Two-way avoidance learning in pigeons after olfactory nerve section

Physiology and Behavior, Vol. 13, pp. 57-62. Brain Research Publications Inc., 1974. Printed in the U.S.A.

Two-Way Avoidance Learning in Pigeons After Olfactory Nerve Section ROBERT S. HUTTON 2 , BERNICE M. WENZEL, THEODORE BAKER AND MARTHA HOMUTH

Department o f Physiology and Brain Research Institute, University o f California, Los Angeles

(Received 8 February 1974) HUTTON, R. S., B. M. WENZEL, T. BAKER AND M. HOMUTH. Two-way avoidance learning in pigeons after olfactory nerve section. PHYSIOL. BEHAV. 13(1) 57-62, 1974. - In two experiments, pigeons with bilaterally sectioned olfactory nerves reached criterion performance in two-way shock avoidance sooner than sham-operated and unoperated control birds. The CS was red light added to a constant hackgzound of green houselight, the US was footshock, and the avoidance response was moving from one compartment to the other through an intervening V-shaped opening. In the first experiment, intertrial responses delayed CS onset; in the second experiment, the ITI was fixed and intertrial responses had no" effect. This contingency made no difference in the results. Differences in rate of extinction were not significant but the lesioned groups showed a consistent tendency toward a slower rate. Heart rate was recorded from each bird while partially restrained during simulated trials in which the CS was presented but not the US. These data suggested that the experimental birds were less likely to habituate to the CS. A similar effect has been found in other experiments with both rats and pigeons that had sustained olfactory lesions. Olfactory nerve section

Two-way shock avoidance

Pigeon

Non-olfactory functions of olfactory system

EXPERIMENT 1

IN PIGEONS, olfactory bulb ablations or bilateral olfactory nerve sections produce changes in orientation and in behavioral reactivity to non-olfactory stimuli [ 17,18]. A n u m b e r of experimenters working with rats have reported similar observations [5, 6, 7, 9, 10, 12]. Olfactory lesions also affect avoidance learning by rats, facilitating two-way active avoidance [2, 8, 9, 10, 11, 14, 15] and impairing passive avoidance learning [1, 8, 9, 10, 11, 14]. Other behavioral effects, seemingly unrelated to olfactory stimulation, have been described after olfactory lesions in mice, hamsters, and rats. This literature has recently been reviewed [ 16]. The experiments reported were conducted to study the effect of an olfactory lesion on two-way avoidance learning by pigeons. In earlier experiments with pigeons, no differences had been found in the behavioral effects of bilateral olfactory lesions produced by cutting the olfactory nerves or by removing the olfactory bulbs. Because nerve section is less traumatic to the central nervous system than is bulb ablation, all lesions in these experiments were bilateral nerve sections.

METHOD

A nimals Seventeen naive adult homing pigeons from our own breeding colony were individually housed in a windowless room. They were given food and water ad lib throughout the experiment. Lighting was controlled automatically with lights on from 2300 to 1100 hr.

Apparatus A Grason-Stadler pigeon enclosure was modified into a two-way shuttle box, 45 cm high x 27.5 cm wide x 70 cm long, containing overhead lighting (CS and house lights) and 2 grid platforms through which scrambled electric shock (0.50 mA) could be administered to the birds feet (see Fig. 1). The platforms were separated by an opaque cardboard partition. A 74 ° V-shaped center opening extended from 10 cm above platform level to the ceiling.

l Supported in part by grant MH 6415 from the National Institute of Mental Health to the Brain Research Institute, under which Dr. Hutton received a postdoctoral traineeship. Technical assistance was provided by Irene Jones and Harold McCaffery. Experiment 1 was reported at the Winter Conference on Brain Research, Snowmass-at-Aspen, Colorado, January 16-22, 1971. Requests for reprints should be addressed to Dr. Bernice M. Wenzel at the University of California, School of Medicine, Department of Physiology, Los Angeles, California 90024. a Now at the School of Physical and Health Education and Department of Psychology, University of Washington, Seattle, WA 98195. 57

HUTTON, WENZEL, BAKER AND HOMUTH

FIG.

