EFFECTS OF PSYCHOTROPIC AGENTS ON SIDMAN AVOIDANCE RESPONSE IN GOOD AND POOR-PERFORMED RATS

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1 EFFECTS OF PSYCHOTROPIC AGENTS ON SIDMAN AVOIDANCE RESPONSE IN GOOD AND POOR-PERFORMED RATS SHUJI TAKAORI, NORIKO YADA AND GENZABURO MORI Department of Pharmacology, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto Received for publication May 27, 1969 The previous paper (1) has shown that effects of psychotropic agents on the explo ratory behavior of rats placed into a symmetrical Y-shaped box are modified by degree of familiarity with the apparatus. In the experienced rats which had received the train ing trials in the box three or four times, chlorpromazine and diazepam produced a pro longation of the start latency with decrease in the ambulation and rearing scores. However, diazepam shortened the start latency and increased the ambulation scores in the inex perienced animals which had never been placed in the apparatus. Although major and minor tranquilizers have been found to interfere with normally trained conditioned avoidance responses (2-5), information about the effects of these agents on poor performance of rats is still insuficient. Rech (6, 7) has reported that am phetamine and scopolamine improve conditioned avoidance behavior for rats which per formed poorly, in spite of extensive training, in a shuttle-box procedure. The present experiments were designed to determine effects of several psychotropic agents on Sidman type avoidance responding in the good and poor-performed rats. METHODS Thirty-eight male rats of Wistar strain, weighing about 150 g (about 60 days of age) at the start of the training schedule, served as the subjects. They were housed in an in dividual cage at the room temperature of 22 + I 'C. The individual daily food-intake after the onset of the training was restricted to 15 g of a commercial diet (CLEA CA-1) allowing free access to drinking water. Sidman-type avoidance apparatus (Kyoto Keisoku Co.) was used. The experimental box, made of wood, had a floor area of 20 cm by 15 cm and an inside height of 28 cm. The floor was a grid composed of 7-mm stainless steel rods at a space of 10 mm. The inside walls of the box were covered with grids of 5-mm stainless steel rods at a space of 10 mm. Both the floor and side wall rods were charged with an alternate current at 300 volts and 5 ma. A press lever was placed at one side wall 2 cm above the floor and pro truded 3 cm into the box. To prevent the lever-presses resulting from jumping responses, a false ceiling of plastic sloped from the top rear of the box to a position 4 cm above the lever.

2 FIG. 1. Examples of recording. P : Lever-press, S : Shock, AR : Avoidance response, ER : Escape response, EF : Escape failure. Time scale : 10 seconds. The rat was placed individually in the box for 30 minutes every day except Sunday, and frequencies of the lever-presses and electroshock performances were recorded on an ink-writing oscillograph (Fig. 1). Whenever the animal pressed the lever in the box, it postponed the onset of shock for 40 seconds. When the animal did not press the lever during 40 seconds after the last lever-press, shock came on to both the floor and side wall rods. The shock duration was fixed at 1.0 second. After a shock, further shocks were presented at 10-second intervals until the rat pressed the lever. When the animal pressed the lever during the first shock or before an arrival of the next one, this lever-press per formence was designated as an escape response. The difference between the total number of lever-presses and escape responses for 30 minutes represented the frequency of avoidance responses, and that between the total number of shocks and escape responses corresponded to escape failure: i.e. Avoidance responses= Number of lever-presses-escape responses Escape failures=number of shocks-escape responses Test agents used were chlorpromazine hydrochloride, haloperidol, diazepam, nitra zepam, chlordiazepoxide, methamphetamine hydrochloride and imipramine hydrochlo ride. Chlorpromazine and methamphetamine were dissolved in physiological saline, and diazepam, nitrazepam and chlordiazepoxide were suspended in 0.5% carboxymethylcel lulose solution. Haloperidol (Serenace, Dainippon Seiyaku Co.) and imipramine (To franil, Fujisawa Co.) were commercial injection preparations. Various doses of the test agents were injected intraperitoneally 30 minutes before the placement of the animals to the experimental box. The number of animals used for one dose was 6 to 7. The ad ministration of the test agents was repeated at the time-interval of at least three days in the rats which had received the training trials more than 20 sessions. The control study

