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Hypnotic activity of N3-benzylthymidine on mice
Life Sciences, Vol. 41, pp. 2791-2797 Printed in the U.S.A.
Pergamon Journal~
HYPNOTIC ACTIVITY OF N3-BENZYLTHYMIDINE ON MICE Ikuo Yamamoto,f Toshiyuki Kimur~ Yuji Tateoka, Kazuhito Watanabe and Ing Kang Ho* Department of Hygienic Chemistry, School of Pharmacy, Hokuriku University, 3-Ho, Kanagawa-machi,Kanazawa 920-11, Japan and * Department of Pharmacology and Toxicology, The University of Mississippi Medical Center, Jackson, Mississippi 39216, U.S.A. (Received in final form October 26, 1987)
Summary The pharmacological effect in mice of N3-benzylthymidine (N3ByTd) was examined by two routes of administration; intravenous ( i . v . ) and intracerebroventricular ( i . c . v . ) , and compared with the effect of administration of N3-benzyluridine (N3-ByUd) previously reported. Hypnotic a c t i v i t y , pentobarbital (PB)-induced sleeping time, motor incoordination and spontaneous a c t i v i t y were used as indices of pharmacological effects. N3-ByTd (0.5-2.0 umol/mou~e, i.c.v.) and N3-ByUd (1.5-3.0 umol/mouse, i . c . v . ) were found to possess dose-dependent hypnotic a c t i v i t y , and N3-ByTd had more potent hypnotic a c t i v i t y than N3-ByUd. Both N3-ByTd and N3-ByUd (0.5 and 1.0 ~mol/mouse, i . c . v . , respectively) showed a synergistic effect on PB-induced sleep, although t h e i r parent compounds, thymidine (Td) and uridine (Ud), did not potentiate the a c t i v i t y at each dose. In motor incoordination, the effect of N3-ByTd (0.5 umol/mouse) continued for 6 hr after i . c . v , injection. All compounds decreased the spontaneous activity of mice by i . c . v , administration. Furthermore, both N3-ByTd and N3-ByUd decreased the a c t i v i t y , when they were administered by i . v . These results reveal that both N3benzylpyrimidine nucleosides have more direct depressant effects on the central nervous system (CNS) than the parent compounds. Among the pyrimidine nucleoside derivatives tested, N3-ByTd was found to be the most potent hypnotic substance. Recently, we reported that N3-ByUd exerts hypnotic a c t i v i t y by i . c . v . administration in mice ( I ) , and that N3-allyluridine and N3-allylthymidine also have CNS depressant effects, although these compounds do not cause hypnotic a c t i v i t y in mice (2, 3). We thus proposed that N3-ByUd is a new type of hypnotic substance because of a characteristic sugar moiety at the N position in i t s structure. To our knowledge, there is no report with regard to hypnotic a c t i v i t y by pyrimidine nucleosides and their analogues. Ud is known as one of the sleep-promoting substances which have been extracted from brainstem of sleep-deprived rats (4-6). Krooth et al. (7) have reported that Ud, Td and other natural pyrimidines decreased spontaneous f To whom all
correspondence should be addressed
0024-3205/87 $3.00 + .00 Copyright (c) 1987 Pergamon Journals Ltd.
