An experimental framework for evaluation of dependence liability of various types of drugs in monkeys

Sections

I. Introduction
II. Purpose and methods of each test
III. Summary of results and discussion
FIGURE 1
Influence of dosing schedule on development of physical dependence on morphine in monkeys (average of 3 monkeys)

Bibliography

Details

Author: T. YANAGITA
Pages: 57 to 64
Creation Date: 1973/01/01

An experimental framework for evaluation of dependence liability of various types of drugs in monkeys 1

T. YANAGITA
Director, Division of Medical Science, Medical Research Laboratory, Department of Psychopharmacology, Central Institute for Experimental Animals, Nogawa, Kawasaki, Japan.

I. Introduction

Drug abuse is a phenomenon developed as a result of the interaction of a drug, an individual, and the social environment. Therefore, for evaluation of the abuse potential of a drug, it is necessary to consider not only pharmacological effects of the drug, but also individual differences in mental and physical susceptibility to the drug and the present status of and social attitude toward the abuse of that type of drug or agent. There is, however, no doubt that the pharmacological effects of a drug play a primary role in the incidence of drug abuse.

For the evaluation of the pharmacological effects of a drug in terms of abuse potential, the first question to be asked is what type of effect should be explored. Tolerance and physical dependence have long been recognized as unique pharmacological phenomena specific to certain groups of drugs, and the ability of a drug to produce these phenomena has naturally been considered to be of great importance in predicting the abuse potential of a drug.

In the last decade, however, the concept of the psychopharmacological etiology of drug dependence and abuse has come to the fore. It is now a widely accepted observation that the pharmacological effects during which a subject develops a psychological desire to re-experience these effects after his first trial or first few trials of drug use plays a primary role in the initiation of drug abuse common to all types of drug. From an experimental viewpoint, this effect is called the "drug-seeking behaviour-reinforcing effect". Since this effect greatly contributes to the development of the primary psychological (or psychic) dependence on a drug, there can be no doubt of the necessity for evaluating the potency of this effect.

Another important aspect is the intensification of drug-seeking behaviour by the withdrawal syndrome when one is physically dependent on the drug. Here, the already existing desire for a drug is negatively reinforced by withdrawal aversion. Thus, one may develop secondary psychological dependence on the drug. Since the potency of this negatively reinforcing effect is a function of a subjective feeling of withdrawal aversion, it is not necessarily in proportion to the ability to produce physical dependence.

A pharmacological effect of a drug that produces unusual or anti-social behaviour during abuse is also important because the danger and harm of drug abuse frequently derive from such an effect. This effect, which can be called a "psychotoxic effect", may not be included in the concept of dependence liability in a strict pharmacological sense, but abuse potential and the danger of abuse of a drug cannot be evaluated without knowledge of this effect.

1

Supported by WHO Research Grant 1969-1972. The paper was read at the 5th International Congress on Pharmacology, San Francisco, July 1972. It is scheduled for publication as: Yanagita, T.: An experimental framework for evaluation of dependence liability of various types of drugs in monkeys. Pharmacology and the Future of Man. Proc. 5th Int. Congr. Pharmacology, San Francisco 1972, vol. 1 (Karger, Basel 1973).

Thus, the answer to the question of what type of effect should be explored is as follows:

  1. Drug-seeking behaviour-reinforcing effect;

  2. Ability to produce tolerance;

  3. Ability to produce physical dependence;

  4. Negatively reinforcing effect during withdrawal;

  5. Psychotoxic effect.

As a principle of the evaluation it must be realized that the route of administration and the dose regimen at which dependence liability of a drug is experimentally evaluated should not be determined by the clinically recommended usage but by the manner in which the drug is abused. Generally speaking, drugs are abused by routes of administration at which rapid absorption and rapid onset of the effects are expected (e.g., intravenous, if a drug is injectable), and at unusually high and frequent doses.

Based on the above considerations, the following comprehensive studies have been established as the routine method of our laboratory for evaluating drug-dependence liability of drugs.

