REFLEX ACTION IN THE FROG

Experiment No. 2

Group 3: Avian

Ibita, Arlex A.
Morante, Althea Louise Rica E.
Lopez, Bryan I.
Natino, Reysabel S.
Lagumen, Romae M.
Gacho, Lester S.
Lamadrid, Rhea B.

BS Biology 4A

Submitted to:

PHOEBIE GAEL ASIS

Instructor, Animal Physiology

November 6, 2023
 ABSTRACT
In this experiment involving frogs, a comparison was conducted between normal frogs,
spinal frogs, and double-pithed frogs to investigate their reflex actions and response times.
Normal frogs exhibit various essential reflexes, while the spinal frog which the brain has
disrupted still exhibits reflexes that considerably occur due to the automatic response of the
spinal reflexes. On the other hand, the double-pithed frog completely abolished their capacity
to react to stimuli. This experiment underscored the central nervous system's vital role in
coordinating frog reflexes and responses, illustrating that pithing disrupts neural pathways
responsible for reflex actions, resulting in a lack of response. In addition, it was also revealed
how disruptions affect to inhibit reflexes, as well as its actions and time; and how stimulation
may implicate or trigger a reaction and its reflex phase. In summary, varying levels of neural
intervention, like spinal pithing, have a profound impact on frog reflexes, highlighting the
central nervous system's significance in their control and response modulation.

INTRODUCTION and adaptation to their environment.

Specifically, the indication of these reflexes is
The functioning of nerves relies on the
due to their physiology. The frogs possess a
combined action of sensory and motor fibers.
highly developed nervous system that allows
However, it is not always necessary for a
them to respond quickly to external stimuli.
sensory impulse to travel to the brain to
This includes reacting to potential dangers,
trigger a motor response. In some cases, a
such as predators, and also ensuring efficient
sensory neuron can directly connect to a
movement and coordination in their daily
motor neuron in the spinal cord, forming a
activities.
reflex arc that operates automatically. An
example of this is the kneejerk response, This experiment aims to observe
where tapping the tendon below the kneecap various reflex actions in frogs, including the
causes the leg to jerk involuntarily. The tap withdrawal reflex, corneal reflex, and righting
triggers an impulse that travels to the spinal reflex, among others. Additionally, the
cord and back to the leg muscle, bypassing the experiment seeks to investigate the effects of
brain. This type of response is known as an single-pithing and double-pithing on these
involuntary reflex action. reflexes. It also intends to examine key
characteristics of reflexes, such as reflex time,
Typically, a reflex arc includes one or
synaptic delay, synaptic fatigue, and the
more connector neurons that modulate the
inhibition and coordination of reflexes.
response, allowing different levels of reaction
based on the strength, weakness, or duration Objectives
of the stimulus. Reflex arcs frequently
In this experiment, the objectives are:
connect with other arcs through nerve fibers
in the spinal cord. These connections enable 1. Observe reflex actions in the frog such
the brain to recognize a sensory stimulus, as withdrawal reflex, corneal reflex,
observe the reflex response, and suppress that and righting reflex;
response if necessary. 2. Study the effect of single-pithing or
decerebration and double-pithing on
Meanwhile, in the case of frogs, these
these reflexes; and
amphibians are known to display various
3. Learn some of the principal
reflex actions that are crucial for their survival
characteristics of reflexes such as

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 reflex time, synaptic delay, synaptic reflex." A small piece of filter paper
fatigue, and inhibition and (alternative: a piece of paper towel)
coordination of reflexes. moistened with 15% acetic acid was
placed on the frog's body near its rear
MATERIALS AND METHODS
end, and the researchers noted the
A. Reflexes in Normal, Spinal, and movement direction of the frog's leg in an
Double-Pithed Frogs attempt to remove the paper.