3 was performed in the animals injected physiological saline alone. The statistical significance of the data was determined by Student's t-test. RESULTS I. Preliminary experiments Thirty-eight rats which received the training trials more than 20 sessions were clas sified into the following four types : 1) good-performed, 2) poor-performed, 3) non-performed and 4) variably responded rats. 1) Good performed rats In 9 out of 38 rats, the frequency of lever-presses increased steeply from the second or third trial and the number of avoidance responses after the 10th trial was always more that 100 during 30 minutes. It was usually 150 to 200. Consequently, the total number of shocks and escape failures decreased proportionally to an increase in the frequency of Sessions Fir 2. Examples of avoidance learning curve of good and poor-performed rats for first 30 sessions.

4 avoidance responses. The frequency of escape failures after the 10th trial was less than 5 per 30 minutes. The upper panel in Fig. 2 demonstrates an example of development of the avoidance responses, shocks and escape failures in a good-performed rat. 2) Poor performed rats Twenty rats showed the frequency of avoidance responses below 50 during 30 minutes, in spite of the repetition of trials more than 20 times. The shocks received in 30 minutes were 30 to 70 of steady level from the 15th trial. The frequency of escape failures decreased gradually until the 15th trial, but thereafter no animal exhibited the number of escape failures less than 5. The lower panel in Fig. 2 illustrates an example of development of the avoidance responses, shocks and escape failures in a poor-performed rat. 3) Non performed rats In 3 out of 38 rats, the frequencies of avoidance responses and escape responses were always below 10 during 30 minutes, in spite of the extensive training more than 20 times. These animals received shocks almost throughout the experimental period. These non performed rats were excluded from further experiments. 4) Variably responded rats Remaining 6 rats showed the variable frequencies of avoidance responses and escape failures even after the repetition of the training trials more than 20 times. This group of the animals was also discarded from further experiments because a clear-cut determina tion of the drug effects could not be allowed. II. Effects of drugs The effects of drugs on the avoidance responses, shocks and escape failures were com pared between the good and poor-performed rats. 1) Chlorpromazine The good-performed rats treated with chlorpromazine in intraperitoneal doses of TABLE 1. Effects of chlorpromazine and haloperidol on frequencies of the avoidance responses, shocks and escape failures in the good and poor-performed rats. Six animals were used in each group. Values are mean±s.e. Significantly lower than corresponding control value : --F P<0.01. Significantly higher than corresponding control value : **P<0.01, *P<0.05.

5 1.25, 2.5 and 5 mg/kg showed a dose-dependent lowering of the avoidance response rate with an increase in the number of shocks (Table 1). These changes were significant at P=0.01 compared with corresponding control value. However, chlorpromazine in doses less than 2.5 mg/kg did not affect the escape failures, though a dose of 5 mg/kg produced a slight increase in the frequency of escape failures. These three parameters were restored to the control level at the next trial which carried out at 24 hours after the drug administ ration. In the poor-performed rats, the frequency of avoidance responses was significantly (P<0.01) decreased in doses of 2.5 and 5 mg/kg of chlorpromazine, and that of escape failures was evidently increased. 2) Haloperidol The intraperitoneal injection of 0.125, 0.25 and 0.5 mg/kg of haloperidol produced a significant decrease (P<0.01) in the frequency of avoidance response with an increase in the number of shocks and escape failures in the good-performed rats (Table 1). In the poor-performed rats, a significant lowering (P<0.01) of the avoidance response rate was observed in doses higher than 0.5 mg/kg, and the frequencies of shocks and escape failures were significantly (P<0.01) increased in dose over 0.25 mg/kg. 3) Diazepam The effects of diazepam on the avoidance behavior of the good-performed rats were almost the same as those of chlorpromazine. As shown in Table 2, the avoidance response TABLE 2. Effects of diazepam, nitrazepam and chlord iazepoxide on frequencies of the avoidance responses, shocks and escape failures in the good and poor-performed rats. Six or seven animals were used in each group. Values are mean -t S.E. Significantly lower than corresponding control value : ++P<0.01, +P<0.05. Significantly higher than corresponding control value : **P<0.01, *P<0.05.

