Species typical display behavior following stimulation of the reptilian striatum

Physiology & Behavior, Vol. 29, pp. 615--620. Pergamon Press, 1982. Printed in the U.S.A.

Species Typical Display Behavior Following Stimulation of the Reptilian Striatum ROBERT

S. TARR

Department of physiology, Chicago College of Osteopathic Medicine 1122 East 53rd Street, Chicago, IL 60615 R e c e i v e d 4 M a r c h 1982 TARR, R. S. Species typical display behavior following stimulation of the reptilian striatum. PHYSIOL. BEHAV. 29(4)

615-620, 1982.--Seventy unanesthetized, unrestrained western fence lizards (Sceloporus occidentalis) were electrically stimulated through implanted electrodes. Behavior elicited included the species typical assertion display, elements of the challenge display and elementary locomotor responses: circling, rolling and curling. The assertion and challenge displays were elicited from telencephalic sites whereas the elementary locomotor effects were eficited from electrodes in the brain stem. Assertion displays were consistently elicited in 25 animals at an average threshold current of 46 p.A. Sites showing the lowest threshold and greatest reliability were tightly clustered in the striatum and nucleus accumbens. Challenge behavior was elicited in eleven animals at an average threshold of 58/xA. Seven of the animals with challenge responses had electrodes in a small area anterior and dorsal to nucleus sphericus. The implications of these results are discussed relative to current views of the comparative neuroanatomy of the basal ganglia and relative to the basic functional organization of the vertebrate central nervous system. Nucleus accumbens

Striatum

Lizard

Stereotyped behavior

S T U D I E S utilizing electrical stimulation of the reptile brain have resulted in conflicting data [26]. This appears to be the consequence of several factors. First, the early (prior to 1968) studies lacked precise quantification of current parameters and histological verification of stimulus site [15,26]. Secondly, most of these early studies were attempts to find a " m o t o r c o r t e x " or subcortical motor area, as predicted from the prevalent anatomical views at that time [15]. Thus some investigators found the dorsal cortex (then thought to be the homologue of mammalian neocortex) an active motor area [1,2], whereas others did not [10,26]. Subcortical nuclei (thought to be the homologue to the basal ganglia) likewise give mixed results [ 10,15]. The third factor that has hindered studies on reptiles has been an inadequate appreciation of the known behavioral complexity of these animals. Thus the behavioral categories tested were often v e r y broad or were components of numerous separate behaviors [7, 10, 26, 29]. Several definitive studies have been done however. Schapiro and Goodman [28], utilizing constant current stimulation parameters and accurate histological marking, demonstrated a rigid stimulus bound circling from stimulation of the optic tectum. Another investigator [17] using geckos elicited threat display related vocalizations from regions around torus semicirculaxis. Stimulation of nucleus profundus mesencephali [30] in collared lizards resulted in low threshold defensive posturing and dewlap (a skin fold of the throat used in iguanid lizard aggressive behavior) extension. The past decade has produced substantial progress and revision in the area o f comparative neuroanatomy [21,26]. The classic view, based on the gross topographic relation-

Assertion display

Social behavior

ships of structures, has been displaced as histochemical and connectional studies reveal a rational basis for determining homologies [9, 11, 25]. Thus functional studies on reptiles utilizing this anatomic model make comparisons with mammalian work a valid and feasible operation. In the present study various targeted structures were systematically electrically stimulated in the iguanid lizard, Sceloporus occidentalis. The use of an iguanid lizard allows for a precise analysis of complex species typical responses since the social behavior of these animals has been thoroughly studied [14, 20, 31]. METHOD

Adult male western fence lizards (Sceloporus occidentalis) were obtained from dealers in California and housed in groups of 15 to 25 in cages 30 by 30 by 60 cm. Animals were anesthetized by intraperitoneal injection of 0.1 cc per 20 grams body weight of 5 mg/cc nembutal solution. The lizard was then positioned in a modified K o p f rat stereotaxic and two electrodes positioned by coordinates relative to the parietal eye [12,31]. Small holes were drilled with a dental drill and the electrodes were lowered into place. A ground electrode was inserted under the skin in the dorsal neck region. The active electrodes and ground were then cemented in place with dental acrylic. Upon recovery the lead wires were attached to a light spring suspended from an arm over the testing cage. Active electrodes were 000 stainless steel insect pins insulated with epoxylite insulator and dried point up in an oven.

