A M . ZOOI.OCIST, 12:507-512 (1972).
Cooling as Reinforcing Stimulus in Aplysia
PAUL DOWNEY AND BEHRUS JAHAN-PARWAR
Biology Department, Clark University, Worcester, Massachusetts 01610
and Worcester Foundation for Experimental Biology,
Shrewsbury, Massachusetts 01545
SYNOPSIS. An experiment was carried out to investigate the role of temperature in the
previously reported reinforcing effect of an increase in sea water level in Aplysia. In
the present experiment, it was found that the reinforcing effect of water level change
on rod-pressing behavior in Aplysia depends on a decrease in temperature associated
with water level change. In order to study modification of rod-pressing behavior produced by contingent increase in water level and decrease in temperature, the rate
and latency of rod-press responses in experimental animals were compared with those
of yoked control animals exposed to non-contingent water level and temperature
change. Higher response rates and shorter response latencies were obtained from experimental over yoked control animals, but only the shorter latencies of experimental
animals were attributed to a behavioral change resulting from contingent water level
and temperature reinforcement..
water level.
In the experiment of Lee (1969), modThere is considerable interest in demon- ification in the behavior of Aplysia was
strating learning behavior in the marine determined by using yoked control procemollusc, Aplysia. The major cause of this dures to compare experimental animals exinterest is the relatively small number and posed to contingent water level change
large size of neurons in the Aplysia ner- with yoked control animals exposed to nonvous system (Frazier et al., 1967), which contingent changes in water level. Obmakes this animal a promising model for served differences between experimental
and control animals were attributed to
studying the cellular basis of learning.
Several investigators have reported clas- conditioning associated with contingent
sical conditioning in Aplysia (Jahan- water level reinforcement.
Parwar, 1970; Lickey, 1968), but there has
In the following experiment, yoked conbeen only one report on operant condi- trol procedures modeled after those of Lee
tioning in this animal (Lee, 1969). It was (1969) were used to further evaluate the
shown in that experiment that an increase conclusion that modification in the behavin sea water depth from a level that only ior of Aplysia occurs when water level rehalf covered the animal to one in which inforcement follows that behavior in time.
the animal was completely covered could
be used to reinforce an operant response in
METHOD
Aplysia. What was not determined was
Subjects. Thirty Aplysia californica,
the particular stimulus aspect of changing
wafer level that produced the observed re- weighing between 120 and 395 grams, were
inforcing effect. The possibility investi- used in this experiment. Between experigated in the present experiment is that the mental sessions, the animals were kept in
reinforcing effect of contingent water individual living chambers supplied with
level change results from a decrease in filtered and aerated sea water, cooled to 14
temperature associated with an increase in ± 0.5°C. The animals were fed ad libitum
portions of dried seaweed (Dulse) which
Supported by P.H.S. Grant NS08868 and Careeer typically caused the animals to gain weight
Development Award K4-HD-5 178 to B. JahanParwar and by N.I.M.H. Research Traineeship at an average rate of 15 grams per week
(range: 0 to 38 grams per week).
MH 10625 to Paul Downey.
INTRODUCTION
507
508
PAUL DOWNEY AND BEHRUS JAHAN-PARWAR
WATER /MLE TS
SPOTLIGHT
HEAT LAMP
I PROGRAMMING
EQUIPMENT
FIG. 1. Schematic overhead view of the single compartment experimental enclosure (A), the two
compartment enclosure (B) , and a side view of the
single compartment enclosure showing the rod
manipulandum and the overhead spotlight and
heal lamp (C) .
Apparatus. The experimental chamber
was a plastic enclosure with an open mesh
floor which was partially submerged in a
30 X 30 X 18 c m t a n k filled w i t h s e a w a ter. A water-filled syringe located below the
enclosure was used in the manner of a hydraulic cylinder to raise or lower the enclosure relative to the surrounding tank
(see Fig. 1). An infusion-withdrawal
pump (Harvard Apparatus Model 902)
was used to drive the syringe to produce
changes in the depth of sea water in the
experimental enclosure between 3 cm and
9 cm in approximately 6 sec. Water in the
tank surrounding the experimental enclosure was continuously filtered, aerated,
and circulated through the chamber at the
rate of 2.5 lifers per minute. The experimental enclosure was located in a temperature controlled room maintained at 14 ±:
0.5°C.
Two experimental enclosures were used
in this experiment. One consisted of a sin-
gle compartment having curved walls and
measuring 18 cm in diameter. Next' to the
curved wall, near the sea water outlet, was
a vertical rod manipulandum extending
down to the floor of the experimental enclosure from a circular contact switch, 12
cm above the enclosure floor. This manipulandum allowed contact closure to be
made by displacement of the vertical rod
in any direction on the horizontal plane.
Each contact closure produced a brief (0.5
sec) flash from a spotlight (American Optical Model 651) located over the manipulandum. This served as a feedback stimulus for rod-press responses. Forty cm above
the tank floor was a heat source (General
Electric Infrared Lamp). Prolonged use of
the heat lamp produced temperatures of
17 to 19=C above the surface of the water
and raised the water temperature at the
surface from 14°C to no more than 163C.