The tests were conducted on B. Reflex Action and Reflex Time

normal, spinal, and double-pithed frogs.
A1% stock solution of HCl was
The initial test was observing the resting
used to prepare different concentrations
postures of the frogs, noting the positions
(0.5%, 0.3%, 0.1%, and 0.05%) using tap
of their head, eyes, forelegs, and hind
water as a diluent. The experiment
legs. It was also observed the closing and
involved a spinal frog, with its longest toe
opening of the nostrils and the raising and
dipped into the lowest concentration of
lowering of the frog's mouth door.
HCl for 1:30 minutes, recording the
The reflex experiments were reflex time. Afterward, the foot was
followed. The first test focused on the immersed in a sodium bicarbonate
frogs' equilibrium maintenance. The solution and rinsed with tap water. This
frogs were placed on a board that gently process was repeated, and the average
revolved in a horizontal plane and also in reflex time was calculated. The
a position where their heads were experiment was replicated with different
elevated. Afterward, swimming acid concentrations, creating a graph
movements and climbing responses were showing reflex time (seconds) plotted
also recorded. The second test examined against acid concentration.
evidence of sight. The researchers
C. Inhibition of Reflexes
elicited the blink/protective eye reflex by
bringing a pen closer to the frog's eye The passage describes a series of
until it blinked. The third test assessed experiments involving frogs. Initially, a
muscular tension and reflexes. One frog is placed on a desk and its reflex time
member of the group restrained the frog is observed. The experiment is repeated
while another member straightened one after the frog's legs have been securely
of its hind legs. The frog was then bound with strings. The inhibition is
prodded to jump, and tension and noted and explained. The cerebral
jumping behavior were recorded. The hemispheres of the frog's brain are then
pain response of the frogs was tested by removed, and the reflex time experiment
pinching their toes and noting their is repeated using a dilute 0.3%
flexion-withdrawal reflex. One foot was hydrochloric acid solution. The average
immersed in cold water and gradually reflex time is computed. The experiment
heated, while the other foot was dipped in continues by exposing the optic lobes to
80°C water. The researchers also sodium chloride crystals and noting the
observed the quieting response and clasp reflex time. The optic lobe is eventually
reflex of male frogs by wetting their removed, and the reflex time is
fingers and gently stroking the frog's determined again after a set time.
belly. The final test was conducted to
observe the "wipe reflex" or "scratch

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D. Synaptic Fatigue Table 1. The reflex reaction of normal, spinal, and
double-pithed frogs. The experiment indicates a
The brain of a frog was pithed. positive reaction towards normal frogs, with least
Both sciatic nerves and the muscles were to no reactions from both spinal and double-pithed
frogs.
exposed. With repeated shocks, the left Double-
sciatic nerve was stimulated causing Reflex/ Normal Spinal
Pithed
Reaction Frog Frog
muscles of both legs to contract. This Frog
stimulation was continued until the right I. Resting Posture
a. Head Upright Tilted Parallel
muscle ceased to contract. The to the
stimulation was then stopped for about 2 board
minutes, and then, the stimulation was b. Eyes Blinking Closed Closed
repeated to make sure of the result. c. Forelegs Bent Slightly Flat on
bent the
RESULTS AND DISCUSSIONS board
d. Hind Legs Bent Bent Flat
A. Reflexes in Normal, Spinal, and but flat
on the
Double-Pithed Frogs surface
II. Respiration
a. Movement
of the + + -
nostrils
b. Oscillation
+ + -
of the throat
III. Test for Maintenance of Equilibrium
a. Frog is
+ - -
turned over
b. Frog is
turned
around in a + - -
horizontal
plane
c. Frog is tilted + - -
IV. Swimming Movement
a. Swimming
on the tank + - -
of water
b. Climbing on
the floating + - -
board
c. Righting
+ - -
Reflex
V. Protective Eye Reflex
a. Blinking of
+ - -
the eye
VI. Muscular Tension
a. Limb
extension + + +
reflex
b. Limb
flexion + - -
reflex
c. Jumping
Figure 1. Frogs in their resting position. Initially, the + - -
behavior
frogs were prepared for the part A experiment. (A) VII. Pain Response
Normal Frog. (B) Spinal Frog: single pithing. (C)
Double-pithed frog.

