THE EFFECTS OF FOREBRAIN ABLATION ON AGGRESSIVE …faculty.uncfsu.edu/tvancantfort/REPRINTS/SFF/Siamese Fighting Fish4.pdffighting fish, Betta splend ens. Fourteen ablated fis h wer

In memorial to Larry Bingham who died from cancer prior to the publication of1

this paper.

THE EFFECTS OF FOREBRAIN ABLATION ON AGGRESSIVEDISPLAY IN MALE SIAMESE FIGHTING FISH (BETTA SPLENDENS)

THOMAS E. VAN CANTFORTDepartment of Psychology

Fayetteville State University

AND

LARRY R. BINGHAM1

Department of PsychologySan Francisco State University

Paper presented at The First Annual Sandhills Regional Psychology Conference,Fayetteville, NC, March 23, 2002

Please consult the authors before citing any portion of this manuscript.

2 FOREBRAIN ABLATION IN SIAMESE FIGHT FISH

The Effects of Forebrain Ablation on AggressiveDisplay in Male Siamese Fighting Fish (Betta Splendens)

ABSTRACTMany investigators have found that the telencephalon in various teleosts plays animportant role in behavior, some concluding that the major role is one of behavioralorganization and others emphasizing its function as one of arousal or facilitation. This study explores the role of the forebrain in aggressive display in male Siamesefighting fish, Betta splendens. Fourteen ablated fish were compared to 14 intact and14 sham operated fish on 10 different behaviors elicited either by a conspecific ormirror stimulus. Ablated fish had longer latencies for fin erection, opercular spreadand air gulps and showed a decrease in frequency of opercular spread comparedto intact and sham operated fish. These results support the hypothesis that theforebrain does not directly mediate specific behavior but serves rather in afacilitatory capacity of arousal and in the integration of the information required toelicit aggressive display behavior.

INTRODUCTION

For many years the early neuroanatomists had categorized the teleost (bony

skeleton fish) forebrain as an olfactory mechanism because of its close relationship with the

olfactory mucosa and its apparent isolation from other sensory systems. Aronson (1963),

however determined that the greater part of the teleost pallium is devoid of secondary

olfactory fibers. This finding has been confirmed by Nieuwenhuys (1970) and is consistent

with the conclusion of the many experimentalists who have found that the forebrain has

multiple functions other than olfaction (Frank, Flood, & Overmier, 1972; Gordon, 1979).

Many investigators (Gordon, 1979; Overmier & Savage, 1974) have found that the

telencephalon in various teleosts plays an important role in behavior, some concluding that

the major role is one of behavioral organization and others (Aronson, 1967) emphasizing

its function as one of arousal or facilitation rather than behavioral organization. Some of


the nonolfactory functions attributed to the teleost forebrain such as deficits in schooling

behavior, active and passive avoidance (Gordon, 1979; Overmier & Savage, 1974), and

interference with reproductive behavior have been reviewed by Ten Cate (1935), Healy

(1957), Segaar (1965), and Aronson (1967).

In most of the experiments reported in the literature, ablation of parts or all of the

forebrain did not result in the loss of any behaviors. Rather, it resulted in changes in the

frequency of occurrence of certain behavioral components or in a loss of efficiency, or

synchrony.

Aggressive behaviors have been implicated as having a specific forebrain mediated

function in the initiation of fighting (Noble & Borne, 1941). It was found that following

partial or complete destruction of the forebrain in the jewel fish, the swordtail, and the

danio, there was a longer latency in fight initiation but that when fighting did occur, it was

just as vigorous as in intact fish. This led Noble and Borne to conclude that actual fighting

behavior must be organized in lower brain levels while the forebrain facilitates initiation

of the behavior. Similar results were obtained by Schönherr (1955) who studied aggressive

behavior in the three-spined stickleback. Decrements in aggressive behavior in this species

were also reported by Segaar (1961) and Segaar and Nieuwenhuys (1963). Hale (1956) also

reported a marked decrease in aggressive behavior following forebrain ablation in the sun

fish.

Shapiro, Schuckman, Sussman, and Tucker (1974) observed that telencephalic lesions

in Siamese fighting fish (Betta splendens)decreased the frequency of gill cover response to


both conspecific stimulus and mirror stimulus. The mean duration of the gill cover

response was not affected by lesioning and did not decrease (habituate) with repeated

stimulus presentation. Shapiro, et al. concluded that behavioral changes may be related

to the extent of the lesion. Since Betta splendens with telencephalic lesions still showed

aggressive displays the centers that control gill cover erection may be located in the

midbrain or hindbrain.