1. Diagram of shuttle box.

Each platform was hinged at the back and was supported by two spring coils under the front portion. Weight of the bird was sufficient to depress either platform and to activate an underlying microswitch, which was connected to programming equipment. Both houselights and stimulus lights were 7 W frosted bulbs. House lighting was provided by one green bulb in each compartment. Two red bulbs in each compartment were used for the CS. The shuttle box was housed in a well-ventilated sound-shielded enclosure. Surgical Procedure Anesthesia was induced by placing the pigeon in a plastic bag containing a small gauze pad dipped in Metofane (Methoxyflurane; Pitman-Moore) and was then maintained by a similar pad taped lightly over the nostrils. Additional Metofane could be dropped onto the pad as needed. Induction typically occurred within l-2 min. Most pigeons were given a supplemental dose of Equithesin (0.1 cc i.m.) before starting surgery. Following Xylocaine injection under the. scalp, a mid-sagittal longitudinal incision was made to expose the cranium. Skin folds were held back by clamps and the cancellous bone anterior to the forebrain was removed by drilling. The olfactory nerves could be seen running through a thin bony sheath between the orbits just anterior to the olfactory bulbs. For the 5 sham-operated animals, surgery ended here and the wound was closed with sutures. For the 6 experimental animals, olfactory nerves were bilaterally sectioned under direct vision and a short segment was removed to prevent possible regeneration. Hemostasis was induced by Gelfoam. No anesthetization or surgery was performed on the 6 members of the unoperated control group. Procedure

Eleven days after surgery, each bird was given a 30-min

exploration session in the avoidance box with no stimulus lights or shocks presented but with houselights on after the first minute. Active avoidance training with a modified FI schedule began the next day. All pigeons were trained to avoid shock (US) by moving through the opening in the partition to the opposite side of the shuttle box within 10 set after onset of the red lights (CS) in the compartment where the bird was located at the time. The green houselight in each compartment remained on throughout the session. The minimum IT1 was 30 set and each anticipatory response, i.e., movement to the other compartment in the absence of CS, increased the ITI by 30 sec. Thirty-nun training sessions were held daily until 90% avoidance responding or better had been achieved in any five sessions or until 10 sessions had been completed. Extinction sessions, in which shock was omitted, began on the day after the last training session and continued for a maximum of 10 sessions or until no better than 10% avoidance responding was reached in three consecutive 30-min sessions. One week later all birds were retrained for a maximum of 5 sessions or until each pigeon’s best avoidance performance during initial training was evidenced in a single session. The experiment was conducted in two replications with all groups represented in both. Training sessions were held at the same time each day during the dark phase of the vivarium light cycle, a contingency that had been found to facilitate avoidance learning in preliminary experimentation with normal pigeons. The birds were tested in the same order in each session, with each group represented in each successive set of 3 birds. Heart rate (HR) was recorded during 3 simulated trials in 2 sessions, one on the day before, and the other on the day after, retraining. For these trials, the pigeon was restrained in a portable holder to which it had been adapted in its home cage on the preceding day. During the sessions, the holder was placed at one end of the shuttle box. Following

O L F A C T O R Y LESION AND AVOIDANCE L E A R N I N G IN PIGEONS a 5-min period in the dark, the houselights came on for 5 rain and then 3 avoidance trials were simulated by presenting the CS for 5 rain with 5-rain ITls o f houselights only. The HR was recorded on a Grass polygraph from insect pin electrodes inserted under the skin of one thigh and the ipsilateral breast. A ground electrode was inserted into the opposite leg. Heart rate was determined for each pigeon by counting the number of beats during the first and last minutes of the initial dark period, of the three periods with houselights only, and of the three periods with the CS.

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RESULTS Group performance curves of percent avoidances for the first 7 training sessions are shown in Fig. 2. After Session 7, the number of pigeons in each group decreased as some met the criterion and the group percentages for those remaining are spuriously low as a result. In all groups, avoidance performance improved in the early sessions. The control groups did not maintain steady improvement, however, and differed significantly from the nerve-sectioned group in Sessions 6 (Mann-Whitney U = 8, p < 0 . 0 1 ) and 7 (U = I0, p<0.05). The sham-operated control group showed the ]east improvement. When compared separately with the nerve-sectioned group, the differences were significant for Sessions 4, 5, 6, and 7 (U = 5.5, 4.5, 3, and 4, p = 0.05, 0.03, 0.02, and 0.03, respectively). The control groups did not differ from each other at any point (p>0.05). All groups showed similar performance in extinction in spite of the differing levels of acquisition. One or 2 pigeons in each group failed to meet the criterion, 2 or 3 in each group met it in the minimum number of sessions, and the remainder t o o k from 5 to 6 sessions. All experimental pigeons were retrained to criterion in 2 sessions, whereas 1 pigeon in the control and sham groups did not reach criterion in 5 sessions. Analysis of the HR data revealed no significant group differences ( p > 0 . 0 5 ) but suggested a possible effect that might have been related to the superior avoidance performance of the nerve-sectioned birds. After retraining, the lesioned pigeons tended to show a larger increase in HR when the houselights came on after 5 min in the dark test box and then to show a consistently smaller drop in HR, as compared with that during the first min in the dark box, each time the CS was presented. ]n other words, the experimental group's HR adapted less than that o f the control groups. The groups did not differ significantly in median HR during the first min in the test box (Nerve-sectioned = 234 bpm, Sham = 244, Unoperated = 278). In the same type of HR recording session after extinction and before retraining, no group differences of any kind were seen. DISCUSSION The results support previous observations that pigeons with olfactory lesions exhibit changes in behavior that appear unrelated to loss of olfactory cues [17,18]. They agree with other data showing improved two-way shock avoidance after bulbectomy in rats [8, 9, 10, 14]. Our pigeons could not be observed directly during training and so it is not possible to say whether the experimental birds exhibited unusual behavior related to CS or US onset that might have facilitated avoidance responses. There were no group differences in the number of intertrial responses in