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 a. Withdrawal particularly in the presence of probable harm.
reflex from + - - The nociceptive flexion reflex, withdrawal
pinching
response or reflex, plays a crucial role in
b. Withdrawal
reflex from + - - safeguarding the body against harmful
hot water stimuli. Additionally, it is an automatic
c. Withdrawal reaction of the spinal cord. (Fischer &Truog,
reflex from + - - 2015; Derderian, Shumway, & Tadi, 2023).
cold water Blinking has been the obvious reaction of the
d. Wipe reflex + - -
normal frog. When irritants or foreign objects
In the first experiment, the frogs have make contact with the eye, they first come
positioned in a resting posture results showed into contact with the cornea. This contact may
that: the positioning of the forelegs and hind trigger two reflexes: the blink reflex (also
legs of those pithed were greatly affected, known as the corneal reflex) and tear
displaying the paralleling of the legs instead production (Peterson & Hamel, 2023).
of an angular positioning; the upright position Maintenance of equilibrium or normal posture
of the head was also inflicted. Respiration was was also a reflex that occurs when a vertebrate
evident in normal frogs, while the slow is triggered. It is a natural and instinctive
movement of the nostrils to the spinal frog. behavior observed in many animal species.
Contrastingly, no respiratory indication was This reflex is specifically triggered when an
seen in the double-pithed frog (Table 1). animal finds itself upside down or placed on
its back, prompting it to swiftly maneuver its
In terms of maintenance of body and regain an upright position. (Berman
equilibrium, swimming movement, protective et al., 2023). Wiping movement of the hind
eye reflex, muscular tension, and pain legs was also observed when stimulated with
response only the normal frog had a positive chemicals. The frog's hind limb produces a
reaction, while those that were pithed had wiping reflex when the distal skin of its
least to no reaction at all (Table 1). forelimb is electrically or chemically
stimulated (Fukson, Berkinblit, & Feldman,
Reflex is an automatic response, most 1980).
especially, it is seen among vertebrate
animals. Reflex is described as a natural and Meanwhile, the absence of reflexes
rapid movement that occurs involuntarily in among pithed frogs was evident. As the
response to a stimulus. It is an automatic rupture to the central sensing sites was
reaction that does not require conscious recognized, immediate disassociation of
thought and is facilitated through a reflex arc. actions was observed, particularly in pain
These reflex arcs effectively act on an impulse response. The withdrawal reflexes were
before it even reaches the brain (Physiopedia, absent since it primarily involved the
2019). The normal frog displays the positive peripheral and central nervous systems, as
reflex, which indicates no obscure to the well as the epidermis and musculoskeletal
sensing pathways from sensing external system. The peripheral nervous system
signals, delivery of electric impulses, includes sensory neurons that respond to
processing of the received impulse, and harmful stimuli and the central nervous
eventually reacting based on the stimulus system aids in communication between
through muscular or behavioral reactions. sensory neurons through the spinal cord,
allowing the withdrawal reflex to occur
In addition, the normal frog in the (Derderian, Shumway, & Tadi, 2023).
experiment exhibited notable reflexes such as
the withdrawal, corneal, righting, and wiping However, there were articles implying
reflexes that were important to the that single pithing may only destroy
understanding of basic reflex action. The consciousness, and reflexes may persist.
withdrawal reflex specifies retreating action, Unlike voluntary activities, reflexes occur

