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Evaluation of Wistar rats' individual sensitivity to the development of physical dependence on morphine
Drug
und Alcohol Dependence,
Elsevier
Scientific
Publishers
69
29 (1991) 69 - 75 Ireland Ltd.
Evaluation of Wistar rats’ individual sensitivity to the development of physical dependence on morphine S.K. Sudakov, M.A. Konstantinopolsky,
L.A. Surkova, I.V. Tyurina,
E.V. Borisova
Laboratory of Narcotics Pharmacology and Experimental Therapy of Drug Abuse. All-Union Research Center on Biomedical Problems of Nurcology, Ministry of Health of the USSR, Mulyi Mogiltesevsky pereulok 3, 121291 Moscow (IJ.S.S.R) (Received
March
25th,
1991)
Behavioural and physiological indices which could be symptoms of individual Wistar rats’ sensitivity to the development of physical dependence on morphine, were studied. This state was induced in three different ways: (1) by i.p. injection of increasing doses of the drug; (2) by i.p. injection of morphine as a reward of a rat’s visit to a certain compartment of the Y-maze; (3) by voluntary drinking a morphine solution. Animals with high sensitivity to the development of dependence induced by the first way initially demonstrated intensive ambulation and low rearing in the open field and low nociception in the tail flick test. In the second case the ‘predisposed’ animals had initially higher ambulation also in comparison with the ‘resistant’ ones, but did not demonstrate significant withdrawal symptoms. The rats imbibing morphine solution exhibited a withdrawal syndrome, low level of initial nociception and received more electrocutaneous stimuli in the Vogel test, Thus, marked differences of sensitivity to the development of morphine genetically and neurochemically Key words: morphine;
physical
dependence determined. dependence;
are correlated
behaviour;
with some initial behavioural
predisposition;
It is known that both animals and human beings demonstrate different tendencies to drug consumption and development of dependence [ l-3 1 probably due to individual genetic patterns determining a peculiarity of the realization of these or those mechanisms of drug pharmacological effect. Different lines of mice respond differently to single or repeated morphine injections [ 4 I. Identification of neurochemical and molecular features of animals resistant to the development of drug dependence as distinct from those of predisposed animals might allow production of pharmacological preparations for pathogenetic therapy of drug dependence. Individual patterns of animals reflecting their genetic distinctions can be discovered by studying their behaviour and physiological, biochemical or pharmacological reactions. Thus, genetic Scientific
Publishers
indices
and may be
rats
lines of mice distinct in their sensitivity to morphine differ in a number of indices such as locomotor activity and nociception [ 5,6]. However, marked genetic distinctions between lines do not allow correlation of distinctions of one character with those in another. This paper deals with the individual comparison of the magnitude of Wistar rats’ nociception, locomotor activity and emotionality with the dynamics of morphine dependence developed in different ways.
Introduction
0376~8716/91/$03.50 0 1991 Elsevier Printed and Published in Ireland
and physiological
Methods The study comprised 140 male Wistar rats of 150-170 g at the start of the experiment. The behaviour of each animal was individually examined under four experimental conditions. (1) The intensity of ambulation and rearing and the number of grooming reactions were analyzed in Ireland
Ltd
70
single injection of morphine was given after 8 h followed by 1 mg/kg i.p. naloxone (Du Pont De Nemours, F.R.G.) to provoke the withdrawal syndrome. Fifteen minutes after naloxone administration animals were brought to the ‘open field’ for 3 min to record an alteration in common behavioural reactions and the presence of specific withdrawal ones. Alterations in ambulation and rearing, presence and recurrence of grooming, wet dog shakes, diarrhea, dyspnea, ptosis, piloerection, writhings, seizures, escape attempts, rhinorrhea, paw shakes, teeth chattering, posture disturbance were recorded [ 71. A total index of physical dependence severity was the arithmetical sum of all the aforementioned alterations (Table I). A group of 20 rats, tested in advance in compliance with the above-mentioned methods, was trained to be rewarded with a 5 mglkg morphine injection for a visit to a certain compartment of the Y-maze. Three times a day each rat was brought to the starting compartment and could freely travel to the right or left wing of the Ymaze. After a visit to the right, ‘stimulated’
the round ‘open field’ of 0.8 m diameter. (2) In the Vogel test (Lick Suppression Monitor, Coulbourn Instruments, U.S.A.) rats, after 72 h water deprivation, were offered water in a drinking tube for 5 min and every tenth licking was accompanied by a 0.5 mA electrocutaneous stimulus of 0.1 s duration. The number of electrocutaneous stimuli and the summed-up latent period of approaches to a drinking tube were calculated. (3) In the tail flick test the latent period of tail withdrawal from water of 56°C was measured. (4) The intensity of locomotor activity for 15 min was determined with the ‘Animex’ (LKB, Sweden). The number of fast and slow movements was separately calculated. After measurement of the above mentioned individual characteristics for each animal, morphine dependence was induced in 60 animals followed by an assessment of the withdrawal syndrome according to the previously proposed method in the ‘open field’ test [7]. The procedure consisted of i.p. injections of increasing morphine doses from 10 to 30 mg/kg twice a day with 8-h intervals during 5 days. On day 6 a
Table I.