The heating effect of the lamp on the
darkly pigmented skin of Aplysia would be
COOLING AS REINFORCEMENT IN
Ht*T Law •
\N\!
AC33»«
AC 34o«
-(•I
V- ^
\.ktip or*)
es.ss;
/V 4
AC 3 7 ^ .
AC 38 oo
CO
2
O
Q.
Ui
It
SESSIONS
FIG. 2. The response rates (responses per minute)
of experimental animals (closed circles) and
control animals (open circles) over sessions in
which the responses oE experimental animals were
followed by one of four changes in heat lamp and
water level stimuli: Increase in water level either in
the presence (heat lamp on) or absence (heat
lamp off) of the heat lamp stimulus, increase in
water level and extinction of the heat lamp, or
extinction of the heat lamp alone.
greater than that indicated by these measurements.
The second type of enclosure used in
these experiments consisted of two oval
compartments measuring 18 X U cnl anc^
each equipped with a rod manipulandum.
The surrounding tank and other peripheral equipment (heat lamp, spotlights, etc.)
were the same as those described above.
Events in the experimental chamber
were remotely controlled by solid state
programming equipment (Massey Dickinson), and responses were recorded on magnetic counters and cumulative recorders.
Procedure. At the start' of daily 1-hr sessions, animals were placed in shallow sea
water (3 cm deep) facing away from the
response rods. For some animals, sessions
began with the overhead heat lamp turned
on, and for other animals the heat lamp
was turned off. A response was recorded
each lime a response rod was pressed, and
each response produced a brief flash from
the overhead spotlight. Each response also
began a 30 sec reinforcement period, and
additional responses extended these peri-
Aplysia
509
ods until 30 sec elapsed without a response. Experimental animals were run
under one of four experimental conditions
of heat lamp and water level reinforcement. In the "heat lamp" condition, water
level remained at the 3 cm level
throughout each session, and the heat
lamp was t'urned off during reinforcement
periods. In the second experimental condition — the "heat lamp and water level"
condition — water depth was increased
from 3 cm to 9 cm, and the heat lamp was
extinguished during reinforcement periods. The third condition was the "water
level (heat lamp off)" condition in which
water depth was increased to 9 cm while
the heat lamp remained off throughout
each session. The last' experimental condition — "water level (heat lamp on)" — was
added to control for possible behavioral
effects of change in level of illumination
associated with extinguishing the heat
lamp stimulus. Under this condition, the
heat lamp remained on throughout each
session. Thus, increase in water level that
accompanied reinforcement periods produced a substantial decrease in temperature but had little effect on level of illumination.
A separate group of four experimental
animals paired with four control animals
were run under the "heat lamp and water
level" condition. In addition, these animals
were run in a series of probe trials used to
determine difference in the response
latencies of experimental and control animals. In probe trials, experimental and
control animals were placed in comparable
locations in the two compartments of the
experimental enclosure, facing away from
the response rods. A relative measure of
response latency was obtained by recording
which of the two animals made the first
response in each probe trial and calculating the percentage of probe trials in which
each animal made the first response.
These trials were carried out' twice daily:
once at the start of each session, and once
about 30 min into the session.
Two of the experimental animals were
run individually in the single compart-
510
PAUL DOWNEY AND BEHRUS JAHAN-PARWAR
TABLE 1. Average response rates (responses per minute) of experimental animals under response-contingent changes in water level alone, heat lamp alone, or in both heat lamp and water
level stimuli; and of control animals exposed to non-contingent stimulus changes. The number
of sessions over which each average was taken is shown in parentheses.
Contingent
Animal
AC12
AC13
AC33
AC34
AC37
AC38
AC21
AC22
AC23
AC24
AC25
AC26
AC15
AC16
AC17
AC1S
AC19
AC20
AC49
AC50
Average
Water level
(heat lamp off)
Heat lamp
Heat lamp and
water level
4.5 (6)
3.9 (8)
3.9 (6)
5.2 (2)
2.3 (6)
4.1 (2)
1.3 (9)
1.4 (9)
Non-contingent
(control)
0.6 (6)
0.7 (6)
2.7 (11)
1.3 (11)
3.5 (7)
1.2(7)
2.3 (8)
1.6 (8)
3.9 (17)
3.7 (17)
6.1 (9)
1.4(9)
2.7 (10)
1.4 (10)
2.2 (11)
1.35
3.06
ment enclosure shown in Figure 1A. The
remaining animals were run in pairs (one
experimental and one control) in the two
compartment enclosure illustrated in Figure IB. The responses of control animals
were recorded, and each response produced a flash of light from the overhead
spotlight. But changes in water level and
heat lamp stimuli depended on the responses of the experimental animal paired with
each control animal.
4.0
1.4 (11)
1.26
(represented by open circles). Figure 2
shows that response rates above control
levels were obtained from experimental
animals under the three conditions that
included exposure to the heat lamp stimulus. These were the "heat lamp," "heat
lamp and water level," and "water level
(heat lamp on)" conditions. In the absence of the heat' lamp stimulus as in the
"water level (heat lamp off)" condition,
response rates remained near control
levels.
