METABOLISM OF DRUGS IN OLD RATS (II) METABOLISM IN VIVO AND EFFECT OF DRUGS IN OLD RATS

METABOLISM METABOLISM

IN

OF DRUGS

VIVO

RYUICHI

AND

KATO

IN OLD RATS (II)

EFFECT AND AKIRA

OF

DRUGS

IN

OLD

RATS

TAKANAKA

Department of Pharmacology, National Institute of Hygienic Sciences,Setagaya-ku, Tokyo Received

for

publication

March

16,

1968

It is well known that the effects of some drugs in old animals quite differ from those in young animals. For example, Farner and Verzar (1) observed that the action of am phetamine to increase moter activity was more marked in young rats than in old rats and the action of hexobarbital to antagonized against the amphetamine action was more ef fective in old rats than in young rats. Moreover, the duration of hexobarbital anesthesia was longer in old rats than in young rats (2). Similarly, Petty and Karler (3) observed that anticonvulsive activity of acetazolamide and phenobarbital was more marked in old rats than in young rats. However, there is no study on the mechanism of such altered drug response in rela tion to the rate of drug metabolism. In a previous paper it was reported from our labo ratory that the activities in the oxidation and reduction of drugs and NADPH-linked elec tron transport system in liver microsomes of male and female rats were progressively decreased with aging (4). It is, therefore, of interest to investigate whether or not the metabolism in vivoof vari ous drugs is decreased in old rats in accordance with the decrease in the metabolic activi ties of liver microsomes observed in the in vitro experiments. Moreover, the relationship between the effects of drugs and the rate of in vivometabolism of drugs in the old rats was studied. MATERIALSAND METHODS Male and female rats of Wistar strain of about 40, 100, 300 and 600 days old were used . In most of experiments, 100 days old rats (young rats) and 600 days old rats (old rats) were used. Pentobarbital sodium, hexobarbital sodium, strychnine sulphate and OMPA (octa methylpyrophosphoramide) were dissolved in distilled water and carisoprodol was sus pended with 1% solution of sodium carboxymethylcellulose and all drugs were given intraperitoneally. The preparation of microsomes and the determinations of pentobarbital and cariso prodol oxidation were carried out according to the methods described in a previous paper (4). The metabolism in vivo of carisoprodol and pentobarbital were determined by the rate of disappearance of the drugs in the serum and brain after intraperitoneal injection . 力rl月 暴1籔 …

・ド石イ 中

正

The determinations of carisoprodol and pentobarbital were carried out according to the method of Kato et al. (5) and Brodie et al. (6). Carisoprodol and zoxazolamine paralysis and pentobarbital narcosis were determined by the duration of loss of the righting reflex. The toxicity of strychnine was evaluated by the convulsion and mortality. The toxicity of OMPA was evaluated by the time required for the onset of the toxic symptoms, convulsion and mortality in 50% of the treated rats. TABLE 1. Metabolism

in vitro

and

in vivo of

carisoprodol

in

young

and

old

male

rats.

RESULTS

1. Metabolismof carisoprodolin young and old rats Table 1 shows that the metabolism of carisoprodol by liver microsomes of the old male rats is markedly slower than in the young. Since the ratio of liver weight per 100 g body weight is demonstrated to be smaller in the old rats than in the young (4), the age difference is more evident in the activity of carisoprodol oxidase per 100 g body weight. The rate of decrease in the concentration of carisoprodol in the brain and serum in young and old male rats is given in Fig. 1. Since the rate of decrease in the concentration of carisoprodol was mainely regulated by the rate of metabolism in vivo, these results indicated that the rate of metabolism in vivo of carisoprodol was slower in the old rats than in the young. The biological half-life of carisoprodol in the young male rats was 97 minutes, while

Fir. 1. The rate of metabolism in vivo of cariso prodol in young and old male rats. Carisoprodol (200mg/kg) was given in traperitoneally. The rats were killed 30, 60. 120, 180 and 300 minutes after the administration and the serum (lower) and brain (upper) concentrations of cariso prodol were determined. The each value is average of 5 rats.

TABLE 2.

Metabolism

in vitro

and

in vivo of

carisoprodol

in

young

and

old

female

rats.

that in the old male rats was 199 minutes. Thus, the difference between the young and old rats in the activity of carisoprodol oxidase per 100 g body weight was similar to that in the rate of in vivo metabolism. Moreover, the metabolism of carisopro dol by liver microsomes in female rats was slower in old rats than in the young (Table 2). The rate of metabolism in vivoof cariso prodol in the young and old female rats is given in Fig. 2. The rate of metabolism in vivoof carisoprodol was faster in the young female rats than in the old female. 2. Metabolism of pentobarbital in young and old rats The rate of metabolism in vivo of pento barbital was slower in the old male rats than in the young male (Table 3). The difference between the young and old rats

FIG. 2.