Gorlick (1990) identified parts of the presumptive neural pathway for gill cover

erection in Siamese fighting fish. Motor, motor integration, and sensory areas were

identified in the medulla and mesencephalon. Dilator opericuli motor neurons appeared

to receive all three types of inputs. Gorlick, concluded that connections between motor

areas, and between parts of the reticular formation, may coordinate the performance of gill

cover erection with other behavioral patterns used during aggressive display. Ma (1995)

reported that the opercular dilator muscle consists of three parts: a deep belly and two

superficial bellies. Innervation of this muscle is derived from the maxillary division of the

trigeminal nerve; all three portions are innervated by axons from the same fascicle. Direct

stimulation demonstrates that all bellies can mediate opercular extension. In addition,

Hollis and Overmier (1982) explored the effects of telencephalon ablation on instrumental

learning and Pavlovian conditioning in Betta splendens. They reported that unconditioned

fin erection, gill erection and tail beating to mirror UCS were less frequent in ablates than

in normals or shams. In summary, these various studies seem to show that aggressive

behavior in teleost fish is organized in lower centers, while the forebrain facilitates


responses to the stimuli and integrates the information required to elicit the behavior.

Hale (1956) reported that forebrain ablation reduces aggressive behavior in some

teleost no one has studied the effects of forebrain ablation on aggressive behaviors in

Siamese fighting fish (Betta splendens) towards it’s conspecific (another male Siamese

fighting fish). Aggressive display responses in the male Betta splendens have been

extensively described (Braddock & Braddock, 1955; Simpson, 1968) and include approach;

extension of the gill covers and gill membranes; erection of the dorsal, ventral, medial, and

caudal fins; intense deepening of body and fin color; and the characteristic orientation and

undulatory movements. The aggressive display can be elicited by a conspecific male or

one’s own mirror image ( Figler 1972; Johnson & Johnson, 1973; Lissman, 1933 Meliska,

Meliska & Peeks, 1980).

A variety of factors appears to influence aggressive display response in the male

Betta splendens. Siamese fighting fish shown a facing posture model (used mainly in

aggressive contexts) are more aggressive than Siamese fighting fish shown a broadside

posture model (used in many social context) (Halperin, Giri, & Dunham, 1997). Social

isolation also plays a role in aggressive display response of male Betta splendens. Socially

isolated Siamese fighting fish showed weaker aggressive display to the first model seen

after social isolation in a series of novel models than nonisolate. The aggressive display

was progressively weaker the longer the social isolation. Whereas, isolates displayed more

strongly than nonisolates to the last model of the series, and display intensity became

monotonically stronger with longer social isolation (Halperin, Dunham & Ye, 1992).


Halperin and Dunham (1994) have also demonstrated that social overstimulation reduces

subsequent aggression in male Betta splendens.

There has been some question about the strengths of conspecific versus mirror

eliciting stimuli. For example, Johnson and Johnson (1973) found that a mirror stimulus

elicited greater responses than did a conspecific stimulus. Figler (1972) reported that

unhabituated male Betta splendens was the strongest releaser of aggressive display in

Siamese fighting fish, followed successively by mirror, habituated male, and model.

Meliska, Meliska, and Peeks (1980) identified two groups of Betta splendens, those that

responded with high levels of reactivity to a mirror or those that responded with low

reactivity to a mirror. Subjects that displayed at the mirror for short durations (low

reactivity) were matched for combat with either other short-duration displayers (the Short-

Short group), or long-duration (high reactivity) displayers (the Long-Short group). In

combat, Short-Short pairs fought less intensely than Long-Short pairs; but within Long-

Short pairs, Short- and Long-duration displayers did not differ in combat vigor.

Correlations between combat behavior and precombat mirror display measures

tended to be positive, but were mostly small and nonsignificant. Correlations with

postcombat mirror displays were greater. These results imply that mirror-elicited threat

behaviors provide a partial predictor of actual combat aggression in Siamese fighting fish

(Meliska, Meliska, & Peeks, 1980). Some of the differences found in the above studies may

have been due to the different experimental paradigm or utilization of different response

measures.


The purpose of this study was to examine the effects of forebrain ablation on the

aggressive display in male Siamese fighting fish. A mirror and conspecific stimuli were

utilized, and the effects of ablation on several different measures of aggressive display

involving latency, frequency, and duration of behavior were compared.

METHODSSubjects

Sixty adult male Betta splendens were used. The subjects had color variation as was

expected and is common even within the same brood. Figler (1972) reported color

variation is not a factor in the elicitation of aggressive responses. Subjects were housed in

specially constructed plexiglass aquaria such that each fish was chemically and visually

isolated from all other fishes. The room that housed these aquaria had a controlled light-

dark cycle, lights on from 08:00 hours to 20:00 hours.