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Hypnotic Activity of N3-ByTd
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a c t i v i t y at high doses in mice. In t h i s case, they indicate that Td is more potent than Ud. Therefore, i t is suggested that the introduction of a benzyl to the N3 position of Td may cause stronger hypnotic effects than N3-ByUd. The present study was thus designed to compare the hypnotic and depressant a c t i v i t y of N3-ByTd and Td with N3-ByUd and Ud. Materials and Methods Animals: Male ddN s t r a i n mice weighing 25 ± 3 g were used. They were kept in controlled l i g h t i n g (7:00 to 19:00 l i g h t period), and ambient temperature (23 ± I°C) throughout the experiments. T h e y were given food and water ad libitum. Chemicals: Td and Ud were purchased from Kohjin Co., Ltd. Tokyo and Wako Pure Chemical Ind., Ltd. Osaka, r e s p e c t i v e l y . Pentobarbital sodium was obtained from Tokyo Kasei Co., Ltd. Tokyo. Preparation of N3-Benzylated Compounds: N3-ByTd and N3-ByUd were prepared according to the method of Sasaki et a l . (8). Drug Administration: N3-ByTd, Td, N3-ByUd and Ud were suspended in 3% Tween80-saline solution because of i n s o l u b i l i t y of N3-benzylated compounds in saline. PB was injected i n t r a p e r i t o n e a l l y ( i . p . ) at the dose of 40 mg/kg. The i . c . v , i n j e c t i o n was performed according to the method of Haley and McCormick (9). In spontaneous a c t i v i t y t e s t , i . v . administration was also used in mice via t a i l vein. Pharmacological Effects: Sleeping time, as hypnotic a c t i v i t y , was measured as the time between the loss and recovery of r i g h t i n g r e f l e x in mice. This endpoint was also used for the PB-induced sleeping time. In the dose response experiments, mice were given compounds, by i . c . v , administration, at the dose of 0.25-3.0 ~mol/mouse. PB-induced sleeping time was measured a f t e r i . p . i n j e c t i o n of sodium PB. The tested compound was usually given by i . c . v . administration 15 min before the PB application. For time-course studies, a l l tested compounds were administered i . c . v . 5, 15 and 45 min p r i o r to the i n j e c t i o n of PB. Throughout PB-induced sleeping e f f e c t , motor incoordination and spontaneous a c t i v i t y t e s t s , 0.5 umol/mouse of N3-ByTd and Td was injected by i . c . v , administration, while 1.0 umol/mouse of N3-ByUd and Ud was injected. Each dose was decided by preliminary t e s t s . The dose of 0.5 umol/mouse N3-ByTd did not show any overt CNS t o x i c i t y . Effect of motor incoordination was measured by a bar t e s t . Compounds were injected, then mice were put on a bar (I cm square, 40 cm height). Motor incoordination a c t i v i t y of compounds tested was expressed as the percent of mice that f e l l from the bar within 30 seconds. Spontaneous a c t i v i t y was examined using a Digiscan A c t i v i t y Monitor, coupled with a Digiscan Computer (Omnitech Electronics Inc., U.S.A.) a f t e r both i . v . and i . c . v , administration, and the treated mice were placed in the center of a square p l a s t i c box with 30 cm sides. Measurement of a c t i v i t y was evaluated as the t o t a l distance (inches) of movement within I hr a f t e r injection. Multiple t-test (ANOVA) was u s e d for calculation of s t a t i s t i c a l significance. Results Hypnotic A c t i v i t y : Sleeping times of animals by i . c . v , administration are shown in Fig. possessed hypnotic a c t i v i t y at the doses of ~mol/mouse,respectively. N3-ByUd f a i l e d to
treated with N3-ByTd and N3-ByUd I. Both N3-ByTd and N3-ByUd 0.5-2.0 ~mol/mouse and 1.5-3.0 induce hypnotic a c t i v i t y at 1.0
Vol. 41, No. 26, 1987
Hypnotic Activity of N3-ByTd
0
2793
0
CH3
.oc OH OH
OH
NS-ByUd
N3-ByTd
100 t-
E= E
5C r Q.
u~
1 2 Dose ( p moll mouse)
3
Fig. 1 Hypnotic a c t i v i t y of N3-ByTd and N3-ByUd on mice. N3-ByTd (•) and N3 -ByUd ( • ) were administered by i . c . v . administration. Each point corresponds to the mean sleeping time ± S.E. Seven or 8 mice were used in each group.
~mol/mouse. Sleeping time of of N3-ByUd at 2.0 ~mol/mouse.
N3-ByTd was longer (about two-fold)
than that
PB-Induced Sleep: Table I summarizes the time course e f f e c t of p r i o r treatment of test compounds on PB-induced sleeping time. N3-ByTd (0.5 ~mol/mouse, i . c . v . ) potentiated the PB-induced sleeping time s i g n i f i c a n t l y at 45 min a f t e r the treatment. Td administration did not produce a s i g n i f i c a n t difference at any time interval as compared to control. NS-ByUd, at all time points, showed s i g n i f i c a n t p o t e n t i a t i o n , but i t s parent compound, Ud, did not change the sleep e f f e c t as well as Td.