The

prodol See TABLE 3.

Metabolism

in vitro

and

in vivo of

pentobarbital

rate in the in

of metabolism young legend

young

and for and

old Fig. old

in vivo of cariso female 1. male

rats.

rats.

TABLE 4.

Metabolism

TABLE 5.

in vitro

Duration

and

of

in vivo of

pentobarbital

pentobarbital

narcosis

in

in

young

young

and

and

old

old

female

rats

rats.

in the rate of metabolism in vivoof pentobarbital was similar to that in the activity of liver pentobarbital oxidase per 100 g body weight. Similarly, the metabolism in vitro and in vivo of pentobarbital was slower in the old female rats than in the young female (Table 4). 3. Duration of pentobarbitalnarcosisin young and old rats The duration of pentobarbital narcosis was longer in the old male rats than in the young male (Table 5). Similarly, a long duration of pentobarbital narcosis was observed in the old female rats than in the young, but the difference in the duration of pentobarbital narcosis between the young and old rats is more marked in the male rats than in the female. The difference in the duration of pentobarbital narcosis of the old male rats is 297% of the young rats, while in the female the difference is 93%. The discrepancy between the rate of in vivo metabolism and the duration of pentobarbital narcosis is probably due to the dose of pen tobarbital employed in the present investigation, and a small difference in the rate of the metabolism between the male and female rats may be exaggerated in the duration of action. Moreover, there was no clear difference in the brain concentration of pentobarbital on awaking. These results suggest that the difference in the duration of pentobarbital narcosis is likely not only due to the difference in the sensitivity of the central nervous

system to the drug, but it is rather due to the decreased rate of the drug metabolism in the old rats (7-10). 4. Duration of carisoprodiland zoxazolamineparalysis in young and old rats The duration of carisoprodol paralysis was longer in the old male rats than in the young male (Table 6). Similarly, the duration of carisoprodol paralysis was longer in the young female rats than in the old female, but the percentage of the prolongation in the duration of carisopro dol paralysis in the old rats is somewhat greater in the male rats than in the female . The duration of zoxazolamine paralysis was longer in the old male rats than in the young male (Table 7). In contrast to the duration of pentobarbital narcosis and cariso prodol paralysis, there was no clear sex difference in the duration of zoxazolamine paraly sis and the percentage prolongation in the zoxazolamine paralysis was almost same in the old male and female rats. TABLE 6.

Duration

young

and

of carisoprodol old

TABLE 8.

5.

paralysis

in

TABLE 7. in

rats.

Toxicity

of

strychnine

in

Duration young

young

of and

and

old

zoxazolamine old

paralysis

rats.

rats.

Toxicityof strychnineand OMPA in young and old rats The toxicity of strychnine was higher in the old male rats than in the young male

(Table 8). Similarly, the toxicity of strychnine was higher in the old female rats than in the young female, but the increase in the toxicity of strychnine in the old rats was somewhat greater in the male rats than in the female. In contrast, the toxicity of OMPA (octamethylpyrophosphoramide) was lower in the old male rats than in the young male (Table 9). The mortality of the young rats caused by 5 mg/kg and 10 mg/kg of OMPA was 40% and 100%, respectively, while killed none

TABLE 9.

Toxicity

of

OMPA

in

young

and

old

male

rats.

of the old rats by the administration of 10 mg/kg. These results are due to the fact that the anti-cholinesterase activity of OMPA is due to an active metabolite (hydroxylated OMPA) and the toxicity of OMPA is closely related to the activity of liver microsomal enzyme which activates OMPA (11, 12). 6. Effect of phenobarbitaltreatmenton the duration of hexobarbitalnarcosisin differentaged female rats In previous papers it was observed that the effects of phenobarbital to increase in the activities of microsomal drug-metabolizing enzymes and NADPH-electron transport system were decreased with the increase in age and in the old rats significant increase of the ac tivities by phenobarbital was not observed any more (13, 14). The effect of phenobarbital, therefore, on the duration of hexobarbital narcosis in different aged female rats was investigated. As shown in Fig. 3, the effect of pheno

FIG. 3. The effect of phenobarbital treatment on the duration of hexobarbital narcosis in different aged female rats. Phenobarbital (60 mg/kg) was given intraperitoneally for 3 days to the rats of 40, 100, 300 and 600 days old and hexobarbital (110mg/kg) was given intraperitoneally 24 hours after the last injection of phenobarbital. The each value is average +standard error of 6-12 rats.