Indices of the withdrawal symptoms. Their arithmetical
sum is an individual total index of the withdrawal syndrome.
Symptom
Index
Condition of its attribution
Ptosis Writhings Seizures Escape attempts ‘Wet dog’ shakes Teeth chattering Dyspnea Piloerection Posture disturbance Rhinorrhea Paw shakes Diarrhea Alterations of locomotor activity (ambulation)
1 1 1 1 1 1 2 2 2 2 3 3-9 0 1 2 3 4 5 6 0 1
If present at least once in 3 min
Alterations
of rearing
Depending on its severity (character of experiments) No change Complete inhibition of locomotor activity after 2.5 min 2.0 min 1.5 min 1.0 min 0.5 min at once Number of rearings during 3 min less than 12 Number of rearings during 3 min more than 11
compartment it was injected with morphine. In case of a visit to the left, ‘non-stimulated’ compartment no injection was given. On day 27 rats were brought to the ‘open field’ to record alterations in common behavioural reactions and presence of specific withdrawal symptoms according to the previously described methods. The third group of 60 rats tested in advance was deprived of water for 48 h and then during 16 days twice a day with an 8-h interval they were put for an hour into individual cages with drinking tubes containing morphine solution for free consumption. The morphine concentration was increased from 0.1 mg/ml (first day) to 0.45 mg/ml (eighth day). On day 10, besides the 0.45 mg/ml morphine solution, a 0.05 mg/ml quinine solution in another drinking tube was available and the morphine and quinine solution consumption under conditions of free choice was recorded during 6 days. On day 17 (within 24 h after the last morphine solution consumption) rats were brought to the ‘open field’ for 3 min to record changes in common behavioural reactions and the presence of specific withdrawal symptoms according to the above-mentioned methods. Findings of the preliminary testing were compared with the total index of physical dependence severity; the rate of habit formation to visit the Y-maze stimulated compartment; and with the amount of freely consumed morphine solution. Differences in the individual sensitivity of rats to the development of physical and psychological dependence were established with the Student criterion for small samples, Wilcoxon-Mann-Whitney criterion and the Fischer criterion as well as by regression analysis.
field’ was characteristic: a high locomotor activity of the majority of animals during the first 30 s of their stay in the ‘field’ was abruptly inhibited up to complete suppression during the following l-2 min. The total index, characterizing all common behavioural and specific reactions of the animals, ranged from 2 to 28 (average 17.4). According to the magnitude of the total index, animals were subdivided into three groups. (1) Sensitive to the development of morphine dependence (total index magnitude exceeded 20). (2) Resistant to the development of dependence (total index magnitude was below 5). (3) The intermediate group. A comparison of the first and second group was most interesting. The comparison of the withdrawal syndrome total index with the findings of preliminary testing showed that animals with a high sensitivity to the development of physical dependence initially demonstrated an intensive ambulation (308 f 30.5 units) and low rearing (16 f 2.1 units) in the ‘open field’ and had low nociception (latent period of tail withdrawal was 8.3 + 0.3 s). Resistant animals demonstrated a high ambulation (276 f 25 units) and rearing (22.5 f 3.7 units) with insignificant grooming activity; nociception was raised (latent period 6.3 * 0.2 s). In comparison with resistant
Results Individual peculiarities of the severity physical dependence following administration increasing morphine doses
of of
After the development of dependence rats demonstrated different degrees of morphine withdrawal symptoms. Thus, the process of alteration in locomotor activity in the ‘open
Fig. 1. Initial number (in %) of electrocutaneous stimuli in the Vogel test to rats predisposed and resistant to the dependence development in case of i,p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution. Average number of electrocutaneous stimuli received in the Vogel test in each group was accepted as 100%.