II. Purpose and methods of each test

  1. Preliminary test of acute CNS effects of drugs in normal monkeys

Acute CNS effects of a drug are observed after single dose administration in normal monkeys. This test is conducted partly as a dose-range-finding test. The preparation form of a drug, drug concentration, route of administration, and dose level are the variables in this test. A standardized protocol is used, with behavioural manifestations, motor functions, autonomic signs, and body temperature as the major parameters of observation. The systemic drug effects and behavioural toxicity characteristic of each group of the principal drugs of abuse are observed, and the minimal effective dose, the toxic and lethal doses, and the duration of the effects are determined.

  1. Single-dose substitution test in monkeys physically dependent on morphine or barbital

The ability of a drug to support physical dependence in place of morphine or barbital (cross-physical dependence) is examined in this test by a single administration of the drug at doses previously determined in preliminary tests in physically dependent monkeys after withdrawal. This test is applicable to analgesic-antitussives and sedativehypnotics and thc testing procedure is the same as that established by Seevers (1936) for morphine in monkeys and similar to that established by Deneau for barbital in dogs (Seevers et al., 1966). When a test drug of the analgesic-antitussive type is found not to support dependence or to aggravate the withdrawal signs, the ability of the drug to precipitate withdrawal signs is examinated in morphine-dependent monkeys.

  1. Test of ability to produce physical dependence and tolerance in normal monkeys

The ability of a drug to produce physical dependence and tolerance is examined by repeated administration of the drug to normal monkeys 1 to 4 times a day, for 31 days or four weeks. The dose regimen is determined by the results obtained in the preliminary test and the single-dose substitution test. The systemic effects of the drug are observed daily, and if development of tolerance occurs, the dose is increased by 25-100 per cent. In the case of analgesic-antitussives, the procedure established by Seevers et al. (1966) is used, i.e., a precipitation test by nalorphine or naloxone on the 14th and 28th day of administration, and a natural-withdrawal test from the 32nd dayfor five days

. In the case of sedative-hypnotics, the precipitation test cannot be conducted, but the natural-withdrawal test is conducted for seven days. If the withdrawal manifestations are obscure in these withdrawal tests, the above procedures are repeated once or twice with increased doses.

  1. Cross self-administration test with a standard drug and saline

The ability of a drug to reinforce drug-seeking behaviour in the monkeys is examined in this procedure. It also serves as a dose-range-finding test for determining unit doses appropriate for the continuous-self-administration test and the progressive-ratio test. A monkey with an implanted intravenous catheter is chronically restrained in an individual cage. The monkey is allowed to self-administer a drug at will by pressing a lever switch on the wall of the cage. Monkeys that have been trained to discriminate between highly reinforcing drugs (e.g. cocaine or other stimulants) and saline are allowed to lever-press for one of the standard drugs, for saline, and for the test drug in turn, three days for each, 4 hours daily. The dose-response relationship is examined in terms of the unit dose versus the number of injections. This test is limited to intravenously injectable drugs.

  1. Continuous, intravenous or intragastric self-administration test

Monkeys are allowed to self-administer a drug without time or dose limitation. They are tested first with saline for one to two weeks, then with a test drug at an appropriate unit dose, which has been determined by the cross self-administration test, the preliminary test, or both. If a monkey initiates self-administration at a meaningful injection rate, the daily response rate and manifestation of drug effects will be observed for four to eight weeks. In this procedure, a 24- or 48-hour drug free period is imposed four weeks after the initiation of drug intake, and the number of lever presses and the severity of withdrawal are observed. If a monkey does not initiate self-administration in a two- to four-week self-administration period, the unit dose is increased or decreased and further observation is made for another two weeks. If the monkey still does not initiate self-administration, programmed administration of the drug is conducted every 2 to 4 hours for a further two weeks, and initiation during or after this programmed-administration period is observed.