barbital the

to decrease

increase

phenobarbital

the duration

in age and could

not

of hexobarbital

significant be observed

decrease

in the

narcosis duration

was markedly of hexobarbital

decreased narcosis

with by

in the old rats. DISCUSSION

In the previous paper (4), it was reported that the activities of drug-metabolizing enzymes and NADPH-linked electron transport system in liver microsomes were lower in old rats than in young rats. The results of the present investigation showed that the metabolism in vivo of cariso prodol and pentobarbital was slower in the old rats than in the young rats in accordance with the decreases in the activity of carisoprodol and pentobarbital oxidation by the liver microsomes (Tables 3 and 4). The differences in the activities of the microsomal enzymes became greater when the lower ratio of liver weight to body weight in the old animals was taken into consideration. The absolute value of the decrease in the metabolisms in vivo of carisoprodol and pentobarbital was greater in old male rats than in old female but there was no clear difference in the percentage of the decrease between male and female old rats. Thus the clear sex difference in the activity of carisoprodol and pentobarbital oxidation was still observed in the old rats. Since the activities of carisoprodol and pentobarbital oxidations are dependent on androgen (10), these results suggest that the androgen dependent regulation mechanism for drug-metabolizing enzymes of liver microsomes is still maintained in the old male rats (15-17). The durations of carisoprodol paralysis and pentobarbital narcosis were longer in the old rats than in the young, but the concentration of pentobarbital in the brain at the end of narcosis was almost same in the old and young rats. These results suggest that the difference in the rate of the metabolism of the drugs are likely a main factor for the determination of the difference in the duration of carisoprodol paralysis and pento barbital narcosis (7-10). Further supports for this view were obtained from the studies on the toxicities of strychnine and OMPA that the toxicity of strychnine was increased in the old rats, while the toxicity of OMPA was decreased. The toxicities of strychnine and OMPA are closely related to the activities of drug-metabolizing enzymes and when the enzyme activities are decreased the toxicity of strychnine is increased (8, 13), whereas the toxicity of OMPA is decreased (8, 11, 12). SUMMARY

1. The metabolism in vivoand effect of drugs in old male and female rats (600 days old) were studied in comparison with those in young male and female (100 days old). 2. The metabolisms in vivo of carisoprodol and pentobarbital were slower in the old rats than in the young rats in accordance with the decrease in the activities of carisoprodol and pentobarbital oxidations by the liver microsomes.

3. in the 4. the

durations

old rats The

toxicity 5.

the

The

than

in the

toxicity of OMPA

These

results

of metabolism

6.

The with

effect the

was

paralysis

and

pentobarbital

narcosis

were

longer

young.

of strychnine

rate

decreased

of carisoprodol

was higher

lower

indicate

that

in the

old

in the old rats than rats

than

the effect of drugs

in the

in the young,

whereas

young.

in the old rats is closely

related

to

of drugs. of

phenobarbital

aging

and

such

to effect

decrease could

the not

duration

be observed

of

hexobarbital

in the

was

old rats.

REFERENCES

1)

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2)

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3) 4)

PETTY, W.C. AND KARLER, R.: J. Pharmac. exp. Ther. 150, 443 (1965) KATO, R. ANDTAKANAKA,A.: Jap. J. Pharmac. 18, 381 (1968)

5) 6)

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7)

KATO, R. AND CHIESARA,E.: Brit. J. Pharmac. C'hemother.18, 29 (1962)

8)

KATO, R., CHIESARA,E. AND VASSANELLI, P.: Biochem.Pharmac. 11, 913 (1962)

9)

KATO, R.: Jap. J. Pharmac. 17, 499 (1967)

Ther. 109, 26 (1953)

10)

KATO, R., CHIESARA,E. AND FRONTINO,G.: Biochem. Pharmac. 11, 221 (1962)

11)

KATO, R.: Arzneim. Forsh. 11, 797 (1961)

12)

KATO, R., TAKANAKA,A. AND OMORI, Y.: Jap. J. Pharmac. 17, 509 (1967)

13)

KATO, R., CHIESARA,E. AND FRONTINO,G.: Experientia 17, 520 (1961)

14)

KATO, R. AND TAKANAKA,A.: J. Biochem. 63, 406 (1968)

15)

KATO, R. AND GILLETTE,J.R.:

J. Pharmac. exp. Ther. 150, 285 (1965)

16) KATO, R. AND TAKAHASHI,A.: Mol. Pharmac. 4, 109 (1968) 17)

KATO, R., TAKANAKA,A. ANDTAKAYANAGI,M.: Jap. J. Pharmac. (in press)