animals rats sensitive to the development of morphine physical dependence received almost three times less electrocutaneous stimuli (P < 0.001) in the Vogel test (Fig. 1) and the total latent period of approaches to a drinking tube was longer (P < 0.01). Findings of the initial locomotor activity did not correlate with the magnitude of the withdrawal syndrome total index. Individual peculiarities of dependence following morphine injections after a voluntary visit to a certain compartment of the Y-maze
When putting rats during 14 days in the maze they visited the right compartment in 31.5% of cases on average. As in the previous experiment animals were subdivided into three groups depending on the frequency of visits to the stimulated compartment. (1) Rats most often visiting the stimulated compartment (‘predisposed’ to the dependence development). (2) Rats extremely seldom or
never visiting the right compartment (‘resistant’ animals); the average latent period of a visit of these groups of animals to the stimulated compartment differed significantly and was 39.0 f 8 s and 103.0 + 12 s, respectively. (3) Animals with intermediate values of the number and latent period of visits to the right compartment. The comparison of results obtained in these groups with findings of the preliminary testing shows that ‘predisposed’ rats initially exhibited higher ambulation in the ‘open field’ in comparison with ‘resistant’ ones (184.5 f 16.8 units and 153.4 f 25.9 units, respectively) as well as lower nociception (latent period of tail withdrawal was 6.2 f 1.6 s and 2.6 + 0.7 s, respectively). Measurement of physical dependence symptoms in the ‘open field’ after the termination of morphine-administration experiments did not reveal significant symptoms in any experimental group.
--e
M
-is-
M+Q
-
Q
6
0
I
I
I
I
1
I
I
I
I
I
1
I
I
I
2
3
4
5
6
7
a
9
10
11
12
13
days
Fig. 2. Amount of morphine consumption (mg) by rats giving preference to morphine solution (M), quinine solution (Q) and by the intermediate group (M + Q). Ordinate - amount of morphine daily consumption. Abscissa - days of the experiment (on the 10th day rats had free access to quinine solution also).
Individual peculiarities of dependence following voluntary consumption of morphine During the first 10 days, when rats could satify their water need by drinking morphine solution in increasing concentrations, individual differences in consumption were not very significant. However, even then a small population of animals (5%) consumed morphine solution in amounts evidently insufficient to keep a normal fluid and electrolyte balance. At the same time another 7% of rats consumed large amounts of morphine solution extremely exceeding the average norms of water consumption. After providing free access to morphine and quinine solutions a sharp division of all animals into 3 groups took place; 43.3% of rats preferred to consume morphine solution; 28.3% gave preference to quinine solution and 28.3% of animals consumed both solutions with equal intensity. The amount of daily morphine consumption by these groups is illustrated in Fig. 2. The majority of rats giving preference to morphine demonstrated a withdrawal syndrome (total index was 11.4 units) during the following measurement of the expression of physical dependence in the ‘open field’ test. At the same time rats preferring quinine consumption or consuming approximately equal amounts of morphine and quinine solutions did not demonstrate withdrawal symptoms. Rats giving preference to morphine solution were possessed of the lowest nociception (latent period of tail withdrawal 4.16 f 0.36 s) in comparison with rats consuming quinine or with the intermediate group (3.43 f 0.3 s and 4.06 f 0.27 s respectively). Besides, rats with morphine preference and the intermediate group received considerably more electrocutaneous stimuli (32.82 SE4.89 and 32.07 f 5.46 respectively) in comparison with rats not consuming morphine (19.53 f 5.3). Discussion
From the results it can be inferred that the population of Wistar rats comprises animals with different responses to morphine administration. Some of the rats are characterized
Fig. 3. Intensity (in ‘70) of the initial rearing in rats predisposed and resistant to the dependence development in case of i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution. Average intensity of rearing in each group was accepted as 100%.