  1. Progressive-ratio test in self-administration of a drug

A monkey is trained to self-administer a standard drug intravenously by pressing a lever 100 times for each dose. Then the standard drug is replaced by saline and observation is continued until the animal reduces the number of daily injections to less than half that for the standard drug. At this point, self-administration of a test drug is begun at a lever-press ratio of 100:1. Twenty-four hours later, a progressive-ratio test is begun in which the lever-press ratio is doubled progressively after a fixed number of injections. When the time interval between injections becomes longer than 24 hours for stimulants or 48 hours for depressants, the monkeys are said to have extinguished self-administration, and the lever-press ratio performed for the last dose is regarded as the final ratio of the test. For observation of intensification of drug-seeking behaviour due to the development of physical dependence and withdrawal aversion, programmed administration of a drug in one group and saline in another group precedes the progressive-ratio test.

III. Summary of results and discussion

Table I is a summary of a single-dose-substitution test on some analgesic antitussives. Our results on morphine, methadone, meperidine, and codeine agreed in general with those of tests conducted previously at the University of Michigan. This method is being used as a quick screening test for predicting physical-dependence liability. However, our recent experience has revealed that some drugs such as thebaine, azabicyclane (Yanagita et al., 1971), and propiram, which are liable to produce strong physical dependence (table II), do not suppress withdrawal but rather aggravate the signs in the withdrawn monkeys in which dependence had been maintained with high morphine levels (3 mg/kg s.c., 4 times/day). The drugs, however, suppressed withdrawal signs in monkeys in which dependence was maintained with a low dose of morphine (0.3 mg/kg s.c., 4 times/day). A similar effect had previously been found in man with propiram and profadol (Jasinski et al., 1971). These findings indicate that the physical-dependence-producing test by repeated administration of a drug to normal monkeys is indispensable for reliable prediction of physical-dependence liability. The use of low doses to maintain monkeys in routine tests is not advisable because of unstable withdrawal manifestations.

TABLE I

Single-dose-substitution test in morphine-dependent monkeys

(3 mg/kg, 4 times/day, s.c.)

 

Withdrawal signs

 

Drug

Suppression

Precipitation

Dose for complete suppression mg/kg s.c.

Morphine
+
-
3.0
Methadone
+
-
3.0
Oxymethebanol
+
-
3.0
Meperidine
+
-
10.0
Codeine
+
-
16.0
d-Propoxyphene
+
-
16.0
Thebaine
-
+
 
Azabicyclane
- (+)
+
 
Propiram
- (+)
+
 
Pentazocine
- (+)
+
 
Naloxone
- (-)
+
 
Saline
- (-)
-
 

( ) = Suppression in monkeys dependent on low dose of morphine (0.3 mg/kg, 4 times/day s.c.).

TABLE II

Development of physical dependence on analgesic antitussives

Drug

Route

Dose, mg/kg x times/day, for 31 days

Grade in natural withdrawal a

Morphine
s.c.
3 x 4
Severe
Oxymethebanol
s.c.
4 x 4
Severe
Meperidine
s.c.
5 x 4
Severe
Codeine
s.c.
16 x 4
Severe
 
p.o.
128 x 4
Intermediate
Thebaine
i.v.
self, 1 x 10-35
Severe
Azabicyclane
s.c.
20 x 4
Severe
d-Propoxyphene
s.c.
10 x 4
Intermediate
Propiram
s.c.
16 x 4
Intermediate
Pentazocine
s.c.
6x4
Mild

a Graded by Seevers' criteria (1936).

FIGURE 1

Full size image: 9 kB

Influence of dosing schedule on development of physical dependence on morphine in monkeys (average of 3 monkeys)

Influence of dosage schedule of morphine on development of physical dependence on the drug was observed by precipitating morphine withdrawal signs with a single dose of levallorphan in monkeys that were receiving morphine by 3 different dosing schedules. Each line represents the average of 3 monkeys. Mild, intermediate, and severe grades of withdrawal signs are indicated by I-II, III-IV, and V-VI, respectively (Seevers, 1936).

FIGURE 2

Full size image: 22 kB, FIGURE 2

Results of cross self-administration test on 5 stimulants with SPA (1-1,2-diphenyl-1-dimethylaminoethan-HCl) and saline in 4 self-administration-experienced monkeys. The ratio represents percentage of number of injections of the drug to that of SPA (0.1 mg/kg/ injection).

