.0 0.0

9 0.0 0.0 0.0

10 0.0 0.0 0.0

Table 1: The time taken against no. of stimulation for the three snails is plotted.

Graph 1: Number of pokes against time taken for eye stalk to re-emerge

0

5

10

15

20

25

30

1 2 3 4 5 6 7 8 9 10

Tim

e ta

ken

fo

r th

e ey

e st

alk

to

re-

emer

ge/

s

Number of pokes

Snail 1

Snail 2

Snail 3

Spearman’s rank, rs correlation coefficient.

Null hypothesis, H0: No significant correlation between the number of stimulations and the time for

eye stalk withdrawal.

Alternative hypothesis, H1: Negative correlation between the number of stimulations and the time for

eye stalk withdrawal.

Level of significance for a one-tailed test, α = 0.05

Reject H0 if test statistics ≤ -0.564

Number of

pokes

Rank

stimulation

Mean time /

seconds

Rank time Difference/D D2

1 1 21.17 10 -9 81

2 2 15.53 9 -7 49

3 3 13.90 8 -5 25

4 4 12.20 7 -3 9

5 5 9.87 6 -1 1

6 6 8.03 5 1 1

7 7 5.47 4 3 9

8 8 1.40 3 5 25

9 9 0.00 2 7 49

10 10 0.00 1 9 81

Table 2 :Spearman’s rank, rs correlation coefficient data.

Spearman’s rank, rs correlation coefficient,

Where

= rank stimulation – rank time

= 10

Based on table of critical value and by using significance value of 5% = 0.05 the critical value was

found out to be -0.5636.

Reject H0 as there is sufficient evidence to say that there is negative correlation between the variables.

Therefore, snails can become habituated to a touch stimulus

0 -0.5636

Rejection

region

DISCUSSION

Data Analysis

From the graph, we can see that all three snails showed the same pattern for the habituation

experiment. All three snails showed negative correlation between the time taken and no. of stimulus

which means as the no. of stimulus increases, the time taken for eye stalk withdrawal decreases.

Therefore, in general, we can conclude that the time taken for the snails to retreat and re-emerge

decreases as the no. of stimulus increases, and at certain point became fully habituated.

Comparisons can also be made between all three snails although the trends are almost the same

generally. This is due to different individuals of snail used and each snail is only tested once. Firstly,

we can see that snail 2 took the longest time to retreat and re-emerge after the 1st stimulation is given

with 27.9s, followed by snail 1 and snail 3. Other than that, it is clear that snail 2 and snail 3 had

become habituated after the 8th

stimulation, while snail 1 needs extra stimulation before being fully

habituated (at 9th

stimulation). Therefore, we can also conclude that different snails need different

amount of stimulation before being habituated.

Evaluation

Habituation is a decrease in response to a stimulus after repeated stimulus. Nervous system is that the

calcium channel in the presynaptic membrane becomes less responsive with repeated stimulations.

With fewer calcium channels open, less calcium ions cross into the presynaptic knob. As a result,

fewer vesicles move towards presynaptic membrane, fuse, and discharge their neurotransmitter. Less

neurotransmitter available to bind to the post-synaptic membrane so the post-synaptic excitatory

potential is not high enough (under threshold level) to trigger an action potential. . Thus, the action

potential generated is not high enough which cause no response.

According to the graph drawn, there is negative correlation between the number of stimulus

and the time taken for the eyestalk to fully emerge. This happen due to habituation has takes place sine

same stimulus is given and identified as not important or not danger. Thus, the snails doesn’t respond

after several seconds.

Limitations

There are several limitations that have been identified throughout this experiment.

Not able to find the average time taken for each snail. This is because, once the snail

become habituated due to the given stimulus. It would not react to the nest stimulus. A

long period of time needed for the snail to restore normal activity and this will lead to

time wastage if the experiment is going to be carried out again. Without extra sets of

reading, the average value cannot be calculated thus affecting the validity of the data

Difference in snail’s original habitat and experiment site (the lightning and the

environment are too artificial during experiment and captive period). The snail may be

unable to adapt to the surrounding, feeling stressful (anthropomorphism), thus affecting

the time taken for the eye stalk to re-emerge. The experiment should be done outdoors in

the field or the habitat of the snails instead of laboratory or class to give more pleasant

and comfortable feeling to the snails toward its environment.

The frequency of stimulus is not consistent. Time for the eye stalk to re-emerge is taken

and recorded by each and every student individually, thus time taken to record the data

and carry out the next stimulus is different per stimulus. Or if touched too frequently, the

tentacle may remain withdrawn throughout the experiment.

Snails can “remember” their immediate previous experiences, if a snail is handled

roughly it may react to touch by completely withdrawing, or if the snail is just waking

up from being dormant it may not respond as quickly. Snails also have individual

differences that may contribute to how they react to this experiment. Thus, each and

every snail response is differently.

Sources of errors

Several sources of error in this experiment were identified and steps were taken to minimize

these errors to make the result more accurate.

Difference in snail’s original habitat and experiment site is too pronounced due to

different light intensity and humidity. This is reduce by making the experiment site

damp to make the snail comfortable.

The strength of stimulus may differ according to the student’s nature. Students that are

scared of snail may give small and light stimulus and the others vice versa. Thus, the

stimulus given is very much controlled by the individual.

Conclusion

From this experiment it is true that the time taken for the snails to retreat and re-emerge decreases with

increasing no. of stimulation. Over a no. of stimulation, the snails had become habituated. Other than

that, we can also conclude that different snails need different no. of stimulus before being habituated.

Thus, the hypothesis is accepted.

References

1. www.animalbehavioronline.com/habituation.html. Accessed on 16th January 2013.

2. http://www.snail-world.com/Snail-Anatomy.html. Accessed on 16th January 2013.

3. C J Clegg, 2008. Edexcel Biology for AS, 209 p . London : Hodder Education.

4. Habituation in snail, Matthew Ferguson

5. http://visual.merriam-webster.com/animal-kingdom/mollusks/snail/Aplysia. Accessed on 16th

January 2013