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Hypnotic Activity of N3-ByTd
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Table I Time course of the effects of N3-ByTd, N3-ByUd and t h e i r parent compounds on PB-induced slee p Compound
Sleepin 9 time (min) Time i n t e r v a l p r i o r to PB administration 5 15 45 (min)
Control (3% TweenSO-saline)
52 ± 7
63 ± 8
53 ± 4
Td
54 ± 7
57 ± 4
50 ± 3
N3-ByTd
70 ± 8
78 ± 6
70 ± 4
Control (3% Tween80-saline)
53 ± 3
46 ± 3
56 ± 3
Ud
51 ± 3
46 ± 3
56 ± 3
NS-ByUd
72 ± 6*
89 ± 3**
85 ± 5**
Results are expressed as the mean sleeping time ± S.E. Td and N3-ByTd were injected at the dose of 0.5 pmol/mouse by i . c . v . Ud and Na-ByUd were injected at the dose of 1.0 pmol/mouse by i . c . v . PB was injected at the dose of 40 mg/kg by i . p . a f t e r compounds pretreated. * and ** indicate s i g n i f i c a n t l y d i f f e r e n t from control level of p
Motor Incoordination: As shown in Fig. 2, N3-ByTd and Na-ByUd e l i c i t e d motor incoordination. B o t h parent compounds showed a l i t t l e e f f e c t at 5 min, but they exhibited almost no e f f e c t , when compared with control, a f t e r t h i s time. Recovery times from motor incoordination induced by N3-ByTd and Na-ByUd were 6 hr and 1.5 hr, r e s p e c t i v e l y . Moreover, although the dose of N3-ByTd administered to mice was h a l f the dose of N3-ByUd, N3-ByTd exhibited a stronger incoordination e f f e c t than did N3-ByUd. Activity: Spontaneous a c t i v i t y of animals treated with N3~a~ OUS ed compounds by i . c . v , and i . v . i n j e c t i o n is summarized in both Table I I and I I I , r e s p e c t i v e l y . A c t i v i t y a f t e r N3-ByTd was reduced to 8% of control level. In addition, administration of Td ( i . c . v . ) also decreased the spontaneous a c t i v i t y , but i t s e f f e c t was very weaker than that of N3-ByTd. Effects of N3-ByUd and Ud were s i m i l a r to Na-ByTd and Td on a c t i v i t y . When N3-ByTd and N3-ByUd were given by i . v . injection (1.0 mmol/kg), the spontaneous a c t i v i t i e s were 4% and 55% of control, r e s p e c t i v e l y . Both i . c . v , and i . v . administration of N3-ByTd s i g n i f i c a n t l y decreased spontaneous a c t i v i t y in mice.
Vol. 41, No. 26, 1987
Hypnotic Activity of N3-ByTd
2795
100
Gv
50
to b_
O
0
~
OA
1
OA
OA
OA
C~
2
3
4
5
OAt
6
Time after administration ( h r ) Fig. 2 Motor incoordination effects of N3-ByTd, NLByUd and t h e i r parent compounds. Results are expressed as percent of mice that f e l l from the bar. Treatments: N3-ByTd (m), Td ( [ ] ) , N3-ByUd (A), Ud (A) and 3% Tween80-saline as control ( 0 ) . N3-ByTd and Td were administered at the dose of 0.5 ~mol/mouse by i . c . v . N3ByUd and Ud were administered at the dose of 1.0 umol/mouse by i . c . v . Eight mice were in each group.
