Lateralized rewarding brain stimulation affects forepaw preference in rats

Physiology&Behavior,Vol. 34, pp. 495--499.Copyright©PergamonPress Ltd., 1985. Printedin the U.S.A.

0031-9384/85$3.00 + .00

Lateralized Rewarding Brain Stimulation Affects Forepaw Preference in Rats N . H E R N A N D E Z - M E S A 1 A N D J. B U R E ~ 2

Institute o f Physiology, Czechoslovak Academy o f Sciences, Prague R e c e i v e d 16 J a n u a r y 1984 HERNANDEZ-MESA, N. AND J. BUREt. Lateralized rewarding brain stimulation affects forepaw preference in rats. PHYSIOL BEHAV 34(4) 495--499, 1985. Rats trained to reach for food pellets into a narrow tubular feeder consistently prefer to perform this stereotype instrumental movement with either the left or the right forepaw. In 16 rats with established handedness electrodes were implanted into both lateral hypothalami. The animals were rewarded by intracraniai self stimulation (ICSS, 300 msec, 50 Hz, 20-60/zA) for reaching into a modified feeder for a plastic ball operandum, the movement of which between the bottom and entrance of the feeder was monitored by mechanical contacts. The rats readily continued to reach when ICSS was delivered immediately after the photoelectrically detected reach or after the displacement of the operandum. Most rats learned in a single session to modify the movement when ICSS delivery was made contingent upon holding the operandum between the bottom and entrance of the feeder for 256 or 512 msec. The efficiency of reaching (ratio of successful reaches to all reaches) decreased with increasing holding time; only a few animals were able to master a 1024 msec delay. Reaching was supported by ICSS of either lateral hypothalamus. Whereas in 8 rats the strongly expressed forepaw preference was not changed by lateralized ICSS, in 8 latently ambidextrous animals stimulation of the lateral hyimtbalamus ipsilateral to the preferred forepaw increased reaching with the normally non-preferred forepaw from 15% to 60%. Stimulation of the lateral hypothalamus contralateral to the preferred forepaw did not change the preference. The preference shift was equally well expressed in simple and difficult versions of the task. It is concluded that lateralization of motivational influences can be reflected in the asymmetry of the neural mechanisms processing the lateralized sensory signals and/or elaborating the lateralized motor output. Handedness Self stimulation Lateral hypothalamus Movement control Skilled movements Motivation

R E C E N T research suggests that mesollmbic and nigrostriatal systems play a pivotal role in incentive motivational learning [21] manifested by the changed ability of reinforcement-related environmental stimuli to elicit operant responses. A fruitful approach to the analysis of the underlying brain mechanisms employs lateralized elimination or stimulation of the corresponding neural circuits. Unilateral damage of lateral hypothalamus [12] or substantia nigra [11] leads to neglect of the contralateral side of the body which is not due to impaired perception but rather to deficient interfacing of the memory input with the motor output. Selfstimulation with electrodes in the medial forebrain bundle is decreased more by lesions of the ipsilateral than of the contralateral nigrostriatal system [3, 10, 15]. Combination of unilateral lesion or stimulation of the reward circuits with unilateral functional decortication showed that the hemicortex ipsilateral to the intact or stimulated reward circuit is more important for operant control of feeding and drinking [1], thermoregulatory behavior [17,18] and brain stimulation reward [16] than the hemicortex overlying the damaged (nonstimulated) side of the brain. Behavioral asymmetry elicited by two lateralized lesions or by lateralized lesion and lateralized stimulation may sim-

Lateralization of brain functions Rat

ply be due to impaired access of the reward circuit to the input or output mechanisms. Lateralization of this kind is trivial. Evidence for the integrative role of reinforcement in incentive motivational learning should be based on observations demonstrating that the shift of the rewarding stimulus from one side to the other side of an intact brain is reflected by a corresponding lateralization of behavioral output. Lateralized operant reaching, the so called handedness [4, 8, 14], is a reaction particularly well suited for the experimental study of this problem. It is conceivable that the preference of the rat for the left or right forepaw is partly due to an asymmetry of motivational influences. If the animal's reaching into a narrow tube is not rewarded by food pellets but by short trains of intracranial self stimulation (ICSS), it is possible to lateralize the reward by applying the electrical stimulus to one side of the brain. The purpose of the present paper is to examine the influence of the shift in ICSS lateralization on the selection of the forepaw used for operant reaching. METHOD Sixteen male hooded rats (Druckray strain) aged 3 months were used throughout. Under pentobarbital anaesthesia (40

~Visiting scientist from ICB Victoria de Giron, University of Havana, Cuba. 2Requests for reprints should be addressed to Dr. Jan Buret, Institute of Physiology, Czechoslovak Academy of Sciences, Videtisk6 1083, 142 20 Prague 4 - Kr6.