C o p y r i g h t © 1982 P e r g a m o n Press--0031-9384/82/100615-06503.00/0

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Exposed tip length ranged from 0.05 mm to 0.1 mm. The ground electrode was an uninsulated 00 insect pin with at least 5 mm of shaft implanted under the skin. The stimulus was a biphasic square wave, 1 msec pulse duration, delivered from a Grass S-88 stimulator through two constant current stimulus isolation units. Current was monitored by an oscilliscope (see [5]). Pulses were delivered at 50 pulses per second and at 5 pulses per second. The pulse train lasted from 10 to 30 seconds with at least a double intertrain rest period. The testing cage was a 30 by 45 by 60 cm glass aquarium with light provided by a 60 watt shielded incandescent bulb suspended in one corner of the cage. The light produced a temperature gradient on the cage floor ranging from 22°C to 60°C. This gradient was used by the lizard to behaviorally maintain its preferred body temperature [18]. Three sides of the test cage were covered with paper, the fourth side was brought next to a one way mirror (angled in a way that prevented the animal from seeing its reflection) for observation from a darkened booth. On day one (at least 24 hours postoperative) the lizard was stimulated, while alone in the cage, until behavior was consistently evoked. Threshold determination for each electrode was verified over a two day period. On day two mealworms and a male conspecific were added to the test cage and stimulation was repeated. Evoked behaviors were categorized according to the following classification scheme: I. Complex, goal oriented behavior a. Eating b. Species typical assertion display (The assertion display is a highly stereotyped species recognition display. It is usually not directed towards a conspecific and is characterized by a repetitious vertical bobbing motion resembling a series of " p u s h u p s " ) c. Species typical challenge display posturing (an aggressive directed display) d. Fighting with the conspecific e. Exploratory behavior (tongue flick, etc.) 2. Elementary locomotor effect a. Circling (ipsiversive or contraversive) b. Rotation in the frontal plane (tilting or rolling) c. Movements in the vertical direction (curling in a ball) A written description of evoked behavior was also obtained for each animal with attention paid to the individual sequence and form of the behavior evoked. After all testing was completed the electrode tip sites were marked with an 80/~A DC current for 30 seconds. The animal was sacrificed and the brain stored in 10% Formalin, embedded in paraffin, sectioned at 14 microns in the frontal plane and stained with the Prussian blue iron stain. Drawings made from a microprojector were used to reconstruct the locations of the electrodes. RESULTS

A total of 70 animals were treated as described above. One hundred twenty marked electrode tip sites were identified. Complex goal oriented behaviors elicited included: assertion display (25 animals); challenge display posturing consisting of a raised posture and either lateral compression of the sides, exposure of the ventral blue aggressive markings or extension o f the dewlap (11 animals); and exploratory behavior without any display (24 animals). In no animal did stimulation elicit eating or fighting with the conspecific. Elementary locomotor effects elicited by stimulation in-

cluded: ipsiversive circling (10 animals), contraversive circling (4 animals), tilting and rolling (10 animalsL and curling (7 animals). An additional 7 lizards manifested combinations of the above locomotor effects, such as curling to the left and moving to the right or such as spiraling up into a position where the front legs were removed from the cage floor. In all cases the behavior was elicited on both testing days. Thresholds were often slightly higher on day two. The particular behavior elicited depended on electrode placement. Since this localization is clarified by other stimulus parameters (such as threshold needed to elicit the behavior) separate behaviors are listed below and electrode placement, current used and the reliability of the response are described for each category of elicited behavior.

Assertion Display The 25 electrode sites giving assertion displays are shown in Fig. 2 and 3. There was a concentration of points along a column of cells in the region near the tip of the lateral ventricle: That is, nucleus accumbens and paleostriatum [21]. Several active sites were found in the posterior telencephalon lateral and dorsal to this cell column. Only 1 site outside of the telencephalon (in the hypothalamus) produced the assertion display. Threshold currents ranged from 5/zA (10 t~A peak to peak) to 100/~A. The average threshold value was 46 tzA. The consistency of the evoked response (percent of applied stimuli that resulted in a display) ranged from 22% to 91% with a mean of 50%. In one animal the assertion display was elicited by each of 22 consecutive stimuli. Analysis of electrode tip site by consistency reveals that the sites that showed a consistency of 50% or greater (12 animals) were tightly clustered near the tip of the ventricle in a column extending from 58% to 75% (where the distance from the anterior pole of the telencephalon to the posterior pole equals 100%) of the way through the telencephalon. In 18 of the animals the display occurred at stimulus offset or a few seconds after termination of the stimulus and was preceded by a generalized arousal characterized by the animal looking around the cage, taking a few steps then displaying. In 4 lizards the display followed termination of current flow but without any looking or walking preceding it. Two lizards gave assertion displays at the start of or during current flow. Table 1 shows a summary of sites that produced the assertion display. Not shown are a few sites in nucleus accumbens, septal nuclei, posterior pole of the telencephalon and hypothalamus.