Table I I Spontaneous a c t i v i t y of animals treated with N3-ByTd, N3-ByUd and t h e i r parent compounds by i . c . v , administration. Compound Control
Dose (~mol/mouse) (3% Tween80-saline)
Total distance (inches) 2258 ± 329
Td
0.5
919 ± 236**
N3-ByTd
0.5
185 ± 65
Control
(3% Tween80-saline)
% of control 100 41 8
2556 ± 250
100
Ud
1.0
1134 ± 96
44
N~-ByUd
1.0
27 ± ~ *
I
Results are expressed as the mean distance of animal movement ± S.E. Compounds tested were injected by i . c . v . ** indicates s i g n i f i c a n t l y d i f f e r e n t from the control level of p
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Hypnotic Activity of N3-ByTd
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Table I I I Spontaneous a c t i v i t y of animals treated with N3-ByTd and N3-ByUd by i . v . administration Compound
Control
Dose (mmol/k~) (3% TweenSO-saline)
Total distance (inches)
% of controT
1393 ± 121
100
N3-ByTd
1.0
58 ± 19"*
4
N3-ByUd
1.0
773 ± 172"
55
Results are expressed as the mean distance of animal movement ± S.E. Compounds tested were injected by i . v . * and ** indicate s i g n i f i c a n t l y d i f f e r e n t from the control of p
Discussion The present study demonstrates that CNS depressant e f f e c t of N3-ByTd, especially those of hypnotic a c t i v i t y and motor incoordination, was the most potent among other compounds tested. N3-ByTd c l e a r l y showed pronounced effects on motor incoordination. Recovery time of Ng-ByTd from motor incoordination (6 hr) was f o u r - f o l d that of N3-ByUd (1.5 hr) in spite of its being given at half the dose. In the case of N3-allyl substituted pyrimidine nucleosides, we already reported that N3-allylated compounds potentiated diazepam-induced motor incoordination, although the compounds themselves could not e l i c i t motor incoordination (3.8 ~mol/mouse, i . c . v . ) (2), i . e . , the benzyl group at N3 position must play an important role to producing the CNS depressant e f f e c t s . Some studies (7, 13) have reported on the CNS depressant a c t i v i t y of pyrimidine nucleosides; however, these compounds have not been reported to possess hypnotic a c t i v i t y . Recently, we found that N3-ByUd has hypnotic activity. At this point, we took i n t e r e s t in the structure of pyrimidine nucleosides as CNS depressants, because the sugar moiety is a constituent of pyrimidine nucleosides, and there are no hypnotics to r e l a t e with pyrimidine nucleosides, to our knowledge. In this regard, Honda et al. (10) revealed the necessity of a sugar moiety in the sleep-promoting e f f e c t . On the other hand, Wenzel and Keplinger (11) reported that some pyrimidine derivatives and related oxypyrimidines prolonged hexobarbital-induced sleep times. As other pharmacological e f f e c t s , Ud, uracil and some related compounds protected against the seizures (12, 13). The present results confirmed the CNS depressant a c t i v i t y of pyrimidine nucleosides, and the difference between Td and Ud analogues on CNS a c t i v i t y . N3-ByTd and N3-ByUd e l i c i t e d hypnotic a c t i v i t y only by i . c . v , i n j e c t i o n , but not i . v . This r e s u l t may be due to the blood brain b a r r i e r , metabolism and/or the a f f i n i t y for a putative receptor. However, spontaneous a c t i v i t y of animals treated with N3-ByTd and N3-ByUd was s i g n i f i c a n t l y decreased by both i . v . and i . c . v , administration. The results suggest that N3benzylated compounds penetrate into the brain in s u f f i c i e n t amounts to induce
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Hypnotic Activity of N3-ByTd
2797
CNS a c t i v i t y . In conclusion, we found that a new hypnotic substance, NS-ByTd, exerts hypnotic action and other CNS depressant e f f e c t s .
References I. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13.