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SESSIONS FIG. 2. Group performance curves for first 7 avoidance training sessions in Experiment 1. spite of the fact that each o f these responses delayed CS onset by 30 sec. This feature of the procedure, therefore, could not have accounted for the superiority of the lesioned birds in meeting the learning criterion. They were neither more nor less spontaneously active under these circumstances. The suggestion in the HR data that the experimental birds were less habituated to the CS could account for the difference in avoidance performance. It is consistent with earlier data [17] that showed progressively larger HR increases to repeated visual stimuli in successive habituation sessions for pigeons with sectioned olfactory nerves or ablated olfactory bulbs. Similar results have been obtained for a u d i t o r y stimuli with rats after olfactory bulbectomy [6] and after olfactory nerve section [7]. A second experiment was designed to study avoidance learning when the reinforcement contingencies were independent of the bird's behavior, i.e., with a fixed ITI and a constant number of CS presentations. In addition, HR w a s monitored before any training was given in order to determine the initial reaction of each group to the lights that would be used as CS. EXPERIMENT 2 METHOD

A nimals Twenty-five naive adult homing pigeons from our own breeding colony were maintained as described above.

60

HUTTON, WENZEL, BAKER AND HOMUTH

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Apparatua and Surgical Procedure The apparatus and surgical procedures were the same as in Experiment 1. Bilateral olfactory nerve sections were performed on 9 pigeons and sham operations on 8, leaving a non-surgicai control group of 8 birds.

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Procedure The training schedule consisted of a maximum of 10 daily training sessions of 30 trials each. The CS-US interval was 10 sec as before and the ITI was 30 sec. Intertrial responses had no effect. The learning criterion was 90% avoidance or better in any three sessions. Extinction training began 3 days after the l Oth training session and continued to a criterion of three consecutive sessions with no more than 10% avoidance. Training began on the twelfth day after surgery with a 30-rain exploration session. On the following day, each bird was adapted, for 30 rain in its home cage, to the restraint device for HR recording. On the next day, HR was recorded in the test box using the same procedure described in Experiment I. Avoidance training began on the following day. A post-training HR session was held on the second day after the 10th training session. On the preceding day, each bird had a 30-min session of restraint and adaptation in its home cage. Extinction sessions, in which shock was withheld, began on the day after the HR session.

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RESULTS Group mean percent avoidances for Sessions I through 7 are shown in Fig. 3. On the basis of the results obtained in Experiment I, it was predicted that the nerve-sectioned group would make more avoidance responses than the control groups. The Mann-Whitney U-test on the data for Sessions 1 through 6 showed that the experimental group was significantly better than the combined control groups i n Sessions5 and 6 ( U = I 2 and 13.5, respectively; p<0.001, one-tailed). By Session 7, 4 birds had met criterion, viz., I nerve-sectioned, I sham-operated and 2 unoperated. The remaining eight nerve-sectioned pigeons continued to perform significantly better than the 13 remaining control pigeons (U = 15, p<0.01 ). In Session 8, 2 more experimental birds reached criterion compared to 1 additional sham-operated bird and no unoperated ones. By Session I0, 2 experimental and 8 control birds had not reached criterion. With the dwindling numbers of birds, these differences in percent avoidance became insignificant, however. In the last session l bird in each of the 3 groups was still performing at less than 20%. The results for the In'st 3 sessions of extinction are given in Table I. The criterion of I0% avoidance or less for 3 consecutive sessions was met by 69% of the control pigeons in the first three sessions as compared to 33% of experimental birds (p>0.05). A difference in this direction would be expected on the basis of the final acquisition perform-