by a rapid development of psychological and physical dependence while other animals are completely resistant to the development of dependence. They also differ in some other physiological indices. In agreement with results of the three experimental series in which morphine dependence was induced in different ways, the highest predisposition to dependence development was observed in animals with a lowered rearing and low nociception (Figs. 3 and 4). However, predisposed rats whose dependence was induced by drinking morphine solution or in the Y-maze in distinction from predisposed animals whose dependence was induced involuntarily, had initially demonstrated lower ambulation in comparison with resistant rats (Fig. 5). This could be due to the experimental conditions when the animals were to manifest locomotor activity in order to receive the drug. It is known that effects of single or repeated administrations of morphine develop differently in different lines of mice. It was shown that the C57 line of mice initially characterized by low locomotor activity increased it after a single morphine administration and rapidly developed tolerance to morphine in the test of increasing locomotor activity, their physical dependence on
Initial duration (in W) of the latent period of tail Fig. 4. flit in rats predisposed and resistant to the dependence development in case of i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution (in W). Average duration of the latent period of tail flick in each group was accepted as 100%.
morphine developed more rapidly and the withdrawal syndrome was more severe [4-61. If, as has been suggested 18-10 1 the brain monoamine system is involved in the locomotor stimulating effect of morphine, the different sensitivity to morphine of specific line animals may be explained by different levels of catecholamine metabolizing enzymes [ 111. It is possible that a particular brain catecholamine metabolism of our rats was responsible also for
their different behaviour in the ‘open field’. Since brain monoamines are involved in the development of dependence [ 121 such metabolic differences could also explain the observed differences in the development of physical dependence on morphine. The initial low nociception of predisposed animsls may indicate an intensified activity of their endogenous opioid system [ 131. Indeed, monkeys prone to developing dependence on morphine had an increased content of preceptors in the nucleus caudatus and globus pallidus and &receptors in the thalamus [ 141. Thus, it may be suggested that genetically determined differences in the monoamine and endogenous opioid systems probably participating in the reinforcement mechanisms can determine the individual sensitivity of an animal to the development of physical dependence on morphine regardless of how it is induced. On the other hand, animals more susceptible to the dependence development after involuntary, i.e. i.p. morphine administration as opposed to animals more actively developing dependence by voluntary drinking of morphine solution, are possessed of more emotionality as measured with the Vogel test. This fact shows that some genetic peculiarities of animals can contribute to or prevent the development of dependence, depending on the mode of drug consumption. Thus, the results of the study in its present stage allow to conclude that there are marked differences in animal sensitivity to the development of morphine dependence. These differences correlate with certain behavioural and physiological indices which, in their turn, are connected with genetic and neurochemical peculiarities of the systems providing them. References
Fig. 5. Intensity (in %) of the initial ambulation in rats predisposed and resistant to the dependence development in case if i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze), and of drinking morphine solution. Average intensity of ambulation in each group was accepted as 100%.
Kolb, L. and Himmelsbach, C.K. Am. J. Psychiat. (1938) 94, 759-767. Nichols, J.R. and Hsiao, S. Science (1967) 157, 561-563. Maiski, AI., Vedernikova, N.N., Chistiakov, V.V. and Lakin, V.V. Biohimicheskie aapekty narkomanii. M. Medicina, 1982.
75
4 5 6 7
8 9
Ericson, K. and Kiianmaa, K. Ann. Med. Exp. Fenn., (1971) 49, 73-78. Olivero, A. and Castellano, C. Psychopharmacol. (1974) 39, 13-Z. Kuribara, H. and Tadokoro, S. Japan J. Pharmacol. (1989) 49, 197-203. Konstantinopolsky, M.A.. Tiurina, I.V. and Surkova. L.A. Medikobiologicheskie problemy alkogolizma. M. 1988, p. 56-60. Rethy, C.R., Smith, C.B. and Villarreal, J.E. J. Pharmacol. Exp. Ther., (1971) 1’76, 472-479. Buxbaum, D.M., Yarbough, G.G. and Garter, M.M. J. Pharmacol. Exp. Ther., (1973) 185, 317-327.
