Psychopharmacology

Psychopharmacology (1987) 92:508 512

© Springer-Verlag 1987

Effects of ethylketazocine and morphine alone and in combination with naloxone on schedule-controlled behavior in pigeons J.L. Katz Preclinical Pharmacology Branch, National Institute on Drug Abuse, Addiction Research Center, PO Box 5180, Baltimore, MD 21224, USA Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine

Abstract. The behavioral effects of morphine and ethylketazocine were compared in pigeons responding under multiple fixed-interval, fixed-ratio schedules of food presentation. Both morphine and ethylketazocine produced dose-related decreases in rates of responding maintained under either schedule. Maximal effects of morphine were observed about 15~45 rain after injection and typically lasted the entire session (about 60 rain). Effects of ethylketazocine had a faster onset (maximal effects were observed within 15 min after injection), and shorter duration (effects diminished within the session). Ethylketazocine and morphine had similar potencies. Dose-effect curves for both drugs were shifted to a similar degree by naloxone. Key words: Ethylketazocine - Morphine Naloxone Competitive antagonism - Schedule-controlled behavior Pigeons

The different effects of several opioid agonists led Martin and co-workers to suggest different types of opioid receptors mediating those effects (Gilbert and Martin 1976; Martin et al. 1976). Mu receptors were proposed as those mediating the effects of morphine-like opioids, kappa receptors were proposed as those mediating the effects of ketazocine-like opioids, and sigma receptors were proposed as those mediating the effect of drugs having effects like Nallylnormetazocine (SKF-10047). Some of the evidence for multiple opioid receptors has come from studies showing different antagonist potencies of a single opioid antagonist against the effects of different agonists. In the guinea-pig ileum, for example, naloxone shifted the dose-effect curve for the mu agonist morphine more than dose-effect curves for the kappa agonists ketazocine or ethylketazocine (Kosterlitz et al. 1974). Behavioral evidence for multiple opioid receptors has come from studies of drug discrimination in laboratory animals. In rhesus monkeys trained with the mu agonist etorphine as a discriminative stimulus, other mu agonists such as morphine produced responding like that produced by etorphine, whereas kappa agonists such as ketazocine and ethylketazocine produced responding like that produced by vehicle (Herling and Woods 1981a). Further, in rhesus monkeys trained with ethylketazocine as the discriminative stimulus, morphine produced responding like that produced

by vehicle, whereas other kappa agonists, such as ketazocine, produced responding like that produced by ethylketazocine (Hein et al. 1981). In contrast to effects observed in primates, studies of drug discrimination in pigeons have found that ketazocine and ethylketazocine produced responding like that produced by morphine in pigeons trained with morphine as the discriminative stimulus (Herling et al. 1980). Further, morphine produced responding like ethylketazocine in subjects trained with ethylketazocine as the discriminative stimulus (Hein et al. 1980). The above results have led to the suggestion that the effects of mu and kappa agonists are mediated differently in primates and similarly in pigeons (Herling and Woods 1981b). If the effects of mu and kappa agonists are mediated similarly in pigeons and dissimilarly in other species, the potencies of an opioid antagonist in antagonizing the effects of mu and kappa agonists should be similiar in pigeons and dissimilar in the other species. There have been few studies directly comparing antagonist effects against different agonists. In rodents, naloxone shifted dose-effect curves for morphine effects on schedule-controlled responding more than dose-effect curves for ethylketazocine (Harris 1980). Similarly, apparent pA2 values for naloxone antagonism of discriminative-stimulus effects in rats (Herling and Shannon 1982) or antagonism of analgesic effects in mice (Ward and Takemori 1983) differed for morphine and ethylketazocine. In pigeons, the doses of naloxone reported to antagonize the decreases in response rates produced by ketazocine or ethylketazocine were higher than the dose required to reverse the rate-decreasing effects of phenazocine, a mu agonist (Leander 1982). In a study of drug discrimination with pigeons, naltrexone shifted the morphine dose-effect curves for rates of responding and discriminative effects less than if shifted the dose-effect curves for ethylketazocine (Herling et al. 1984). Thus, the antagonism data in pigeons are inconsistent, with only some results similar to those obtained in other species, suggesting that the actions of mu and kappa agonists are mediated by distinct receptors. The present study was designed to study further the behavioral effects of prototypic mu and kappa agonists and their interaction with the opioid antagonist, naloxone. Pigeons were trained to respond under a multiple fixed-interval, fixed-ratio schedule of food reinforcement and the effects of morphine and ethylketazocine were studied alone and in combination with naloxone.