TABLE 1 PERCENT AVOIDANCE RESPONSES BY MEMBERS OF EACH GROUP DURING THE FIRST THREE EXTINCTION SESSIONS

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Median differences from the HR during the first min in the dark box are shown in Fig. 4 for the onset and offset of each CS and each houselight during the pretraining session ( H R c s or HRHL - HRDark). These differences are plotted as percentages because the initial HR for the nervesectioned group before training (176 bpm) differed significantly (U = 20.5, p
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O L F A C T O R Y LESION AND AVOIDANCE L E A R N I N G IN PIGEONS

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HLI c'sl H'LzCSz HL3 C'S3 HL4 CSl HLZ CS2 Hi'3 CS3 STIMULUS FIG. 4. Median % difference in HR between 1st rain in unlighted shuttle box and 1st rain of each 5-min period of houselighting (HL) and of CS presentation before and after training in Experiment 2. control groups (Sham = 240, Unoperated = 218, Mean = 229). There was no such difference in initial HR after t r a i n i n g (Nerve-sectioned = 246 bpm, Sham ffi 259, Unoperated ffi 315). The same trend observed in Experiment 1 can be seen here, viz., less habituation o f HR in the experimental birds than in the controls. Before training, the lesioned birds were somewhat more reactive, relative to their initial HR, to the third presentation of the CS than to the second. At th{s point, they differed significantly from the combined control groups (U ffi 25, p < 0 . 0 5 , one-tailed). It is unlikely that the smaller responses o f the control groups before training reflect merely their hisher initial HR. At the start o f the posttraining recording session their HRs were even higher, indicating that they had not reached a s y m p t o t e in the f'Lrst session. DISCUSSION Both experiments have shown faster learning of two-way shock avoidance by pigeons with sectioned olfactory nerves than by control pigeons. The consistency o f this finding for both rats and pigeons strongly supports the hypothesis that removal of the normal olfactory input to other areas of the forebrain significantly alters behavioral function. Nothing is known about the role o f central neural structures in mediating avoidance learning by the pigeon. Various Limbic structures have been implicated in the rat, however, and neural pathways have been demonstrated between them and the olfactory bulb. It remains to be seen how the olfactory system exerts its influence and what differences there may be among species. The possible role of orientation and habituation in learning active avoidance appears to be worth exploring. Marks e t al. [5] reported poorer postural orientation of

bulbectomized rats toward the door between compartments in one-way avoidance but greater reactivity of the same rats to the sound of the food dispenser while learning a leverpressing task. Pigeons with olfactory lesions showed both o f these responses during magazine training for a key-pecking task, i.e., b_igh reactivity to the sound of the food hopper followed b y turning away from it [ 17 ]. In testing Startle responses of bu]bectomized rats [9,10], it was noted that their intertrial activity failed to increase in the second session as much as that of controls. This difference was interpreted as indicative of less habituation on the part of the experimental rats. The fai/ure o f HR to habituate in olfactory lesioned rats has been mentioned above [6,7]. No other workers have reported HR after olfactory lesions. Although the present data on HR provide only suggestive evidence, they are consistent with an interpretation of altered orienting behavior in the animals with olfactory lesions. In these experiments any tendency toward less habituation and greater alertness would have increased the impact o f the CS and US, thus facilitating acquisition of the avoidance response-if the increased subjective intensities were within the optimal range. It would also account for a greater resistance to extinction. In further work along these lines, it would be desirable to record HR in the actual training procedure and to make more detailed analysis in order to evaluate the role of orientation level more directly. The different training procedures in the two experiments did not produce strikingJy different results and the experiments were not designed to test this question. The important point is that the effect of the lesion was clear in both procedures regardless of the variation in ITIs, in numbers of trials per session, and in the effects of intertrial r e s p o n s e s . Performance of the nerve-sectioned group improved consistently in both experiments and reached

62

HUTTON, WENZEL, BAKER AND HOMUTH

much higher levels than that of the control birds in the later

sessions. Until recently, active avoidance learning was considered virtually impossible for the pigeon. Acquisition and extinction of one-way shock avoidance were described in 1968 [4]. The 3 out of 4 pigeons that met criterion in that experiment were then successfully trained in two-way avoidance. A later report [13], not available at the time the first of the present experiments was being designed, showed that free-operant shock avoidance behavior could be established in pigeons using depression of a foot-treadle as the response. Our o w n procedure was perfected afterprotracted preliminary work with a variety of alternatives.Each of the successful demonstrations is based on the principle of allowing a natural component of escape behavior to serve as the avoidance response. In our case, the base of the Vshaped opening in the partition between compartments was far enough above the floor grid that the bird was forced to flutter through it. Its shape and thinness prevented perching. T w o other novel features of the procedure should be noted. One was the successful use of footshock rather than the customary method of delivering shock through electrodes implanted under the pubis bones. It could be argued that the group differences in avoidance behavior were