10 11 12 13 14
Kuschinsky, K. and Hornykiewicz, 0. Eur. J. Pharmacol. (1974) 26, 41-50. Eleftheriou, B.E. Neuroendocrinology (1971) 7, 329-336. Anokhina, I.P., Kogan, B.M. and Hristoliubova, J. nevropatologii i psihiatrii, (1979) 79, 751-758. Ollat, H., Parvez, S. and Parvez, H. Biogenic amines (1989) 6, 381-410. Neil, A., Terenius, L. and Ternes, I.W. Eur. J. Pharmacol (1986) 122, 143-147.
und Alcohol Dependence,
Elsevier
Scientific
Publishers
69
29 (1991) 69 - 75 Ireland Ltd.
Evaluation of Wistar rats’ individual sensitivity to the development of physical dependence on morphine S.K. Sudakov, M.A. Konstantinopolsky,
L.A. Surkova, I.V. Tyurina,
E.V. Borisova
Laboratory of Narcotics Pharmacology and Experimental Therapy of Drug Abuse. All-Union Research Center on Biomedical Problems of Nurcology, Ministry of Health of the USSR, Mulyi Mogiltesevsky pereulok 3, 121291 Moscow (IJ.S.S.R) (Received
March
25th,
1991)
Behavioural and physiological indices which could be symptoms of individual Wistar rats’ sensitivity to the development of physical dependence on morphine, were studied. This state was induced in three different ways: (1) by i.p. injection of increasing doses of the drug; (2) by i.p. injection of morphine as a reward of a rat’s visit to a certain compartment of the Y-maze; (3) by voluntary drinking a morphine solution. Animals with high sensitivity to the development of dependence induced by the first way initially demonstrated intensive ambulation and low rearing in the open field and low nociception in the tail flick test. In the second case the ‘predisposed’ animals had initially higher ambulation also in comparison with the ‘resistant’ ones, but did not demonstrate significant withdrawal symptoms. The rats imbibing morphine solution exhibited a withdrawal syndrome, low level of initial nociception and received more electrocutaneous stimuli in the Vogel test, Thus, marked differences of sensitivity to the development of morphine genetically and neurochemically Key words: morphine;
physical
dependence determined. dependence;
are correlated
behaviour;
with some initial behavioural
predisposition;
It is known that both animals and human beings demonstrate different tendencies to drug consumption and development of dependence [ l-3 1 probably due to individual genetic patterns determining a peculiarity of the realization of these or those mechanisms of drug pharmacological effect. Different lines of mice respond differently to single or repeated morphine injections [ 4 I. Identification of neurochemical and molecular features of animals resistant to the development of drug dependence as distinct from those of predisposed animals might allow production of pharmacological preparations for pathogenetic therapy of drug dependence. Individual patterns of animals reflecting their genetic distinctions can be discovered by studying their behaviour and physiological, biochemical or pharmacological reactions. Thus, genetic Scientific
Publishers
indices
and may be
rats
lines of mice distinct in their sensitivity to morphine differ in a number of indices such as locomotor activity and nociception [ 5,6]. However, marked genetic distinctions between lines do not allow correlation of distinctions of one character with those in another. This paper deals with the individual comparison of the magnitude of Wistar rats’ nociception, locomotor activity and emotionality with the dynamics of morphine dependence developed in different ways.