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without conscious thought. Therefore, The results showed the speed of reflex
reflexes can persist even when higher-level time of the pithed frog to different
brain functions are absent (Purves et al., concentrations of Sodium Chloride. In 0.05%
2023). For instance, in the spinal frog, which concentration of Sodium Chloride, the frog
spinal reflex is notable. The mentioned reflex reacted on the second trial after 87 seconds
does not require any input from the central (Table 2). In 0.1% concentration, it reacted on
nervous system. One example is the
the third trial after 80 seconds (Table 2). At
withdrawal reflex, where stimulation of the
0.3% concentration, the frog reacted on the
skin between the claws causes the limb to
flex. However, sensory information can also third trial after 87 seconds (Table 2). Lastly,
be to the brain, leading to additional behaviors in 0.5 concentration, it reacted on the first trial
(Nordquist, Meijer, van der Staay, Arndt, after 52 seconds and on the second trial after
2017). This withdrawal reaction in the spinal 78 seconds with an average of 65 seconds
frog was seen in the latter spinal frog (Table 2). Note that no response was observed
experiment; although, in the current section, on the first trial of all the concentrations
no visible reactions were recorded. except the 0.5%, second trial on 0.1 and 0.3%,
B. Reflex Action and Reflex Time and third trial on 0.05% and 0.5% of the
sodium chloride concentration.
Table 2. Pithed frog’s average reflex time to
hydrochloric acid (HCl) in different concentrations. The findings depict the relationship
Frog Reaction between the reflex time of a singly pithed
Concentration Within 90 Seconds specimen and the concentration of the
of Sodium (Seconds) Average
Chloride (%) Trial Trial Trial solution it's exposed to. Reflex actions are
1 2 3 immediate, simple responses by the body to
0.05 - 87 - 87 external stimuli. Typically, these impulses
0.1 - - 80 80 travel through a reflex arc, where a sensory
0.3 - - 87 87
receptor receives the stimulus and conveys
the information to a sensory neuron, which
0.5 52 78 - 65
then carries the impulse to the central nervous
system (CNS). Within
the CNS, an
Average reflex time vs stimulus strength
interneuron processes
100
and relays the signal to
90
80
a motor neuron, which
70 in turn transmits it to an
Time (seconds)

60 effector, resulting in a
50 suitable response to the
40 stimulus (Betts et al.,
30 2016).
20
10 As the
0 concentration of the
0.05 0.1 0.3 0.5
irritant compound
% HCl
increases, the reflex
Reaction Linear (Reaction) time shortens, as
depicted in Figure X.
Figure 2. Graph of average reflex time vs stimulus strength. A direct This is because higher
relationship between the reflex time and the strength of the stimulus has been concentrations
showed.

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generate stronger impulses, prompting the dilutions of HCl resulted in an average of 2
body to respond swiftly to the stimulus seconds per reflex in 0.1%, oppositely to
(Matthew and Rushworth, 1956). 0.5% which has a 1-second average reflex of
2 seconds per reaction (Table 3). On the other
C. Inhibition of Reflexes
hand, the exposure of the exposed optic lobe
in NaCl resulted in a much lower reflex, or at
least none after 5 minutes of observation.
The righting reflex is an innate
behavior observed in many animals, including
frogs, that helps them regain an upright
posture when placed in an inverted or
abnormal position (Libretexts, 2021). It is
caused by the force that draws a reflex
inhibition which makes the legs relax or
suppress the other reflexes that may occur
(Niedergerke, R., & Orkand, R. K. (1966).
When the string loosely binds, it may exert a
mild pressure or stimulation on the frog's
body or limbs. This external stimulus triggers
a sensory response, which is transmitted
through the peripheral nervous system (PNS)
to the central nervous system (CNS). As a
result of these adaptive processes, the reflexes
gradually become evident, suggesting that the
frog's nervous system can compensate for the
Figure 3. Righting reflexes of a frog that had been leg- initial disruption caused by the loosely bound
tied. Immediate reaction after inhibiting the reflexes. string (McCorry, 2007).
Table 3. Normal frog in inhibition of reflexes When exposed to HCl, the frog
experiment. Reflex time in different HCl experiences pain or discomfort, which
concentrations.
produces an instant reaction or withdrawal of
Frog Reaction
Concentration Within 90 Seconds the frog’s legs as a defensive response to the
of Sodium (Seconds) Average irritant. This pattern, in the experiment,
Chloride (%) Trial Trial Trial indicates that higher HCl concentrations may
1 2 3
lead to shorter reflex time in the frog. The
0.1 3 2 1 2
0.3 3 3 - 2
CNS and PNS of the frog play crucial roles in
0.5 - 4 1 1 mediating reflexes the CNS, consisting of the
brain and spinal cord, processes sensory
The frog was placed dorsal (back) information and coordinates motor responses.
down on a desk and its legs were tied with In the context of reflexes, sensory receptors in
strings, the righting reflexes of the frog the frog's PNS detect stimuli, such as tactile
involved attempts to reorient its body into an or environmental cues, and transmit signals
upright position (Figure 3). However, it has via sensory neurons to the CNS responses
the least attempts compared to the non-tied (Bio, 2023). Upon receiving these signals, the
experimentation of the reflex. Meanwhile, the CNS processes the information and generates
reflex time of frog exposure to various motor responses through motor neurons that