A summary of the physical-dependence-producing test on analgesic antitussives is shown in table II. As found in the table , the highest tolerable dose level was used in general in accordance with the concept described previously. The crucial role of dose regimen in this test is demonstrated with morphine in figure 1. It was also illustrated with barbiturates in a previous paper (Yanagita and Takahashi, 1970). d-Propoxyphene was ineffective by the oral route in rhesus monkeys. Our preliminary study on absorption and metabolism of the drug indicated a species difference between monkey and man when the drug was administered orally. Special attention should be paid to this factor when dependence liability is evaluated in animals. A summary of the physicaldependence tests on barbiturates and some other sedative hypnotics is shown in table III. Many other benzodiazepine derivatives were found to be similar to diazepam or chlordiazepoxide, although some of them were less potent in producing physical dependence.

The results of the single-dose-substitution test with sedative-hypnotics corresponded qualitatively to those of a physical-dependence-producing test using the same drugs, but quantitatively the ability of a drug to suppress withdrawal did not always parallel the ability to produce physical dependence. Chronic maintenance of physical dependence on barbital for the single-dose-substitution test was more difficult than on morphine because of complications such as liver damage, marasmus, and accidental suffocation.

Results of the cross self-administration test on five stimulants are shown in figure 2. In this case, SPA ( l-1-2-diphenyl-1-dimethylaminoethan HCl), a CNS stimulant, was used, and the reinforcing effect was clearly demonstrated with these drugs at appropriate unit doses. As the unit dose decreases the injection ratio increases, but if the unit dose becomes too low to reinforce the response, the ratio will drop down to the saline level. Thus, the ratio is a function of the unit dose and duration of the pharmacological effect, in addition to being a function of the reinforcing capacity. In the case of depressants, using SPA as the standard, a high ratio was obtained with meperidine, codeine, and d-propoxyphene, but not with morphine, pentobarbital, or diazepam. A relatively high ratio, however, was obtained with the latter drugs when one of the weak depressants was used. We are seeking a standard drug that would be appropriate for this test on depressants.

In the continuous-self-administration test, a state of psychological dependence upon many principal drugs of abuse in man was demonstrated by persistent drugseeking behaviour, daily intake of high dose levels, and marked psychotoxic manifestation of drug effects (Deneau et al., 1969). Many principal drugs of abuse were actively self-administered by monkeys with the exception of hallucinogenic drugs and tetrahydrocannabinol. Marked withdrawal manifestations and increase of lever presses were observed with the narcotics, pentobarbital, and alcohol in this test when the drugs were withdrawn for 24 to 48 hours.

TABLE III

Physical-dependence tests on sedative hypnotics

 

Suppression of withdrawal

Physical dependence producing

Drug

signs

mg/kg

dose, a mg/kg x times day

weeks

withdrawal grade b

Barbital
+
(100 p.o.)
150 p.o. x 1
4
Severe
Phenobarbital
+
untested
100 p.o. x 1
6
Severe
Pentobarbital
+
(25 i.v.)
self, 5 x 59-85
2
Severe
Alcohol
+
(4,000 p.o.)
self, 1 g x 6.5
4
Severe
Meprobamate
+
(200 p.o.)
400 p.o. x 2
8
Severe
Diazepam
+
(5 p.o.)
20 p.o. x 1
8
Severe
Chlordiazepoxide
+
(20 p.o.)
113 p.o x 1
12
Severe
Chlorpromazine
-
       

( ) = Dose for complete suppression (mg/kg).

a Final dose.

b Graded by Yanagita's criteria (1970)

TABLE IV

Results of progressive-ratio test on CNS stimulants

 

Drugs and unit dose, mg/kg

Monkey No.

Cocaine 0.11

SPA 0.25

Methamphetamine 0.02

Amphetamine 0.03

243 6,400 1,600 800 400
  (16) (4) (2)  
248 3,200 1,600 400 400
  (8) (4) (1)  
281 3,200 3,200 1,600 800
  (4) (4) (2)  
334 3,200 1,600 800 400
  (8) (4) (2)  
Mean of ( )
9.0 4.0 1.8
1.0

( ) = Ratio against amphetamine in each monkey.