6 rate was markedly reduced in doses higher than 2.5 mg/kg of diazepam. A significant increase (P<0.01) in the number of shocks and escape failures was obtained in doses over 5 mg/kg. In the poor-performed rats, however, the effects of diazepam were quite different from those of chlorpromazine. A significant increase (P<0.01 or P<0.05) in the frequency of avoidance responses and a slight decrease in the number of shocks and escape failures were found in doses from 1.25 to 10 mg/kg, though the animals ex hibited a relaxation of the skeletal muscle. 4) Nitrazepam The effects of nitrazepam on the avoi dance behavior were the same as those of diazepam in both good and poor-performed Fin. 3. Effects of nitrazepam (N) in a daily rats (Table 2). The frequency of avoidance dose of 2.5 mg/kg on the frequency of avoidance responses and shocks in 7 poor responses was reduced in proportion to doses performed rats. Each point represents of nitrazepam in the good-performed rats, mean E S. E. S : Saline, R : Reserpine (0.5 mg/kg but it was markedly increased in the poor i.p.) was injected immediately after the third trial. performed ones. Therefore, the frequencies of shocks and escape failures were increased in the former type of animals and they were decreased in the latter one. These changes in the avoidance responses and shocks caused by 2.5, 5 and 10 mg/kg were significant at P=0.01 or P=0.05 in both good and poor performed rats. Fig. 3 shows the effects of repetitive administration of nitrazepam in a daily dose of 2.5 mg/kg on the frequencies of avoidance responses and shocks in the poor-performed rats. Nitrazepam produced a marked increase of avoidance response rate with decreased shock frequencies. These responses of the avoidance behavior to nitrazepam were not modified by previous treatment of the animals with reserpine in an intraperitoneal dose of 0.5 mg/kg 24 hours before the injection of nitrazepam. 5) Chlordiazepoxide The good-performed rats received 5 to 40 mg/kg of chlordiazepoxide showed the decreased avoidance response rate wita increased shock frequencies (Table 2). The frequency of escape failures was also sig:zi`icantly (P<0.01) increased in doses higher than 20 mg/kg. In the poor-performed rats, however, the doses of chlordiazepoxide increased slightly the frequency of avoidance responses. The effect of chlordiazepoxide on the shocks and escape failures was variable and it did not allow the existence of a dose-response relationship.

7 TALE 3. Effects of methamphetamine and imipramine on frequencies of the avoidance responses, shocks and escape failures in the good and poor-performed rats. Six or seven animals were used in each group. Values are mean -?-S.E. Significantly lower than corresponding control value : +P<0.05. Significantly higher than corresponding control value : **P<0.01, *P< ) Methamphetamine As shown in Table 3, the intraperitoneal injection of methamphetamine enhanced the frequency of avoidance responses in both good and poor-performed rats, and the in crease caused by 0.25, 0.5 and 1 mg/kg was significant at P=0.01 or P=0.05. Therefore, the number of shocks was slightly reduced by the administration of methamphetamine. The decrease in the frequency of escape failure caused by 0.25 mg/'kg was also significant at P=0.05 in the poor-performed rats. 7) Imipramine Imipramine in doses less than 5 mg/kg did not affect the number of avoidance re sponses, shocks and escape failures in both good and poor-performed rats (Table 3). However, a dose of 10 mg/kg produced a lowering of the avoidance response rate with an increase in the number of shocks and escape failures, especially in the poor-performed animals. DISCUSSION The avoidance procedure presented by Sidman (8-10) is a non-discriminated avoidance or continuous avoidance, which have no explicit warning stimulus before delivary of shock. In this avoidance schedule, there are two temporal variables: the avoidance response shock and shock-shock intervals. The response-shock interval is the period by which each avoidance response postpones the onset of shock, and the shock-shock interval is the time lapse between two consecutive shocks if no avoidance response occurs between them. Sidman (9) and Rey et al. (11) have reported the effects of these intervals on the establish ment and maintenance of the avoidance behavior. In the present experiments, the re sponse-shock interval of 40 seconds and the shock-shock interval of 10 seconds were adopted according to the method of Randall et al. (5). Many investigators have utilized the