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extend from the CNS to the muscles involved removal of the cerebral hemisphere may
in the reflex action. The efficiency and speed disturb the balance of neurotransmitters
of this neural processing contribute to the within the frog's brain. Neurotransmitters are
observed reflex time (Thau, 2022). The effect chemical messengers that facilitate
of HCl concentration on reflex time may be communication between neurons, and
attributed to several factors. Firstly, HCl is alterations in their levels or availability due to
known to be corrosive and can have irritant the removal of the cerebral hemisphere could
effects on tissues (Wallulis, 2017). Higher impact the functioning and responsiveness of
concentrations of HCl may elicit stronger the remaining neural circuits (Guy-Evans,
sensory stimuli, leading to faster transmission 2021). These combined factors contribute to
of signals to the CNS and subsequently the observed lack of response from the frog in
shorter reflex times. Furthermore, HCl can this experimental context.
potentially affect the excitability of neurons
D. Synaptic Fatigue
within the CNS and PNS. Changes in the
chemical environment caused by HCl may The repetitive stimulation of electric
influence the firing thresholds and response charge in the spinal frog resulted in
properties of neurons involved in reflex temporary loss of reflexes, particularly
pathways, thereby impacting the speed of the affecting the withdrawal reflex. However,
reflex response (Shellhaas, 2019). after some time, a regain of the reflexes
occurred.
Meanwhile, the lack of response from
the frog to the application of sodium chloride As a result of synaptic fatigue, the
on its exposed optic lobes after the removal of reflex activity in the frog may become
its cerebral hemisphere can be attributed to diminished or even temporarily inhibited.
several factors. Firstly, the cerebral The amplitude and speed of the reflex
hemisphere is a critical component of the response may decrease, leading to a slower or
frog's central nervous system, responsible for weaker reaction. However, after a period of
higher-level sensory processing, integration, rest, the synapses can replenish their
and decision-making (Bhavya, 2017). Its neurotransmitter supply, allowing the reflex
removal results in the loss of sensory input, activity to return to its full potential. When
impairing the frog's ability to perceive and the muscle is continuously stimulated to
respond to stimuli. Secondly, the removal of contract, it eventually becomes fatigued and
the cerebral hemisphere likely disrupts the is accompanied by impaired synaptic
neural pathways connecting the optic lobes to transmission (Boyas & Guével, 2011).
the rest of the central nervous system. Synaptic fatigue, also known as short-term
Without intact connections and processing synaptic depression, is a type of short-term
centers, the transmission and interpretation of synaptic plasticity wherein neurons
sensory information from the optic lobes are temporarily lose the ability to fire and
compromised, leading to a lack of response transmit input signals due to activity-
(36.2: Sensory Processes - Transduction and dependent factors. It is believed to function
Perception, 2018). Additionally, the cerebral as a negative feedback mechanism,
hemisphere plays a vital role in coordinating regulating specific forms of nervous system
voluntary motor movements. Its absence activity at a physiological level (Kilpatrick,
causes a loss of motor control, making it 2010). This condition occurs when there is a
challenging or impossible for the frog to temporary reduction in the number of
execute a response to the sodium chloride synaptic vesicles containing
stimulation (Guy-Evans, 2021). Lastly, the neurotransmitters in the synapse. This

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reduction is typically a result of ongoing and Meanwhile, the observed
frequent stimulation of neurons at a high characterization of reflexes was also notable.
frequency. The release of neurotransmitters The highlights from the experiments were:
from the synapse is necessary to transmit the the reflex time became faster as the
signal to the postsynaptic cell (Simons- concertation level of the stimuli increased;
Weidenmaier, Weber, Plappert, Pilz, & reflex inhibition occurs due to existing
Schmid, 2006; Zhang, Gong, Liu, Fa, Ninan, factors such as physical or internal damage,
Staniszewski, & Arancio, 2005). these factors help to impede a reaction; and
lastly, overstimulation was concluded to
Synaptic fatigue occurs when
induce temporary pause on reflex.
neurotransmitter vesicles in the presynaptic
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