Surgery

Subjects were anesthetized in Sandos MS 222 (m-Aminobenzoic acid ethyl ester-

Methansulfate salt), 1 part to 4 parts water, and then placed in the stereotaxic apparatus,

the scales were scrapped away and a medial incision approximately 2 mm corresponding

with the cranial and caudal limits of the eye was made. A transverse incision was made

approximately 3 mm corresponding with the caudal limits of the eye and at 15 degrees to

the coronal plane of the subjects and about 2 mm deep to sever the forebrain from the

midbrain. The forebrain was then aspirated by means of positively applied suction from

a water aspirator. Sham operated fish were treated in the same fashion except their brains


were left intact, only the skull incisions were made.

Apparatus

The test aquaria had a clear plexiglass partition in which one compartment housed

the stimulus and the other compartment housed the experimental fish. Recording of

latencies, frequencies and durations of behaviors was accomplished with a Rustrack-4-

channel event recorder.

Procedures

Subjects were randomly assigned to one of three experimental groups; ablated

(N=30), sham operated (N=15), and intact (N=15). The subjects from each experimental

group were randomly assigned to one of two stimulus group; mirror stimulus or

conspecific stimulus and observed for 10 minutes.

Table 1 lists the behavioral components of the subject’s threat display that were

simultaneously recorded during the ten minute observation period. Also included in these

Table 1. Measurement and definition of behaviors

Latency of fin erection (LFE) latency in seconds to caudal fin erectionLatency of opercular spread (LOS) latency in seconds to opercular spreadLatency of air gulps (LAG) latency in seconds to air gulpsFrequency of fin erection (FFE) number of times the caudal fin was erect during the sessionFrequency of opercular spread (FOS) number of times the operculum was spread during

the sessionFrequency of air gulps (FAG) number of times the fish gulped air during the sessionDuration of fin erection (DFE) total time in seconds that the caudal fin was erect during the

sessionDuration of opercular spread (DOS) total time in seconds that the operculum was spread

during the session

measurements were two derived scores. Mean duration of fin erection, mean time in

seconds that the caudal fin was erected during each occurrence in a session. The mean time


was obtained by dividing total duration by frequency of occurrence in a session. And

mean duration of opecular spread, mean time in seconds that the operculum was during

each occurrence in a session. The mean time was obtained by dividing total duration by

frequency of occurrence in a session.

At the end of testing the subjects were sacrificed in the anesthesia solution for

approximately thirty minutes and then placed in a 10 percent formalin solution for 48

hours. The fish were then placed in the stereotaxic apparatus, their skulls opened and the

brains photographed to determine the extent of ablation.

RESULTS

After the surgery, 14 ablated Betta splendens in their home aquaria exhibited

behaviors that were indistinguishable from the behaviors of the control fish. That is, their

fleeing from the net, location and consumption of food, and general locomotor activity

were equal to the intact and sham operated fish. Those ablated fish that showed any gross

sensory or motor impairment were excluded from this study. The results from this study

were obtained from those 14 ablated fish, 14 randomly selected sham operated, and 14

randomly selected intact fish.

A two-way analysis of variance (see Table 2) was used to determine the effects of

surgery (ablation, sham operated, and intact) and eliciting stimuli (conspecific vs mirror)

between the groups. Removal of the telencephalon in male Siamese fighting fish resulted

in significantly greater latencies to respond as compared with intact and sham operated

control groups on fin erection, opercular spread, and air gulps (figures 1 - 3).


Table 2. Summary table for two-way analysis of variance.

Effect LFE LOS LAG FFE FOSSurgery df 2/36 2/36 2/36 2/36 2/36

F 5.04 3.90 10.77 1.47 10.71 p .05 .05 .01 NS .01

Stimulus df 1/36 1/36 1/36 1/36 1/36 F 1.44 9.61 0.25 0.18 3.23 p NS .01 NS NS NS

Surgery df 2/36 2/36 2/36 2/36 2/36 X F 0.17 1.12 0.81 0.86 1.09Stimulus p NS NS NS NS NS

Effect FAG DFE DOS MDFE MDOSSurgery df 2/36 2/36 2/36 2/36 2/36

F 11.81 17.69 6.10 10.02 0.08 p .01 .01 .05 .01 NS

Stimulus df 1/36 1/36 1/36 1/36 1/36 F 0.65 1.77 2.73 4.80 0.43 p NS NS NS .05 NS

Surgery df 2/36 2/36 2/36 2/36 2/36 X F 0.10 0.11 0.14 0.91 0.22Stimulus p NS NS NS NS NS

F .05 (2/36) = 3.26 F .05 (1/36) = 4.11F .01 (2/36) = 5.25 F .01 (1/36) = 7.40


Figure 1. Mean latency to fin erection.

Figure 2. Mean latency to opercular spread.


Figure 3. Mean latency to air gulp

Frequency of air gulp and frequency of opercular spread was significantly greater

in the intact and sham group as compared to the ablated group (figures 4 & 5). The

decrease in the frequency of opercular spread for the ablated group is consistent with the

findings of Shapiro et al. (1974). A two-way analysis of variance shows no significant

difference between ablated and the two other groups on frequency of fin erection (figure

6).