I. YAMAMOTO, T. KIMURA, Y. TATEOKA, K. WATANABE and I. K. HO, Chem. Pharm. B u l l . , 33, 4088-4090 (1985). I. YAMAMOTO, T. KIMURA, Y. TATEOKA, K. WATANABE and I. K. HO, Res. Commun. Chem. Pathol. Pharmacol., 5_22, 321-332 (1986). I. YAMAMOTO, T. KIMURA, Y. TATEOKA, K. WATANABE and I. K. HO, J. Med. Chem., 30, in press (1987). S. INOUE, K. HONDA, K. UCHIZOND, R. UENO and O. HAYAISHI, Proc. Natl. Acad. Sci. U.S.A., 81, 6240-6244 (1984). Y. KOMODA, M. ISHIKAWA, H. NAGASAKI, M. IRIKI, K. HONDA, S. INOUE, A. HIGASHI and K. UCHIZONO, Biomed. Res., 4, 223-228 (1983). K. HONDA, Y. KOMODA, S. NISHIDA, H. NAGASAKI, A. HIGASHI, K. UCHIZONO and S. INOUE, Neurosci. Res., I, 243-252 (1984). R. S. KROOTH, W. L, HSIAO an-d G. F. M. LAM, J. Pharmacol. Exp. Ther., 207, 504-514 (1978). T. SASAKI, K. MINAMOTO and H. SUZUKI, J. Org. Chem., 3__88, 598-607 (1973). T. J. HALEY and W, G. McCORMICK, B r i t . J. Pharmacol., 12, 12-15 (1957). K. HONDA, Y, KOMODA and S. INOUE, Report of the i n s t i t u t e for Medical and Dental Engineering (Tokyo Medical and Dental U n i v e r s i t y ) , 18, 93-95 (1984). D. G. WENZEL and M. L. KEPLINGER, J. Am. Pharm. Assoc., 44, 56-59 (1955). D. G. WENZEL, J. Am. Pharm. Assoc., 44, 550-553 (1955). C. A. ROBERTS, Brain. Res., 5__55,291-3~8 (1973).
Pergamon Journal~
HYPNOTIC ACTIVITY OF N3-BENZYLTHYMIDINE ON MICE Ikuo Yamamoto,f Toshiyuki Kimur~ Yuji Tateoka, Kazuhito Watanabe and Ing Kang Ho* Department of Hygienic Chemistry, School of Pharmacy, Hokuriku University, 3-Ho, Kanagawa-machi,Kanazawa 920-11, Japan and * Department of Pharmacology and Toxicology, The University of Mississippi Medical Center, Jackson, Mississippi 39216, U.S.A. (Received in final form October 26, 1987)
Summary The pharmacological effect in mice of N3-benzylthymidine (N3ByTd) was examined by two routes of administration; intravenous ( i . v . ) and intracerebroventricular ( i . c . v . ) , and compared with the effect of administration of N3-benzyluridine (N3-ByUd) previously reported. Hypnotic a c t i v i t y , pentobarbital (PB)-induced sleeping time, motor incoordination and spontaneous a c t i v i t y were used as indices of pharmacological effects. N3-ByTd (0.5-2.0 umol/mou~e, i.c.v.) and N3-ByUd (1.5-3.0 umol/mouse, i . c . v . ) were found to possess dose-dependent hypnotic a c t i v i t y , and N3-ByTd had more potent hypnotic a c t i v i t y than N3-ByUd. Both N3-ByTd and N3-ByUd (0.5 and 1.0 ~mol/mouse, i . c . v . , respectively) showed a synergistic effect on PB-induced sleep, although t h e i r parent compounds, thymidine (Td) and uridine (Ud), did not potentiate the a c t i v i t y at each dose. In motor incoordination, the effect of N3-ByTd (0.5 umol/mouse) continued for 6 hr after i . c . v , injection. All compounds decreased the spontaneous activity of mice by i . c . v , administration. Furthermore, both N3-ByTd and N3-ByUd decreased the a c t i v i t y , when they were administered by i . v . These results reveal that both N3benzylpyrimidine nucleosides have more direct depressant effects on the central nervous system (CNS) than the parent compounds. Among the pyrimidine nucleoside derivatives tested, N3-ByTd was found to be the most potent hypnotic substance. Recently, we reported that N3-ByUd exerts hypnotic a c t i v i t y by i . c . v . administration in mice ( I ) , and that N3-allyluridine and N3-allylthymidine also have CNS depressant effects, although these compounds do not cause hypnotic a c t i v i t y in mice (2, 3). We thus proposed that N3-ByUd is a new type of hypnotic substance because of a characteristic sugar moiety at the N position in i t s structure. To our knowledge, there is no report with regard to hypnotic a c t i v i t y by pyrimidine nucleosides and their analogues. Ud is known as one of the sleep-promoting substances which have been extracted from brainstem of sleep-deprived rats (4-6). Krooth et al. (7) have reported that Ud, Td and other natural pyrimidines decreased spontaneous f To whom all
correspondence should be addressed
0024-3205/87 $3.00 + .00 Copyright (c) 1987 Pergamon Journals Ltd.