RESULTS
Listed in Table 1 are the mean response
rates obtained from animals exposed to
Figure 2 shows response rates (responses three of the experimental (contingent)
per minute) obtained under each of the conditions, and control (non-contingent)
four experimental conditions. The upper conditions. Included in this table are data
graphs show results from experimental an- from animals shown in Figure 1 and simiimals AC 12 and AC 13 run in the single lar data obtained from additional animals.
compartment enclosure. These graphs are These data indicate the magnitude of
divided by vertical lines into segments and difference in response rates obtained from
labeled as to the reinforcing stimulus con- animals exposed to the heat lamp stimulus
ditions used in sessions falling within each (columns labeled "Heat lamp" and "Heat
segment. The lower graphs are from ex- lamp and water level") and response rates
perimental animals AC33, AC37, and obtained from animals run under "water
AC 13 (represented by closed circles) and level (heat lamp off)" and control condicontrol animals AC34, AC38, and AC44 tions. The latter group of animals pro-
COOLING AS REINFORCEMENT IN
TABLE 2. The percent of probe trials in which experimental animals designated in Column 1 produced a shorter latency from the start of a session
to the first response in that session (Probe 1), or
from the start of a probe within a session (Probe
2) and the total percent of probes in which shorter
latencies were obtained from, experimental animals.
Experimental
animals
AC 71
AC 75
AC 77
AC 81
Number of
probe trials
Percent of
Probe 1
trials
Percent of
Probe 2
trials
22
28
22
14
64
57
29
71
91
57
55
57
duced response rates of 1.7 responses per
minute or less (average: 1.26 to 1.35 responses per minute), while animals exposed to the heat lamp stimulus produced
rates of 2.2 responses per minute or greater
(average: 3.06 to 4.0 responses per minute).
Table 2 contains the results of probe
trials used to compare response latencies of
experimental and control animals after a
period of exposure to contingent (experimental) or noncontingent (control) conditions. A difference in response latency is
indicated by shorter latencies from experimental animals in more than the 50% of
probe trials that would occur on the basis
of chance. Table 2 shows that' three of the
four of experimental animals in Probe 1
trials and all four experimental animals in
Probe 2 trials had shorter latencies in
more than 50% trials.
DISCUSSION
This experiment shows that the reinforcing effect of increase in water level on
rod-pressing behavior in Aplysia depends
on the cooling effect produced by increasing water level. Under all conditions that
included decrease in temperature as one of
the stimulus changes following a response, the rate of rod-pressing behavior
was substantially higher than that of control animals exposed to non-contingent
changes in temperature and water level.
Contingent water level change in the absence of a drop in temperature produced
response rates near control levels.
Aplysia
511
Extinguishing the heat lamp stimulus in
the present experiment produced a decrease in level of illumination as well as a
drop in temperature. The "water level
(heat lamp on) " condition was included
in the present experiment to control for
possible behavioral effects of change in illumination. This procedure allowed substantial reduction in temperature (by immersing the animal in sea water), while
only slightly decreasing the level of illumination. The high response rate obtained
under these conditions shows that the
effect of the heat lamp stimulus on the
behavior of Aplysia results from its effect
on temperature.
Evidence for behavior modification in
Aplysia was obtained in measures for bofli
response rate and response latency in the
present experiment. A change in response
rate is shown by the higher responses per
minute of experimental over yoked control
animals. However, it is possible that such
differences are the result of an unconditioned change in activity level associated
with changes in temperature and water
level. For example, increase in activity
level occurring at the termination of reinforcement could produce higher response
rates in experimental animals. Since the
occurrence of reinforcement depends on
the proximity of experimental animals to
the response manipulandum, the likelihood of random responses resulting from
an increase in activity level is greater for
experimental than for control animals.
Differences between experimental and
control animals in response latency, however, could not be attributed to nonspecific
effects of reinforcement since the measured
behavior (latency) is completed prior t'o
reinforcement. Moreover, since the latencies obtained in Probe 1 trials in the
present experiment were carried out at the
start of each session, the latency measurement was not closely preceded by reinforcement. Thus, the effect of reinforcement
on behavior in Probe 1 trials results from
conditioning rather than from short' term
unconditioned effects of reinforcing stimuli.
512
PAUL DOWNEY AND BEHRUS JAHAN-PARWAR
REFERENCES
Frazier, W. T., E. R. Kandel, I. Kupfermann, R.
Waziri, and R. E. Coggeshall. 1967. Morphological and functional properties of identified neurons in the abdominal ganglion of Aplysia californica. J. Neurophysiol. 30:1289-1351.
Jahan-Parwar, B. 1970. Conditioned responses in
Aplysia californica. Amer. Zool. 10:287. (Abstr.)
Lee, R. M. 1969. Aplysia behavior: effects of
contingent water level variation. Conimun. Behav.
Biol. 3:157-164.
Lickey, M. 1968. Learned •behavior in Aplysia vaccaria. J. Comp. Physiol. Psychol. 66:712-718.