509 Methods

Subjects. Four adult male White Carneaux pigeons (Columba livia) were studied. The subjects were maintained at 80-85% of their unrestricted-feeding weights by adjusting their access to food between experimental sessions. Water and grit were always available in the home cages. All subjects had been studied previously under schedules identical to those described below and had received various drugs, but not more frequently than twice per week. Subjects used in these studies were maintained in accordance with guidelines of the Animal Care Committee of the Addiction Research Center and the Committee on Care and Use of Laboratory Animals of the Institute of Laboratory Animal Resources, National Research Council (Department of Health, Education and Welfare Publication No. (NIH) 78-23, revised 1978).

Apparatus. Experimental sessions were conducted with the pigeon in a chamber similar to the one described by Ferster and Skinner (1957), which was provided with white noise to mask extraneous sounds. A translucent response key (model B, R. Gerbrands Co.) was mounted on the front wall of the chamber. Each peck on the key with a minimal force of 0.15 N produced an audible click of a relay within the chamber and was recorded as a response. A solenoidoperated food magazine (model B, R. Gerbrands Co.) could provide 4-s access to mixed grain through an aperture in the front wall which was illuminated during food delivery. Colored lamps could transilluminate the response key and serve as visual stimuli.

Procedure. Responding was maintained by presentation of food under a multiple schedule with fixed-interval and fixed-ratio components. Sessions started with the fixed-interval component during which green lights transilluminated the response key and the first response after the lapse of 300 s produced food. Each fixed-interval component was followed by a fixed-ratio component during which blue lights transilluminated the response key and the 30th response produced food. If a response was not emitted within 60 s of the lapse of the fixed interval, or if 30 responses were not emitted within 60 s during the fixed-ratio component, the component automatically switched to the other without food presentation. Sessions ended after 12 components of each type (about 65 rain).

Drugs and injection procedures. Ethylketazocine methane sulfonate was dissolved in sterile water to which lactic acid was added; if necessary, sodium hydroxide was added to the solution to adjust the pH to above 4. Morphine sulfate and naloxone hydrochloride were dissolved in a 0.9% saline solution. Drug solutions were diluted so that each dose could be injected in a volume of no more than 1.0 ml/kg body weight. Control injections were similar volumes of vehicle. Drugs were injected IM (pectoral muscle); when two drugs were studied in combination, each was injected in a different side of the breast. Injections were given 5 rain before the session was started. Doses are expressed as the salts. Experimental sessions were conducted daily, Monday through Friday. At least 6 days intervened between drug administrations which were typically given on Tuesday or Friday. Vehicle-control sessions were conducted every Thursday. Drugs and drug combinations were studied in

different orders with different subjects, and a complete dose-effect curve was usually determined for one drug or combination before another drug or combination was studied. Doses of morphine studied ranged from 0.1 to 30.0 mg/ kg (0.13-39.54~mol/kg); doses of ethylketazocine also ranged from 0.1 to 30.0 mg/kg (0.25 75.74 ~mol/kg). Doses of each drug or drug combination were studied once or twice in each of three pigeons.

Measurement of effects. Average rates of responding were computed each session by dividing total responses by elapsed time for individual subjects. Effects of each drug or drug combination are expressed as a percentage of the average rate of responding during control sessions that preceded sessions in which that drug or drug combination was studied. Since effects in individual subjects were well represented by the average of all subjects, results are presented as averages of all subjects. For assessment of the time course of effects of each drug, response rates were calculated for successive groups of three fixed intervals. The pattern of responding during the fixed interval was quantified with the quarter-life statistic, which estimates the per cent of the interval elapsed when 25% of the responses have been emitted (Herrnstein and Morse 1957; Gollub 1964). Quarter-life values of 25% indicate a constant rate of responding throughout the interval; values greater than 25% indicate responding predominantly in the latter parts of the fixed interval. Results