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mg/kg) the animals were bilaterally implanted with two arrays of 4 stainless steel wire electrodes (100 tzm diameter) arranged in the coronal plane to form a rhomb with 1.0 mm sides. The electrode assemblies were directed to the medial forebrain bundle at the level of lateral hypothalamus according to the atlas by Fitkov~i and MarSala [5]. They were used in order to increase the probability of obtaining comparable ICSS from either hemisphere. The coordinates for the deepest electrode were 2.5 mm posterior to bregma, 1.4 mm lateral from midline and 9.0 mm below skull surface, with lambda 1.0 mm below bregma. Other two electrodes were about 0.7 mm higher and 0.7 mm more lateral or medial, respectively. The fourth electrode was about 1.4 mm above the f'trst one. The electrodes were connected to miniature transistor sockets which were secured to the skull bones with anchoring bolts and acrylate. Two days after surgery, the animals were put on a 24 hr food deprivation schedule, gradually reduced to 85% of free feeding weight and trained to reach for food pellets into a horizontal tubular feeder (11 mm internal diameter). The training was continued until the animals were able to get pellets placed 2 cm from the feeder orifice. During the same period, the implanted electrodes were tested for self stimulation in another testing box equipped with a nosing hole guarded by a photoelectric sensor. The animal was connected with a counterbalanced flexible cable to the output of a constant current stimulator which delivered a train of 300 msec, 50 Hz sine waves to the selected electrodes whenever the animal activated the nosing hole sensor, A CRO was used to monitor the intensity of the stimulating current as the voltage drop across a precision resistor connected in series with the stimulating electrodes. All rats learned to use the nosing hole operandum in a single session, lasting for 1-2 hr. The threshold was established for each of the 6 electrode pairs (1-2, 1-3, 1-4, 2-3, 2-4, 3-4) on each side of the brain and the electrode pairs yielding the lowest threshold were used in the further course of the experiment. The actual experiment started only after the animals had fully mastered the reaching-for-food reaction and after their self stimulation sites had been reliably established. The rat was placed into the experimental box, the tubular feeder of which was equipped with a photoelectric sensor monitoring

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FIG. 2. Average rates (ordinate) of reaching (white columns) and of ICSS-reinforced reaching (shaded columns) during schedule a (Ph) and during 30 min exposures to schedule c, with the delay set to 256, 512 and 1024 msec, respectively. The vertical bars denote SEM values in this and subsequent figures.

the movement of the forepaw across a point 5--6 mm inside to feeder. The food pellet at the bottom of the feeder could be replaced with a plastic ball (4 mm in diameter) connected to a spring mounted needle (Fig. 1). The animal could grasp the plastic pellet and pull it to the feeder entrance. As soon as the pellet was released, it was returned by the spring to the resting position at the bottom of the feeder. Movement of the operandum between these two extreme positions was monitored with two sets of mechanical on/off contacts, which were connected together with the output of the photoelectric sensor to solid state programming equipment, lntracranial self stimulation (300 msec, 50 Hz, AC) was delivered whenever one of the following conditions was satisfied: (a) immediately after activation of the photoelectric sensor; (b) immediately after the operandum was displaced from the resting position; (c) after the operandum was held between the resting position and feeder entrance for a preset interval (256-t024 msec). The delivery of ICSS under the above reinforcement contingencies was recorded with a cumulative recorder (100 responses per maximum deflection). All reaches detected with the photoelectric sensor during a period of stimulation were simultaneously counted with an electronic counter. Reaches performed with left or fight forepaw were separately encoded by a trained observer and the preference was expressed by the ratio of the left-right counts during the analyzed epoch. After conclusion of the experiments the animals were deeply anesthetised with pentobarbital and perfused with physiological saline followed by 10% formalin. Anatomical control showed that the electrodes were localized in and around the medial forebrain bundle at the level of lateral hypothalamus. RESULTS Out of the 16 rats six preferred to reach for food with left and 7 with the fight forepaw. Two animals used both