2792
Hypnotic Activity of N3-ByTd
Vol. 41, No. 26, 1987
a c t i v i t y at high doses in mice. In t h i s case, they indicate that Td is more potent than Ud. Therefore, i t is suggested that the introduction of a benzyl to the N3 position of Td may cause stronger hypnotic effects than N3-ByUd. The present study was thus designed to compare the hypnotic and depressant a c t i v i t y of N3-ByTd and Td with N3-ByUd and Ud. Materials and Methods Animals: Male ddN s t r a i n mice weighing 25 ± 3 g were used. They were kept in controlled l i g h t i n g (7:00 to 19:00 l i g h t period), and ambient temperature (23 ± I°C) throughout the experiments. T h e y were given food and water ad libitum. Chemicals: Td and Ud were purchased from Kohjin Co., Ltd. Tokyo and Wako Pure Chemical Ind., Ltd. Osaka, r e s p e c t i v e l y . Pentobarbital sodium was obtained from Tokyo Kasei Co., Ltd. Tokyo. Preparation of N3-Benzylated Compounds: N3-ByTd and N3-ByUd were prepared according to the method of Sasaki et a l . (8). Drug Administration: N3-ByTd, Td, N3-ByUd and Ud were suspended in 3% Tween80-saline solution because of i n s o l u b i l i t y of N3-benzylated compounds in saline. PB was injected i n t r a p e r i t o n e a l l y ( i . p . ) at the dose of 40 mg/kg. The i . c . v , i n j e c t i o n was performed according to the method of Haley and McCormick (9). In spontaneous a c t i v i t y t e s t , i . v . administration was also used in mice via t a i l vein. Pharmacological Effects: Sleeping time, as hypnotic a c t i v i t y , was measured as the time between the loss and recovery of r i g h t i n g r e f l e x in mice. This endpoint was also used for the PB-induced sleeping time. In the dose response experiments, mice were given compounds, by i . c . v , administration, at the dose of 0.25-3.0 ~mol/mouse. PB-induced sleeping time was measured a f t e r i . p . i n j e c t i o n of sodium PB. The tested compound was usually given by i . c . v . administration 15 min before the PB application. For time-course studies, a l l tested compounds were administered i . c . v . 5, 15 and 45 min p r i o r to the i n j e c t i o n of PB. Throughout PB-induced sleeping e f f e c t , motor incoordination and spontaneous a c t i v i t y t e s t s , 0.5 umol/mouse of N3-ByTd and Td was injected by i . c . v , administration, while 1.0 umol/mouse of N3-ByUd and Ud was injected. Each dose was decided by preliminary t e s t s . The dose of 0.5 umol/mouse N3-ByTd did not show any overt CNS t o x i c i t y . Effect of motor incoordination was measured by a bar t e s t . Compounds were injected, then mice were put on a bar (I cm square, 40 cm height). Motor incoordination a c t i v i t y of compounds tested was expressed as the percent of mice that f e l l from the bar within 30 seconds. Spontaneous a c t i v i t y was examined using a Digiscan A c t i v i t y Monitor, coupled with a Digiscan Computer (Omnitech Electronics Inc., U.S.A.) a f t e r both i . v . and i . c . v , administration, and the treated mice were placed in the center of a square p l a s t i c box with 30 cm sides. Measurement of a c t i v i t y was evaluated as the t o t a l distance (inches) of movement within I hr a f t e r injection. Multiple t-test (ANOVA) was u s e d for calculation of s t a t i s t i c a l significance. Results Hypnotic A c t i v i t y : Sleeping times of animals by i . c . v , administration are shown in Fig. possessed hypnotic a c t i v i t y at the doses of ~mol/mouse,respectively. N3-ByUd f a i l e d to
treated with N3-ByTd and N3-ByUd I. Both N3-ByTd and N3-ByUd 0.5-2.0 ~mol/mouse and 1.5-3.0 induce hypnotic a c t i v i t y at 1.0
Vol. 41, No. 26, 1987
Hypnotic Activity of N3-ByTd
0
2793
0
CH3
.oc OH OH
OH
NS-ByUd
N3-ByTd
100 t-
E= E
5C r Q.
u~
1 2 Dose ( p moll mouse)
3
Fig. 1 Hypnotic a c t i v i t y of N3-ByTd and N3-ByUd on mice. N3-ByTd (•) and N3 -ByUd ( • ) were administered by i . c . v . administration. Each point corresponds to the mean sleeping time ± S.E. Seven or 8 mice were used in each group.