8 Sidman-type avoidance procedure for measurement of potency of psychotropic agents, but the rats used are limited to the good-performed ones which learn well to press the lever at a steady rate in order to avoid shock. In the present experiments, the frequencies of avoidance responses, escape responses and escape failures were calculated for 30 minutes every day. After the training trials more than 20 sessions, about 1 /4 of the rats tested showed the pattern of the good-performed ones but about half of them remained in the group of the rats which performed poorly in spite of the extensive training. Table 4 summarized the effects of psychotropic agents on the avoidance responses and escape failures in the good and poor-performed rats. In the good-performed ones, chlorpromazine was shown to suppress the avoidance responses at doses which had no effect on the escape failures. These results coincide well with the reports described by Ader and Clink (2) and Low et al. (12), though the methods of the avoidance procedures are different each other. In the poor-performed rats, the major tranquilizers produced a lowering of the avoidance response rate with an increase in the number of escape failures. The effects of the minor tranquilizers such as diazepam, nitrazepam and chlordia zepoxide on the avoidance behavior of the good-performed rats were as same as those of the major tranquilizers. In the poor-performed rats, however, the minor tranquilizers except larger doses of chlordiazepoxide produced the increased avoidance response rate with a decrease in the frequency of escape failures. These results have something in com mon with the previous findings (1) that diazepam produced a marked increase in the exploratory activities of naive rats placed into a Y-shaped box. Sachs et al. (13) have reported that the rats trained after the injection of chlordiazepoxide acquire the conditioned avoidance response significantly faster than saline controls. Carlton (14) and Stein (15) have maintained that an increase of the rate of behavioral responses is mediated by stimu lating the adrenergic mobilizing system in the brain or by blocking the central cholinergic inhibitory system. In the present experiments, the pretreatment of the poor-performed rats with reserpine did not modify the increased pattern of the avoidance responses in duced by nitrazepam. Therefore, it is hard to consider that the enhancement of the

9 avoidance responses caused by the minor tranquilizers is directly related to the central adrenergic mobilizing system. Centrally active anticholinergic agents such as atropine and scopolamine have been found by Rech (7) to improve the conditioned avoidance behavior of the poor-performed rats in a shuttle-box procedure. Leaf and Muller (16) have demonstrated that scopolamine results in higher response rates and lower shock rates than controls during the acquisition of the Sidman-type avoidance responses. Improvement of the avoidance behavior in the rats treated with the anticholinergic agents may be ex plained on the basis of blocking the central cholinergic inhibitory system, but the precise mechanism of enhancement of the avoidance responses caused by the minor tranquilizers is still unknown. Methamphetamine was found to increase the frequency of the avoidance responses in both good and poor-performed rats. The agent has been considered by Stein (15) and Moor (17, 18) to stimulate the central adrenergic mobilizing system probably by facilitating release of brain noradrenaline. On the other hand, the significance of the results obtained with imipramine in the present experiments was not apparent. Scheckel and Boff (19) have reported that imipramine potentiates the effect of amphetamine on the Sidman-type avoidance response in rats, and the previous paper (1) has also confirmed a potentiation by imipramine in the effect of methamphetamine on the exploratory be havior. Further investigations into effects of imipramine on the Sidman-type avoidance situation are required in connection with changes in the metabolism of brain noradrenaline. SUMMARY The effects of several psychotropic agents on the Sidman-type avoidance response were compared between the good and poor-performed rats. 1. Major tranquilizers such as chlorpromazine and haloperidol produced a signifi cant decrease in the frequency of avoidance responses with an increase in the number of shocks in both good and poor-performed rats. The inhibitory effects of these agents on the avoidance behavior were more striking in the former group of animals than in the latter one. 2. The good-performed rats treated with minor tranquilizers such as diazepam, nitrazepam and chlordiazepoxide showed a significant lowering of the avoidance response rate with an increase in the number of shocks. In the poor-performed rats, however, these agents enhanced markedly the frequency of avoidance responses with a decrease in the number of shocks and escape failures. The marked increase in the avoidance response rate caused by nitazepam was not modified by pretreatment of the animals with reserpine. 3. Methamphetamine produced a significant increase in the frequency of avoidance responses in both good and poor-performed rats. Imipramine in lower doses did not affect the avoidance behavior, but the higher doses reduced the frequency of avoidance responses with an increase in the number of shocks.

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