Introduction
0376~8716/91/$03.50 0 1991 Elsevier Printed and Published in Ireland
and physiological
Methods The study comprised 140 male Wistar rats of 150-170 g at the start of the experiment. The behaviour of each animal was individually examined under four experimental conditions. (1) The intensity of ambulation and rearing and the number of grooming reactions were analyzed in Ireland
Ltd
70
single injection of morphine was given after 8 h followed by 1 mg/kg i.p. naloxone (Du Pont De Nemours, F.R.G.) to provoke the withdrawal syndrome. Fifteen minutes after naloxone administration animals were brought to the ‘open field’ for 3 min to record an alteration in common behavioural reactions and the presence of specific withdrawal ones. Alterations in ambulation and rearing, presence and recurrence of grooming, wet dog shakes, diarrhea, dyspnea, ptosis, piloerection, writhings, seizures, escape attempts, rhinorrhea, paw shakes, teeth chattering, posture disturbance were recorded [ 71. A total index of physical dependence severity was the arithmetical sum of all the aforementioned alterations (Table I). A group of 20 rats, tested in advance in compliance with the above-mentioned methods, was trained to be rewarded with a 5 mglkg morphine injection for a visit to a certain compartment of the Y-maze. Three times a day each rat was brought to the starting compartment and could freely travel to the right or left wing of the Ymaze. After a visit to the right, ‘stimulated’
the round ‘open field’ of 0.8 m diameter. (2) In the Vogel test (Lick Suppression Monitor, Coulbourn Instruments, U.S.A.) rats, after 72 h water deprivation, were offered water in a drinking tube for 5 min and every tenth licking was accompanied by a 0.5 mA electrocutaneous stimulus of 0.1 s duration. The number of electrocutaneous stimuli and the summed-up latent period of approaches to a drinking tube were calculated. (3) In the tail flick test the latent period of tail withdrawal from water of 56°C was measured. (4) The intensity of locomotor activity for 15 min was determined with the ‘Animex’ (LKB, Sweden). The number of fast and slow movements was separately calculated. After measurement of the above mentioned individual characteristics for each animal, morphine dependence was induced in 60 animals followed by an assessment of the withdrawal syndrome according to the previously proposed method in the ‘open field’ test [7]. The procedure consisted of i.p. injections of increasing morphine doses from 10 to 30 mg/kg twice a day with 8-h intervals during 5 days. On day 6 a
Table I.
Indices of the withdrawal symptoms. Their arithmetical
sum is an individual total index of the withdrawal syndrome.
Symptom
Index
Condition of its attribution
Ptosis Writhings Seizures Escape attempts ‘Wet dog’ shakes Teeth chattering Dyspnea Piloerection Posture disturbance Rhinorrhea Paw shakes Diarrhea Alterations of locomotor activity (ambulation)
1 1 1 1 1 1 2 2 2 2 3 3-9 0 1 2 3 4 5 6 0 1
If present at least once in 3 min
Alterations
of rearing
Depending on its severity (character of experiments) No change Complete inhibition of locomotor activity after 2.5 min 2.0 min 1.5 min 1.0 min 0.5 min at once Number of rearings during 3 min less than 12 Number of rearings during 3 min more than 11
compartment it was injected with morphine. In case of a visit to the left, ‘non-stimulated’ compartment no injection was given. On day 27 rats were brought to the ‘open field’ to record alterations in common behavioural reactions and presence of specific withdrawal symptoms according to the previously described methods. The third group of 60 rats tested in advance was deprived of water for 48 h and then during 16 days twice a day with an 8-h interval they were put for an hour into individual cages with drinking tubes containing morphine solution for free consumption. The morphine concentration was increased from 0.1 mg/ml (first day) to 0.45 mg/ml (eighth day). On day 10, besides the 0.45 mg/ml morphine solution, a 0.05 mg/ml quinine solution in another drinking tube was available and the morphine and quinine solution consumption under conditions of free choice was recorded during 6 days. On day 17 (within 24 h after the last morphine solution consumption) rats were brought to the ‘open field’ for 3 min to record changes in common behavioural reactions and the presence of specific withdrawal symptoms according to the above-mentioned methods. Findings of the preliminary testing were compared with the total index of physical dependence severity; the rate of habit formation to visit the Y-maze stimulated compartment; and with the amount of freely consumed morphine solution. Differences in the individual sensitivity of rats to the development of physical and psychological dependence were established with the Student criterion for small samples, Wilcoxon-Mann-Whitney criterion and the Fischer criterion as well as by regression analysis.