~mol/mouse. Sleeping time of of N3-ByUd at 2.0 ~mol/mouse.
N3-ByTd was longer (about two-fold)
than that
PB-Induced Sleep: Table I summarizes the time course e f f e c t of p r i o r treatment of test compounds on PB-induced sleeping time. N3-ByTd (0.5 ~mol/mouse, i . c . v . ) potentiated the PB-induced sleeping time s i g n i f i c a n t l y at 45 min a f t e r the treatment. Td administration did not produce a s i g n i f i c a n t difference at any time interval as compared to control. NS-ByUd, at all time points, showed s i g n i f i c a n t p o t e n t i a t i o n , but i t s parent compound, Ud, did not change the sleep e f f e c t as well as Td.
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Hypnotic Activity of N3-ByTd
Vol. 41, No. 26, 1987
Table I Time course of the effects of N3-ByTd, N3-ByUd and t h e i r parent compounds on PB-induced slee p Compound
Sleepin 9 time (min) Time i n t e r v a l p r i o r to PB administration 5 15 45 (min)
Control (3% TweenSO-saline)
52 ± 7
63 ± 8
53 ± 4
Td
54 ± 7
57 ± 4
50 ± 3
N3-ByTd
70 ± 8
78 ± 6
70 ± 4
Control (3% Tween80-saline)
53 ± 3
46 ± 3
56 ± 3
Ud
51 ± 3
46 ± 3
56 ± 3
NS-ByUd
72 ± 6*
89 ± 3**
85 ± 5**
Results are expressed as the mean sleeping time ± S.E. Td and N3-ByTd were injected at the dose of 0.5 pmol/mouse by i . c . v . Ud and Na-ByUd were injected at the dose of 1.0 pmol/mouse by i . c . v . PB was injected at the dose of 40 mg/kg by i . p . a f t e r compounds pretreated. * and ** indicate s i g n i f i c a n t l y d i f f e r e n t from control level of p
Motor Incoordination: As shown in Fig. 2, N3-ByTd and Na-ByUd e l i c i t e d motor incoordination. B o t h parent compounds showed a l i t t l e e f f e c t at 5 min, but they exhibited almost no e f f e c t , when compared with control, a f t e r t h i s time. Recovery times from motor incoordination induced by N3-ByTd and Na-ByUd were 6 hr and 1.5 hr, r e s p e c t i v e l y . Moreover, although the dose of N3-ByTd administered to mice was h a l f the dose of N3-ByUd, N3-ByTd exhibited a stronger incoordination e f f e c t than did N3-ByUd. Activity: Spontaneous a c t i v i t y of animals treated with N3~a~ OUS ed compounds by i . c . v , and i . v . i n j e c t i o n is summarized in both Table I I and I I I , r e s p e c t i v e l y . A c t i v i t y a f t e r N3-ByTd was reduced to 8% of control level. In addition, administration of Td ( i . c . v . ) also decreased the spontaneous a c t i v i t y , but i t s e f f e c t was very weaker than that of N3-ByTd. Effects of N3-ByUd and Ud were s i m i l a r to Na-ByTd and Td on a c t i v i t y . When N3-ByTd and N3-ByUd were given by i . v . injection (1.0 mmol/kg), the spontaneous a c t i v i t i e s were 4% and 55% of control, r e s p e c t i v e l y . Both i . c . v , and i . v . administration of N3-ByTd s i g n i f i c a n t l y decreased spontaneous a c t i v i t y in mice.