Challenge Behavior Eleven sites produced forms of challenge behavior. The locations of these sites is shown in Fig. 3. Seven of the eleven points were tightly clustered in the medial portion of the posterior basal telencephalon, in a region just anterior and dorsal to nucleus sphericus. The average threshold current needed to elicit challenge behavior was 58 ~ A , with a range of 30/~A to 120 ~A. The form of challenge behavior seen varied somewhat in the different animals. In all cases the lizard rose up on all four limbs within 3 seconds of stimulus onset and remained in this raised posture until after stimulus offset. Often this posture was held f o r s e v e r a l seconds after termination of the stimulus. Additional behaviors were at times present: in 6 animals the sides were drawn in (described as "lateral compression" [20]) resulting in an enlargement of the body profile and in exposure of blue ventral aggressive marking; 3 animals turned a quick half circle

STRIATAL STIMULATION

IN LIZARDS

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RO'r

FIG. 1. Cross section reconstruction through the brain of Sceloporus occidentalis. ABBREVIATIONS ACC ADVR AT CER DL DM GEN HYPO LFB LN LPA MC MPA NIII NIV NDC NPM

Nucleus Accumbens Anterior Dorsal Ventricular Ridge Area Triangularis Cerebellum Dorsal Lateral Thalamus Dorsal Medial Thalamus Geniculate Hypothalamus Lateral Forebrain Bundle Lentiform Nuclei Lateral Preoptic Area Medial Cortex Medial Preoptic Area Nucleus of Third Nerve Nucleus of Fourth Nerve Nuclei of the Dorsal Column Nucleus Profundus Mesencephali

NR NS NTOL OC OT PDVR PT RAPH RET ROT S ST TEC TEG TOL TS VM

Nucleus Ruber Nucleus Sphericus Nucleus of the Lateral Olfactory Tract Optic Chiasma Optic Tract Posterior Dorsal Ventricular Ridge Pretectal Nuclei Raphe Nuclei Reticular Nuclei Rotundus Septal Nuclei Striatum (Paleostriatum) Tectum Tegmentum Lateral Olfactory Tract Torus Semicircularis Nucleus Ventromedialis

m"

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6

FIG. 2. Location of electrode tips that elicited the following behaviors: O=assertion displays; A=circling; [3=rolling; and ©=curling.

identical to the normally seen "face off" posturing seen in this species [20,31] and 1 animal showed a pronounced extension of the dewlap. Furthermore five of the lizards superimposed the display action pattern (push up) on these postures resulting in a nearly complete challenge display. In no case, however, did the stimulated animal orient towards the conspecific in the cage. The challenge behavior was even seen when the test animal was alone in the cage. Interestingly, the conspecific responded to the elicited challenge behavior with the normal responses [31] of flight, submission or return challenge. Table 1 shows a summary of sites that produced challenge behavior. Not shown are the sites in the dorsal aspects of PDVR.

Elementary Locomotor Effects Figure 2 shows the location of tip sites giving elementary locomtor effects: circling, rolling and curling. Fourteen sites elicited a stimulus bound circling that began with stimulus onset and stopped immediately on stimulus offset. As cur-

rent intensity was increased the circling became more rapid. progressing finally to a tonic curling of the body in the direction of circling. 71% (10/14) of the sites produced ipsilateral circling and 29°~ produced contralateral circling. As seen in Fig. 2 all electrodes producing circling were posterior to the telencephalon, in the dorsal thalamus or brain stem reticular formation. Ten animals manifested tilting and rolling when stimulated. As with circling the response started immediately upon stimulus onset and ceased upon termination of the stimulus. At low current (less than 50 tzA) the animal would tilt his head and look straight up in the air. At higher currents the whole body would be rolled up onto one side. Finally (average of 129/zA) the animal would roll sideways across the cage floor. As shown in Fig. 2 the sites that produced rolling were found in a column of cells in the ventral medial brain stem. Seven animals manifested a curling response when stimulating, consistently curling into a ball with no bias to curl either to left or to the fight. The threshold for curling was 68

STRIATAL STIMULATION IN LIZARDS

619

FIG. 3. Location of telencephalic electrode tips. O=assertion display; &=challenge behavior; and I-]=arousal but no display. TABLE1

Behavior Elicited

Location Striatum (21 animals) Anterior 2h of ADVR (13 animals) Posterior Ih of ADVR (13 animals)

Display

Challenge

Exploration

13" 1

3 1

5 11t

4

5*

4

Test for Table I: Xz(4)=17.8, p<0.005. Test for each elicited behavior: *X2(2)= 10.5, p<0.01. t×2(2) = 13.2, p<0.005. *X2(2)= 11.9, p <0.005.

/xA. Again, the behavior was stimulus bound. As shown in Fig. 2 all the sites were located in the midline of the diencephalon and mesencephalon.