field’ was characteristic: a high locomotor activity of the majority of animals during the first 30 s of their stay in the ‘field’ was abruptly inhibited up to complete suppression during the following l-2 min. The total index, characterizing all common behavioural and specific reactions of the animals, ranged from 2 to 28 (average 17.4). According to the magnitude of the total index, animals were subdivided into three groups. (1) Sensitive to the development of morphine dependence (total index magnitude exceeded 20). (2) Resistant to the development of dependence (total index magnitude was below 5). (3) The intermediate group. A comparison of the first and second group was most interesting. The comparison of the withdrawal syndrome total index with the findings of preliminary testing showed that animals with a high sensitivity to the development of physical dependence initially demonstrated an intensive ambulation (308 f 30.5 units) and low rearing (16 f 2.1 units) in the ‘open field’ and had low nociception (latent period of tail withdrawal was 8.3 + 0.3 s). Resistant animals demonstrated a high ambulation (276 f 25 units) and rearing (22.5 f 3.7 units) with insignificant grooming activity; nociception was raised (latent period 6.3 * 0.2 s). In comparison with resistant
Results Individual peculiarities of the severity physical dependence following administration increasing morphine doses
of of
After the development of dependence rats demonstrated different degrees of morphine withdrawal symptoms. Thus, the process of alteration in locomotor activity in the ‘open
Fig. 1. Initial number (in %) of electrocutaneous stimuli in the Vogel test to rats predisposed and resistant to the dependence development in case of i,p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution. Average number of electrocutaneous stimuli received in the Vogel test in each group was accepted as 100%.
animals rats sensitive to the development of morphine physical dependence received almost three times less electrocutaneous stimuli (P < 0.001) in the Vogel test (Fig. 1) and the total latent period of approaches to a drinking tube was longer (P < 0.01). Findings of the initial locomotor activity did not correlate with the magnitude of the withdrawal syndrome total index. Individual peculiarities of dependence following morphine injections after a voluntary visit to a certain compartment of the Y-maze
When putting rats during 14 days in the maze they visited the right compartment in 31.5% of cases on average. As in the previous experiment animals were subdivided into three groups depending on the frequency of visits to the stimulated compartment. (1) Rats most often visiting the stimulated compartment (‘predisposed’ to the dependence development). (2) Rats extremely seldom or
never visiting the right compartment (‘resistant’ animals); the average latent period of a visit of these groups of animals to the stimulated compartment differed significantly and was 39.0 f 8 s and 103.0 + 12 s, respectively. (3) Animals with intermediate values of the number and latent period of visits to the right compartment. The comparison of results obtained in these groups with findings of the preliminary testing shows that ‘predisposed’ rats initially exhibited higher ambulation in the ‘open field’ in comparison with ‘resistant’ ones (184.5 f 16.8 units and 153.4 f 25.9 units, respectively) as well as lower nociception (latent period of tail withdrawal was 6.2 f 1.6 s and 2.6 + 0.7 s, respectively). Measurement of physical dependence symptoms in the ‘open field’ after the termination of morphine-administration experiments did not reveal significant symptoms in any experimental group.
--e
M
-is-
M+Q
-
Q
6
0
I
I
I
I
1
I
I
I
I
I
1
I
I
I
2
3
4
5
6
7
a
9
10
11
12
13
days
Fig. 2. Amount of morphine consumption (mg) by rats giving preference to morphine solution (M), quinine solution (Q) and by the intermediate group (M + Q). Ordinate - amount of morphine daily consumption. Abscissa - days of the experiment (on the 10th day rats had free access to quinine solution also).