TABLE V

Results of progressive-ratio test on different unit doses of cocaine

 

Cocaine, unit dose, mg/kg

Monkey No.

Saline

0.03

0.12

0.48

243 800 400 6,400 6,400
  (2)   (16) (16)
248 400 800 3,200 6,400
 
(1/2)
  (4) (8)
281 200 800 6,400 12,800
 
(1/4)
  (8) (16)
334 200 1,600 3,200 12,800
 
(1/8)
  (2) (8)
Mean of ( )
0.72
1.0
7.5 12.0

() = Ratio against 0.03 in each monkey.

TABLE VI

The highest lever-press ratio for the final dose before extinction

Drug

Monkey No.

Dependent or its analogous state (A)

Non-dependent state (B)

A/B ratio

Morphine
174
1,600 a
1,600 1
  234 12,800
1,600 a
8
  248
12,800 a
6,400 2
  254 6,400
200 a
32
Cocaine
281 800
1,600 a
1/2
  334
3,200 a
6,400
1/2

a Tested first.

TABLE VII

Dependence liability of principal drugs

 

Reinforcement

Drugs

Physical dependence

Total

Secondary

Behavioural toxicity

Morphine
+ + +
+ + +
+ + +
+
Meperidine
+ + +
+ + +
(+ + +)
+
Codeine
+ +
+ + +
(+ +)
+
Pentazocine
+
+ +
?
+
Pentobarbital
+ +
+ + +
(+ +)
+ + +
Alcohol
+ +
+ +
(+)
+ + +
Diazepam
+ +
+
?
+
Cocaine
-
+ + +
-
+ + +
Amphetamines
-
+ +
(-)
+ + +
Nicotine
-
+
(-)
-
LSD
-
-
(-)
+ + a
THC
-
-
(-)
+ a

+ + + = very remarkable; + + = remarkable; + = positive; - = negative.

() = Predicted from continuous self-administration.

a By manual administration

In the progressive-ratio test in self-administration of some stimulants, the intensity of drug-seeking behaviour was found to be a function of the drug and the unit dose, as is shown in tables IV and V. These results may indicate the usefulness of the test for quantitative evaluation of the reinforcing effect of a drug. Table VI shows the results of the progressive ratio tests on morphine and cocaine. The tests were conducted when monkeys were in a physically dependent state or in a state analogous to physical dependency and also in a dependence-free state. An intensification of morphineseeking behaviour by development of physical dependence on the drug was demonstrated in three out of four monkeys, but no intensification was observable with cocaine. Thus, when a new drug is found to be liable to produce physical dependence, it may be worthwhile to explore the influence of physical dependence on the reinforcing effect of that drug.

Profiles of several major dependence-liable drugs, obtained by the above tests, are shown in table VII. It should be noted that the abuse potential of LSD, STP (DOM), mescaline, and THC is difficult to predict by these tests, since monkeys did not self-administer these drugs.

Bibliography

Deneau, G.A.; Yanagita, T., and Seevers, M.H.: Self-administration of psychoactive substances by the monkey. A measure of psychological dependence. Psychopharmacologia 16:30-48 (1969)

Jasinski, D.R.; Martin, W.R., and Hoeldtke, R.: Studies of the dependence producing properties of GPA 1657, profadol, and propiram in man. Clin. Pharmacol. Ther. 12: 613-649 (1971)

Seevers, M. H.: Opiate addiction in the monkey. I. Methods of study. J. Pharmacol. exp. Ther. 56: 147-156 (1936)

Seevers, M. H.; Deneau, G.A., and Yanagita, T.: Animal screening for drug dependence; in Knighton Pain, p. 197 (Little, Brown Boston 1966)

Yanagita T. and Takahashi S.: Development of tolerance to and physical dependence on barbiturates in rhesus monkeys. J. Pharmacol. exp. Ther. 172: 163-169 (1970).

Yanagita, T.; Takahashi, S. and Oinuma, N.: 4β-Methoxy-l-methyl-4α-phenyl-3α,5α-propanopiperidine hydrogen citrate (azabicyclane) tested in rhesus monkeys. Jap. J. clin. Pharmacol. 2: 126-134 (1971).