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FIG. 3. Cumulative recordings of reaching for food pellets (F) and of ICSS delivery upon activation of the photoelectric sensor (Phcondition a) or when the pellet was held between the resting position and feeder entrance for more than 256, 512 or 1024 msec (condition c). Above: successful learning at the maximum delay. Below: failure to master the 1024 msec delay. Calibration: 100 events, 10 min. The numbers below the recordings denote the number of all photoelectrically recorded reaches during the corresponding interval.

forepaws alternately with a preference for the right paw. One rat failed to acquire the reaching for food habit but learned later to reach for the operandum with the left forepaw when ICSS was used as reinforcement. The nosing hole test showed that all animals displayed well expressed ICSS which attained the rate of up to 2 Hz with stimulus intensity about 20% above threshold (42---5/~A). In 10 animals, self stimulation could be elicited from either hemisphere, in 6 rats only from one hemisphere. Transfer of reaching for food to reaching reinforced by ICSS was immediate, when ICSS was delivered after activation of the photoelectric sensor (condition a). The average reaching rate was 1.13-+0.26 Hz (n= 16). The reaching rate remained the ~ame when displacement of the plastic operandum was used to trigger the electrical stimulation (condition b), but the reinforcement delivery rate dropped by 0.39-+0.12 Hz, because not all photoelectrically detected reaches were successful. In the main experiment all rats were given two sessions in which 5 min reaching for food and 10 min of photoelectrically triggered ICSS was followed by three 30-min periods of ICSS delivered when the rat succeeded to hold the pellet between the resting position and the feeder entrance for more than 256, 512 or 1024 msec. ICSS ipsilateral and contralateral to the preferred forepaw was used in the first and second sessions, respectively, in half of the animals and the sequence was reversed iri the remaining rats. The overall results are shown in Fig. 2. The task proved to be considerably more difficult than the simple displacement of the operandum. Whereas the photoelectrically

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FIG. 4. Forepaw preference in the group of 8 latently ambidextrous rats during reaching rei~orced by ICSS contralateral (ICSS C) or ipsilateral (ICSS I) to the forepaw preferred when reaching for food (FOOD). P and N: preferred and nonpreferred forepaw.

triggered ICSS rate remained stable at the 1.07_+0.1 Hz (n--16), already the introduction of the 256 msec holding delay decreased ICSS rate by 75% although the reaching rate dropped only by 10%. Most animals continued to reach and were eventually able to master the 512 msec delay but only 7 rats showed some improvement with 1024 msec delay. Since many rats stopped reaching altogether the average reaching rate dropped to 0.41 Hz in this case. Typical cumulative recordings are shown in Fig. 3. Eight animals displayed throughout the experiments 100% forepaw preference which was not changed when food was replaced by stimulation of the ipsilateral or contralateral hypothalamus and when ICSS was delivered at various delays. The forepaw preference in the remaining 8 rats was less rigid and was significantly changed with the change of the stimulus side. As shown in Fig. 4, the average forepaw preference established in these 8 animals when they were reaching for food remained unchanged when reaching was reinforced by ICSS of the hemisphere contralateral to the preferred forepaw, but was reversed when ipsilateral ICSS was used as reward (t(7)=3.7, p<0.01, paired t-test). The effect was independent on the order of testing (ipsilateral ICSS preceding the contralateral ICSS or vice versa). As shown in Fig. 5 the change was equally well expressed with the photoelectrically triggered ICSS and with various holding times. A two-way analysis of variance of the data of Fig. 5 revealed significant effect of the stimulation side, F(1,49)=14.12, p<0.01, but no difference between reinforcement schedules, F(3,49)=0.1 l, n.s. Significant side × schedule interaction, F(3,49=5.91, p<0.01, was mainly due

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FIG. 5. The change of forepaw preference induced in the group of 8 ambidextrous rats during reaching reinforced by ICSS ipsilateral (I) or contralateral (C) to the limb preferred during reaching for food. Ordinate: average change±SEM from the preference observed during food-reinforced reaching (interrupted horizontal line). Abscissa: reinforcement contingencies.