dependent on a correlated difference in sensitivity to footshock, and that this latter difference was somehow related to the olfactory lesion. No deliberate determinations of shock thresholds were made because in preliminary work all birds trained in this apparatus appeared to react to the shock with similar latencies. That our data agree with those obtained from rats and that no difference in shock threshold was found in rats after an olfactory lesion [12,14] are inconsistent with an interpretation of altered footshock sensitivity. The final new feature was the scheduling of all sessions during the dark phase of the diurnal cycle when the pigeons were normally inactive. We have no explanation for the improved performance exhibited by intact birds when this procedure was instituted during pilot work with the training paradigm. It is analogous to the procedure, presumably followed in most experiments with rats, of training them during the light phase when they tend to be inactive. Further research on this question, using different species, would be of interest. There is no reason to postulate any relation between this aspect of the training and the effect of nerve section. Indeed, the same facilitation of active avoidance by bulbectomy in rats was reported whether training was done in the light [ 14 ] or dark [ 10] phase.

REFERENCES 1. Brown, (3. E., E. Harrell and N. R. Remley. Passive avoidance in septal and anosmic rats using quinine as the aversive stimulus.Physiol. Behm,. 6: 543-546, 1971. 2. Brown, (3. E. and N. R. Remiey. The effects of septal and olfactory bulb lesions on stimulus reactivity. Physiol. Behav. 6: 497-501, 1971. 3. Bugbee, N. M. and B. S. Eichelman, Jr. Sensory alterations and af~ve behavior in the rat. Physiol. Behav. 8: 981-985, 1972. 4. Macph~ E. M. Avoidance responding in pigeons. J. exp. Analysis Behav. 11: 629-632, 1968. 5. Marks, tL E., N. 1t. Remley, J. D. Seago and D. W. Hastings. Effects of bilateral lesions of the olfactory bulbs of rats on measures of learning and motivation. Physiol. Behav. 7: 1-6, 1971. 6. Phillips, D. S. and G. K. Martin. Effects of olfactory bulb ablation upon heart rate. P~ysiol. Behav. 7: 535-538, 1971. 7. Phillips, D. S. and G. K. Martin. Heart rate conditioning of anosmic rats. Physiol. Behav. 8: 33-36, 1972. 8. Sieck, M. H. The role of the olfactory system in avoidance learning and activity. Physiol. Behav. 8: 705-710, 1972. 9. Sieck, M. H. Selective olfactory system lesions in rats and changes in appetitive and averJive behavior. Physiol. 8ehav. 10: 731-739, 1973. 10. Sieck, M. H. and B. L. Gordon. Selective olfactory bulb lesions: Reactivity changes and avoidance learning in rats. Physiol. Behav. 9: 545-552, 1972.

11. Sieck, M. H. and B. L. Gordon. Anterior olfactory nucleus or lateral olfactory tract destruction in rats and changes in aversive and appetitive behavior. Physiol. Behav. 10: 1051-1059, 1973. 12. Sieck, M. H., J. F. Turner, B. L. Gordon and R. G. Struble. Some quantitative measures of activity and reactivity in rats after selective olfactory lesions. Physiol. Behav. 11: 71-79, 1973. 13. Smith, R. F. and F. R. Keller. Free-operant avoidance in the pigeon using a treadle response. J. exp. Analysis Behav. 13: 211-214, 1970. 14. Thomas, J. lk Some behavioral effects of olfactory bulb damage in the rat. J. comp. physiol. Psychol. 83: 140-148, 19~3. 15. Ueki, S. and H. Sugano. Effect of olfactory bulb lesion on behavior. Abstracts o f Papers. 23rd Int. Cong. Physiol. Sci., Tokyo, Japan, 1965, Abstract No. 1095. 16. Wenzel, B. M. The olfactory system and behavior. In: Advances in Limbic and Autonomic Research, Vol. 1, edited by L. V. DiCara. New York: Plenum, in press. 17. Wenzel, B. M., P. F. Albritton, A. Salzman and T. E. Oberjat. Behavioral changes in pigeons after olfactory nerve section or bulb ablation. In: Olfaction and Taste 3, edited by C. Pfaffmann, New York: Rockefeller Univ. Press, 1969, pp. 278-287. 18. Wenzel, B. M. and A. Salzman. Olfactory bulb ablation or nerve section and behavior of pigeons in nonolfactory learning. ExplNeurol. 22: 472-479, 1968.