Generalized Arousal Some points yielded a nonspecific pattern of arousal, exploratory behavior and escape behavior. That is, stimulation of these sites did not elicit any specific locomotor response or display. At low current (usually less than 50 ~A) the animal would open his eyes and look around the cage. At higher current flow (between 50 and 100 t~A) the animal would move around the cage and lick the cage floor and objects in the cage. Often, as current levels went above 100 /~A these animals would manifest escape behavior such as running and jumping at the walls of the cage. The electrode sites producing this behavior were primarily located in the telencephalon. Figure 3 shows the distribution of these sites. Table 1 shows a summary of the telencephalic sites that gave this exploratory behavior. DISCUSSION

Histochemical analysis of reptile telencephalon has re-

vealed that the basal telencephalic structures are rich in cholinesterase and monoamines [4, 21, 22, 23, 25]. These structures have been shown to receive ascending catecholaminerglc fibers from the midbrain tegmentum [24]. On the basis of these histochemical findings and on the basis of known circuitry (such as appropriate thalamic connections) the basal cell masses of the reptile brain are now thought to be the homologue to the mammalian basal ganglia [4, 9, 21, 22, 25]. In the lizard these ventral cell groups are usually referred to as the paleostriatum (or striatum) and nucleus accumbens. This region also includes the nucleus of the lateral olfactory tract [21]. Close examination of the histochemical findings reveals that the greatest density of cholinesterase and catecholamine activity if found in the immediate area surrounding the tip of the lateral ventricle (see [4], p.442 and [21 ], p. 20). Stimulation of the caudal part of this region in the present study produced a consistent low threshold social display in 18/24 animals. Two areas of mammalian behavioral work provide some insight into this finding. First, bilateral lesions in the globus pallidus of the squirrel monkey abolishes the species typical social penile display [19]. The lesions needed are small and the loss only occurs when the globus pallidus or its outflow fibers are damaged. The implication is that selective stimulation of this area in the squirrel monkey could elicit this display. Secondly, amphetamine induced stereotyped behavior in rats has been clearly traced to a dopaminergic response of the striatum [13]. The stereotyped behavior elicited by amphetamine is not a complete species specific response such as a social display [13], however, iguanid lizard assertion displays are characterized by a repetition of a simple motor act (the push up movement) and in this respect is similar to the stereotypy reported in rats. It should be noted, however, that electrical stimulation of the striatum in rats fails to produce stereotyped behavior unless the stimulation is bilateral [34]. Previous neuroethological studies on reptiles provide a base for the present study. Bilateral lesions of the posterior telencephalon (nucleus sphericus, nucleus ventromedialis, PDVR and posterior regions of ADVR) in Sceloporus occidentalis abolished all challenge activity [31]. Stimulation of this area is now seen to elicit challenge behavior. The tight clustering of points in a small region just anterior and dorsal to nucleus sphericus (Fig. 3) suggests that this region is the critical locus in the posterior telencephalon for the integration of challenge display behavior. The earlier reported abolishment of challenge behavior following striatal lesions in both Anolis and Sceloporus [12,32], however, indicates that anterior structures play some role in the production of the challenge display. It is possible that connections between the striatum and more caudal structures play a role in the performance of challenge behavior. The absence of dewlap extension following stimulation of the nucleus profundus mesencephali (NPM) is difficult to reconcile with the conclusion [29,30] that in the collared lizard the NPM is the principle integrator of dewlap extension. This may represent a species difference. The absence of elementary locomotor responses from telencephalic sites is in accord with the bulk of previous studies on reptiles [26] but is at variance with work done on mammals showing turning and circling following electrical stimulation of the basal ganglia [34,35]. The present results reinforce the suggestion [21,26] that the reptile telencephalon is primarily concerned with the integration of complex behavior. The present results demonstrate a degree of localization

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of function that was not manifest in previous reptile stimulation studies [6, 7, 26, 30]. As mentioned earlier, the failure to use precise electrode marking methods [7,26] and to test for complex social behavior [26,30] are possible factors contributing to the noticeable lack of structure-function correlates in earlier studies. Current spread when using monopolar electrodes in a small animal is seen to not be as serious a problem as suggested by the earlier studies. This is in accord with work using quantitative electrophysiological [3] and behavioral [33] methods to determine the degree of spread. Both these studies revealed an effective spread of less than 0.5 mm at 100 /zA. In Sceloporus occidentalis a 0.5 mm

radius drawn from the tip of the ventricle (at 50cybthrough the telencephalon) forms a circle that intersects the septal nuclei, the ventral portions of A D V R the lateral forebrain bundle and the nucleus ventromedialis. Thus the levels of current employed in the present study resulted in the relatively clear functional localization summarized in Fig. 3 and in Table 1. Finally, the present results confirm that reptiles share a basic vertebrate neural organization and suggests that iguanid lizards may provide a simple model system for the study of stereotyped display behavior and striatal function.

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