to results obtained with the 1024 delay, when low rate of reinforcement probably decreased the ICSS effect. DISCUSSION

The main result of the present paper is the finding that lateralized ICSS facilitates the use of the contralateral forepaw in a lateralized operant task. The effect was relatively weak because it could not be demonstrated in rats with clearly expressed handedness but was strong enough to elicit consistent changes of forepaw preference in ambidextrous and latently ambidextrous amimals. It must be emphasized that reaching is a highly stereotype movement [14] consisting of a rapid extension (60 reset) followed by grasping and slow retraction of the forelimb (200--400 msec). Whereas the first component has the character of a ballistic movement, preprogrammed by cerebellum [9] and little influenced by tactile and proprioceptive feedback, retraction is a more variable phase which can be more easily affected by conditioning. It was expected that lateralization of reward may differentially influence performance of an established response and its modification by changed movement-reinforcement contingency. This assumption was not confirmed. The preference shift was equally well expressed with the overtrained reactions (simple reaching) and with the acutely modified reactions (prolonged holding of the operandum). In the latter case the preference ratios obtained for all reaches were equally distributed between non-successful and successful movements. Since the ICSS is applied after completion of the reach, the changed forepaw preference cannot be due to stimulation-induced interference with the elaboration of the movement but rather reflects prolonged facilitation of cortical and subcortical structures ipsilateral to the stimulated reward site. Increased excitability can amplify the relevant sensory stimuli and improve their access to output mechanisms or can directly influence the motor centers.

Important evidence supporting the former possibility was obtained in experiments with hypothalamic stimulation eliciting predatory attack of the cat on a rat [6]. In absence of the rat, hypothalamic stimulation does not elicit an attack. Touch around the mouth, ineffective without stimulation, evokes in a restrained cat head turning toward the stimulus and, when the lips contact the stimulus, mouth opening and biting [12]. The response can only be evoked from the side contralateral to the hypothalamic stimulation and the extent of the sensory field is directly related to the intensity of the central stimulus. As pointed out by Teitelbaum [21] presence or absence of motivational influence expands or shrinks the peripheral sensory fields and facilitates or blocks the reflexive components of the behavioral pattern, respectively. More relevant in the present context are stimulus-induced excitability changes reported by Zimmerberg and Glick [22], who used lateralized stimulation of the caudate nucleus to change side preference in rats bar-pressing for water reinforcement in a two-lever operant chamber. While continuous subthreshold stimulation of the caudate nucleus contralateral to the preferred side did not change the rat's preference, stimulation of the ipsilateral caudate nucleus induced the rat to switch the levers for the duration of the stimulus. It is conceivable that the ICSS-induced switching of handedness is accomplished not only at the cortical level but also at the level of candate nucleus [4,8] which participates in the elaboration of reaching together with the cerebellum [9]. The limb preference has often been ascribed to cortical asymmetries, e.g., to unequal number of pyramidal tract axons [2]. The present experiments indicate that forepaw preference can be due to imbalance of subcortical mechanisms. Glick et al. [7] demonstrated asymmetry of dopamine content of the two striata in normal rats and reported that the animals display in the T-maze or in two-lever operant tasks preference toward the striatum with lower dopamine level. Siegfried and Bure~ [19] reported that rats with 6-OHDA lesions of right substantia nigra displayed, when tested for handedness 35 days later, a 100% preference for the right forepaw. In rats with preoperatively established handedness reaching was uneffected by 6-OHDA lesion of the ipsilateral but was severely impaired or reversed by similar lesion of the contralateral substantia nigra. The unilateral dopaminergic denervation of caudate nucleus did not cause paralysis of the contralateral paw which was used normally for walking, food holding and grooming, but considerably decreased the animal's capability to employ this limb in discrete instrumental reactions. The experimental evidence obtained in both lesion and stimulation studies points toward the conclusion that motor asymmetries can be a consequence of asymmetries of motivational and attentional mechanisms. Reward does not affect the sensory and motor functions directly but rather influences their integration into elements of operant responses. The links between the reward systems and the related behavioral mechanisms are preferentially established within the same hemisphere. This organizational principle implies that asymmetry of incentive motivational learning may account for lateralization of higher nervous functions in animals and can model some aspects of hemispheric specialization of the human brain.

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