Figure 4. Mean frequency of opercular spread.

Figure 5. Mean Frequency of air gulp.


Figure 6. Mean frequency of fin erection.

Duration of fin erection and opercular spread were significantly greater in the intact

and sham groups as compared to the ablated group (figures 7 & 8).

Figure 7. Total duration of fin erection.


Figure 8. Total duration of opercular spread.

Removal of the forebrain resulted in a significant decrease in mean duration of fin

erection per occurrence and the conspecific stimulus elicited a significantly higher mean

duration of fin erection than that exhibited to the mirror stimulus (figure 9). No significant

difference was found between groups on mean duration of opercular spread (figure 10).

Again the results on mean duration of opercular spread for the ablated group are similar

to the results found in Shapiro et al. (1974) study.


Figure 9. Mean duration of fin erection per occurrence.

Figure 10. Mean duration of opercular spread per occurrence.

Finally, there was no significant difference on eight of the 10 dependent measures

for the effect of the eliciting stimuli. Latency to opercular spread was consistently longer

for mirror stimulus as compared to conspecific stimulus across ablated, intact and sham


operated fish.

DISCUSSION

In agreement with other studies, no obvious sensory or motor impairment was

observed in the ablated fish in their home aquaria, and their behavior was

indistinguishable from the behavior of the control fish. That is, fleeing from the net,

location and consumption of food, and general locomotor activity were equal to the intact

and sham operated fish. Therefore, deficits in the ablated fish to be described below could

not be attributed to any gross sensory or motor impairment.

The latency data from this study indicates that the forebrain plays a role in the

initiation of aggressive display, since the ablated group showed significantly higher

latencies on all three dependent measures. The forebrain also seems to facilitate aggressive

display as revealed by the low frequency of opercular spread and low frequency of air gulp

in the ablated group when compared to the intact and sham control groups. This

facilitation is also supported by the duration scores since the ablated fish showed a shorter

total duration on all of the measured components of aggressive display. Interestingly, the

ablated group showed a shorter duration of fin erection despite their somewhat higher

frequencies.

Of the eight dependent measures involving latency, frequency, and duration, the

ablated group differed significantly in seven of these eight comparisons. This strong

concordance among the various responses would indicate a basic underlying deficit

resulting from forebrain lesion, rather than a discrete response impairment. It should also


be noted that these behaviors, although disrupted, were not eliminated and this is

consistent with earlier findings (Hosch, 1936; Kamrian & Aronson, 1954; Hale, 1956).

Furthermore, the results support the hypothesis of Aronson (1967) that the forebrain

does not directly mediate specific behavior but serves rather in a facilitatory capacity of

arousal and integrates the information required to elicit aggressive display behaviors. This

hypothesis would also fit in with the parallels drawn by many authors between the

anatomical make-up of the teleost forebrain and the limbic system of mammals. In general

terms, the limbic system in mammals is thought of as a nonspecific modulator of behavior

patterns organized in other parts of the brain (Gloor, 1960) and as a regulator of awareness

(Douglas & Pribram, 1966).

Gloor (1960) wrote that the limbic system provides “motivational mechanisms

which normally allow selection of behavior appropriate to a given situation.” Aronson

continues this line of thinking by postulating a neural mechanism whereby reverberating

circuits in the lower centers deteriorate when no longer primed by the forebrain

descending influences.

Deterioration of behavior as a result of forebrain ablation might also be the result

of depletion of a biogenic substance which is normally secreted by the forebrain and which

impinges on lower brain centers. For example, Schildkraut and Kety (1967) suggest that

norepinephrine is a mediator of a nonspecific state of arousal. In addition Kusunoki and

Masai (1966) have found substantial quantities of monoamine oxidase in the olfactory

bulbs, and in certain pallia and subpallial portions of the goldfish brain. The monoamine


oxidase in the forebrain may indicate the presence of norepinephrine or other

catecholamines, and ablating the forebrain may disrupt a feedback system involved in a

neurohumoral arousal process.

Ablation studies, in general, remove large portions of the brain to learn what

function that particular part of the brain regulates. In the present study the entire frontal

lobes were removed and we found disruption in the aggressive display of male Siamese

fight fish. It is important to note that there was no loss of any aggressive display behavior,

but the behavior was disorganized. By using discrete lesioning technique we may better

understand the nature of the role that the forebrain plays in the regulation of aggressive

display in the male Siamese fighting fish.

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THE EFFECTS OF FOREBRAIN ABLATION ON AGGRESSIVE …faculty.uncfsu.edu/tvancantfort/REPRINTS/SFF/Siamese Fighting Fish4.pdffighting fish, Betta splend ens. Fourteen ablated fis h wer