Controlperformances. Performances after vehicle were similar to those described previously under the multiple schedule (Ferster and Skinner 1957). Under the fixed-interval schedule, little or no responding occurred early in the interval and was followed by increased response rates later in the interval (Fig. 1, Control). Under the fixed-ratio schedule, a brief pause was followed by a high response rate that was sustained until food presentation. Response rates under the fixed-ratio schedule were uniformly higher than rates under the fixed-interval schedule (Fig. 1). Effects of ethylketazocine and morphine. Ethylketazocine produced dose-related decreases in response rates. Under the fixed-interval schedule, rates were decreased at doses of 1.0 mg/kg and higher (Fig. 2, upper panel, open circles); under the fixed-ratio schedule, rates were decreased at doses of 0.3 mg/kg and higher (Fig. 2, lower panel, open circles). At the highest doses, responding of pigeons was virtually eliminated for the entire session (Fig. 1). Ethylketazocine produced dose-related decreases in quarter-life values only at doses that also decreased response rates (Fig. 3, open circles). Morphine also produced dose-related decreases in response rates in pigeons. Decreases in response rates occurred at doses of at least 1.0 and 0.3 mg/kg under the fixed-interval and fixed-ratio schedules, respectively (Fig. 2, open triangles). As with ethylketazocine, morphine decreased quarter-life values in a dose-related manner in pigeons (Fig. 3, open triangles). Morphine and ethylketazocine were of approximate equal potency in the pigeon. For either drug, the lowest doses that decreased response rates were 1.0 and 0.3 mg/kg under the fixed-interval and fixed-ratio schedules, respec-

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Fig. 2. Effects of ethylketazocine and morphine on responding of pigeons under the multiple fixed-interval, fixed-ratio schedule of food presentation. Abscissae: dose of drug in mg/kg, log scale. Ordinates: rate of responding expressed as a percentage of the average rate during vehicle-control sessions. Vertical lines above C show the average of individual __SD values for vehicle control sessions for three pigeons. Top panel: effects on rates of responding under the fixed-interval component. Bottom panel: effects on rates of responding under the fixed-ratio component of the schedule. Circles: effects of ethylketazocine. Triangles: effects of morphine. Open symbols: effects of the drugs given alone. Filled symbols: effects of the drugs given with naloxone (1.0 mg/kg)

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Fig. 1. Cumulative records showing effects of ethylketazocine and morphine alone and in combination with naloxone on responding maintained by food presentation under the multiple fixed-interval fixed-ratio schedule in pigeon P-464. Abscissae, time; ordinates, cumulative responses. Diagonal marks show presentations of food. The response pen reset after each 1100 responses. The event line is displaced downward during the fixed-ratio component of the schedule. Note that both ethlylketazocine and morphine decreased rates of responding and that naloxone antagonized the decreases in response rates and restored the temporal patterns of responding to resemble control performances

tively. A t the higher doses, morphine decreased response rates to a degree somewhat less than ethylketazocine. W h e n considering the potency on the basis of pmol/kg, the two drugs were also o f approximate equal potency, with morphine slightly more effective than ethylketazocine at the lower doses. Further, the two drugs were not appreciably

LLH 7 0 - 1 ~ 60504"0rv 30W I-- 20,~ IO:D o 0 TI I I I I C 0.1Q'3 1.0 3.0 I0.0 30,0 DRUG DOSE (mg/kg) Fig. 3. Effects of ethylketazocine and morphine on patterns of responding under the fixed-interval schedule of food presentation in pigeons. Abscissae: dose of drug in mg/kg, log scale. Ordinates: quarter-life value as percentage of the interval. Verical lines above C show the average of individual 4- SD values for vehicle-control sessions for three subjects under each condition. Quarter-life values are not shown if average response rates were less than or equal to 15% of control response rates. Symbols are as in Fig. 2 different in potency when comparing only the portions o f the sessions in which the maximal effect occurred (data not shown).

Effects of ethylketazoeine and morphine in combination with naloxone. N a l o x o n e (1.0 mg/kg) shifted the dose-effect

511 curves for both ethylketazocine and morphine under both schedules to the right (Fig. 2; compare open and filled circles and triangles) with the ethylketazocine dose-effect curve shifted to the right slightly more than the morphine curve. Figure 1 shows performances after doses of ethylketazocine or morphine that produced marked decreases in response rates and altered the temporal patterning in responding. When naloxone was also administered, neither response rates nor the temporal patterning of responding were different from those observed after vehicle injection. Figure 3 shows that the effects of ethylketazocine and morphine on quarter-life values were also shifted by naloxone to the right (compare open and filled circles and triangles).