Individual peculiarities of dependence following voluntary consumption of morphine During the first 10 days, when rats could satify their water need by drinking morphine solution in increasing concentrations, individual differences in consumption were not very significant. However, even then a small population of animals (5%) consumed morphine solution in amounts evidently insufficient to keep a normal fluid and electrolyte balance. At the same time another 7% of rats consumed large amounts of morphine solution extremely exceeding the average norms of water consumption. After providing free access to morphine and quinine solutions a sharp division of all animals into 3 groups took place; 43.3% of rats preferred to consume morphine solution; 28.3% gave preference to quinine solution and 28.3% of animals consumed both solutions with equal intensity. The amount of daily morphine consumption by these groups is illustrated in Fig. 2. The majority of rats giving preference to morphine demonstrated a withdrawal syndrome (total index was 11.4 units) during the following measurement of the expression of physical dependence in the ‘open field’ test. At the same time rats preferring quinine consumption or consuming approximately equal amounts of morphine and quinine solutions did not demonstrate withdrawal symptoms. Rats giving preference to morphine solution were possessed of the lowest nociception (latent period of tail withdrawal 4.16 f 0.36 s) in comparison with rats consuming quinine or with the intermediate group (3.43 f 0.3 s and 4.06 f 0.27 s respectively). Besides, rats with morphine preference and the intermediate group received considerably more electrocutaneous stimuli (32.82 SE4.89 and 32.07 f 5.46 respectively) in comparison with rats not consuming morphine (19.53 f 5.3). Discussion
From the results it can be inferred that the population of Wistar rats comprises animals with different responses to morphine administration. Some of the rats are characterized
Fig. 3. Intensity (in ‘70) of the initial rearing in rats predisposed and resistant to the dependence development in case of i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution. Average intensity of rearing in each group was accepted as 100%.
by a rapid development of psychological and physical dependence while other animals are completely resistant to the development of dependence. They also differ in some other physiological indices. In agreement with results of the three experimental series in which morphine dependence was induced in different ways, the highest predisposition to dependence development was observed in animals with a lowered rearing and low nociception (Figs. 3 and 4). However, predisposed rats whose dependence was induced by drinking morphine solution or in the Y-maze in distinction from predisposed animals whose dependence was induced involuntarily, had initially demonstrated lower ambulation in comparison with resistant rats (Fig. 5). This could be due to the experimental conditions when the animals were to manifest locomotor activity in order to receive the drug. It is known that effects of single or repeated administrations of morphine develop differently in different lines of mice. It was shown that the C57 line of mice initially characterized by low locomotor activity increased it after a single morphine administration and rapidly developed tolerance to morphine in the test of increasing locomotor activity, their physical dependence on
Initial duration (in W) of the latent period of tail Fig. 4. flit in rats predisposed and resistant to the dependence development in case of i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze) and of drinking morphine solution (in W). Average duration of the latent period of tail flick in each group was accepted as 100%.
morphine developed more rapidly and the withdrawal syndrome was more severe [4-61. If, as has been suggested 18-10 1 the brain monoamine system is involved in the locomotor stimulating effect of morphine, the different sensitivity to morphine of specific line animals may be explained by different levels of catecholamine metabolizing enzymes [ 111. It is possible that a particular brain catecholamine metabolism of our rats was responsible also for
their different behaviour in the ‘open field’. Since brain monoamines are involved in the development of dependence [ 121 such metabolic differences could also explain the observed differences in the development of physical dependence on morphine. The initial low nociception of predisposed animsls may indicate an intensified activity of their endogenous opioid system [ 131. Indeed, monkeys prone to developing dependence on morphine had an increased content of preceptors in the nucleus caudatus and globus pallidus and &receptors in the thalamus [ 141. Thus, it may be suggested that genetically determined differences in the monoamine and endogenous opioid systems probably participating in the reinforcement mechanisms can determine the individual sensitivity of an animal to the development of physical dependence on morphine regardless of how it is induced. On the other hand, animals more susceptible to the dependence development after involuntary, i.e. i.p. morphine administration as opposed to animals more actively developing dependence by voluntary drinking of morphine solution, are possessed of more emotionality as measured with the Vogel test. This fact shows that some genetic peculiarities of animals can contribute to or prevent the development of dependence, depending on the mode of drug consumption. Thus, the results of the study in its present stage allow to conclude that there are marked differences in animal sensitivity to the development of morphine dependence. These differences correlate with certain behavioural and physiological indices which, in their turn, are connected with genetic and neurochemical peculiarities of the systems providing them. References
Fig. 5. Intensity (in %) of the initial ambulation in rats predisposed and resistant to the dependence development in case if i.p. morphine injections in growing doses (withdrawal), of morphine administration in the stimulated compartment (Y-maze), and of drinking morphine solution. Average intensity of ambulation in each group was accepted as 100%.
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