Time course of effects of ethylketazocine and morphine. Rate of responding during the fixed-interval schedule was decreased by ethylketazocine most prominently in the first three portions of the session after doses of 1.0 or 3.0 mg/kg. Since each portion was approximately 15 rain and injections were given 5 rain before the start of the session, the first three portions of the session correspond approximately to minutes 5 through 50. Rates of responding approached or recovered to control values at each of those doses in the last portion (approximately minutes 50 through 65) of the session. Naloxone (1.0 mg/kg) antagonized the decreases in rates throughout the approximate 65-rain session. At an ethylketazocine dose of 10.0 mg/kg with naloxone, response rates were decreased by about 50% with little change throughout the session (data not shown). Morphine at a dose of 1.0 mg/kg decreased fixed-interval response rates after the first portion of the interval (approximately rain 20 through 65) to about 50% of control rates. The decreases in response rates were antagonized by 1.0 mg/kg naloxone. A higher dose of morphine (3.0 mg/kg) produced decreases earlier in the session, with effects increasing as the session progressed throughout the first 50 rain. The decreases produced by 3.0 mg/kg were antagonized by naloxone after the first 20 min of the session. Morphine at 10.0 mg/kg with naloxone produced decreases in response rates that were uniform throughout the 65-min session (data not shown). Discussion

In the present study, responding of pigeons was maintained by food presentation under a multiple fixed-interval, fixedratio schedule. The effects of morphine and ethylketazocine on behaviors were qualitatively similar. Both ethylketazocine and morphine decreased rates of responding as has been shown in several previous studies (Harris 1980; Herling et al. 1980; Hein et al. 1981). The two drugs decreased responding comparably under both fixed-interval and fixedratio schedules as has also been reported previously (McMillan and Morse 1967; Leander 1982). The primary difference between the effects of ethylketazocine and morphine on schedule-controlled behavior was their time courses. For example, effects of ethylketazocine generally occurred within 20 rain of injection. In contrast, maximal effects of morphine were typically observed between 20 and 50 rain after injection. Effects of ethylketazocine were also of shorter duration, with effects often noticeably diminished within 50-65 rain after injection. The effects of morphine at intermediate and higher doses showed no evidence of diminishing during the session.

Effects of both opioid agonists on response rates were antagonized in a surmountable manner by the opioid antagonist naloxone as has been reported in earlier studies in this and other species (e.g., Harris 1980; Goldberg et al. 1981). Further, large disruptions in the temporal patterns of responding that were produced by either drug alone were absent when naloxone was also administered. Temporal patterns of responding after combinations of selected doses of the agonists with naloxone resembled control performances in all important aspects. In the present study, dose-effect curves for morphine were shifted by 1.0 mg/kg naloxone to the right by about 1 log unit. Previous studies have reported comparable shifts in morphine dose-effect curves after the same (Goldberg et al. 1981) or lower (McMillan et al. 1970; Downs and Woods 1976) naloxone doses. Naloxone shifted the ethylketazocine dose-effect curves to the fight by more than 1 log unit. In contrast, a previous study reported a smaller shift of the ethylketazocine dose-effect curve produced by the same dose of naloxone used in the present study (Leander 1982; shifts of about 1 log unit were estimated from his Fig. 3). The differences in potency of naloxone as an antagonist of ethylketazocine and mu agonists lead to the conclusion that the effects of ethylketazocine on response rates in the pigeon are mediated by different receptor mechanisms than are the effects of mu-receptor agonists (Leander 1982). The present results, however, did not show a different potency of naloxone as an antagonist of morphine and ethylketazocine. In the study by Leander (1982), drugs were given twice per week, a frequency that may have allowed tolerance to develop. Indeed, ethylketazocine was more potent in the present study when drugs were administered no more frequently than once per week. Further, while ethylketazocine was more potent in the present study than in the study by Leander, the dose-effect curves for ethylketazocine in combination with naloxone were not appreciably different in the two studies. Thus, when agonists are administered only once per week, results of antagonism of effects of morphine and ethylketazocine by naloxone are consistent with results of studies of drug discrimination that suggest that the effects of the two agonists are mediated by similar mechanisms.

Acknowledgments. The author thanks R.D. Clark, C. Edwards and S. Fowler for technical assistance, and D. Shelton and M. Hawkes for help in preparation of the manuscript. This work was supported in part by US Public Health Service Grants DA00154, and DA 03113 awarded to the University of Michigan Medical School, and DA 03505 awarded to the University of Maryland School of Medmine.

References

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Received June 16, 1986 / Final version February 2, 1987