Vol. 41, No. 26, 1987
Hypnotic Activity of N3-ByTd
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100
Gv
50
to b_
O
0
~
OA
1
OA
OA
OA
C~
2
3
4
5
OAt
6
Time after administration ( h r ) Fig. 2 Motor incoordination effects of N3-ByTd, NLByUd and t h e i r parent compounds. Results are expressed as percent of mice that f e l l from the bar. Treatments: N3-ByTd (m), Td ( [ ] ) , N3-ByUd (A), Ud (A) and 3% Tween80-saline as control ( 0 ) . N3-ByTd and Td were administered at the dose of 0.5 ~mol/mouse by i . c . v . N3ByUd and Ud were administered at the dose of 1.0 umol/mouse by i . c . v . Eight mice were in each group.
Table I I Spontaneous a c t i v i t y of animals treated with N3-ByTd, N3-ByUd and t h e i r parent compounds by i . c . v , administration. Compound Control
Dose (~mol/mouse) (3% Tween80-saline)
Total distance (inches) 2258 ± 329
Td
0.5
919 ± 236**
N3-ByTd
0.5
185 ± 65
Control
(3% Tween80-saline)
% of control 100 41 8
2556 ± 250
100
Ud
1.0
1134 ± 96
44
N~-ByUd
1.0
27 ± ~ *
I
Results are expressed as the mean distance of animal movement ± S.E. Compounds tested were injected by i . c . v . ** indicates s i g n i f i c a n t l y d i f f e r e n t from the control level of p
2796
Hypnotic Activity of N3-ByTd
Vol. 41, No. 26, 1987
Table I I I Spontaneous a c t i v i t y of animals treated with N3-ByTd and N3-ByUd by i . v . administration Compound
Control
Dose (mmol/k~) (3% TweenSO-saline)
Total distance (inches)
% of controT
1393 ± 121
100
N3-ByTd
1.0
58 ± 19"*
4
N3-ByUd
1.0
773 ± 172"
55
Results are expressed as the mean distance of animal movement ± S.E. Compounds tested were injected by i . v . * and ** indicate s i g n i f i c a n t l y d i f f e r e n t from the control of p
Discussion The present study demonstrates that CNS depressant e f f e c t of N3-ByTd, especially those of hypnotic a c t i v i t y and motor incoordination, was the most potent among other compounds tested. N3-ByTd c l e a r l y showed pronounced effects on motor incoordination. Recovery time of Ng-ByTd from motor incoordination (6 hr) was f o u r - f o l d that of N3-ByUd (1.5 hr) in spite of its being given at half the dose. In the case of N3-allyl substituted pyrimidine nucleosides, we already reported that N3-allylated compounds potentiated diazepam-induced motor incoordination, although the compounds themselves could not e l i c i t motor incoordination (3.8 ~mol/mouse, i . c . v . ) (2), i . e . , the benzyl group at N3 position must play an important role to producing the CNS depressant e f f e c t s . Some studies (7, 13) have reported on the CNS depressant a c t i v i t y of pyrimidine nucleosides; however, these compounds have not been reported to possess hypnotic a c t i v i t y . Recently, we found that N3-ByUd has hypnotic activity. At this point, we took i n t e r e s t in the structure of pyrimidine nucleosides as CNS depressants, because the sugar moiety is a constituent of pyrimidine nucleosides, and there are no hypnotics to r e l a t e with pyrimidine nucleosides, to our knowledge. In this regard, Honda et al. (10) revealed the necessity of a sugar moiety in the sleep-promoting e f f e c t . On the other hand, Wenzel and Keplinger (11) reported that some pyrimidine derivatives and related oxypyrimidines prolonged hexobarbital-induced sleep times. As other pharmacological e f f e c t s , Ud, uracil and some related compounds protected against the seizures (12, 13). The present results confirmed the CNS depressant a c t i v i t y of pyrimidine nucleosides, and the difference between Td and Ud analogues on CNS a c t i v i t y . N3-ByTd and N3-ByUd e l i c i t e d hypnotic a c t i v i t y only by i . c . v , i n j e c t i o n , but not i . v . This r e s u l t may be due to the blood brain b a r r i e r , metabolism and/or the a f f i n i t y for a putative receptor. However, spontaneous a c t i v i t y of animals treated with N3-ByTd and N3-ByUd was s i g n i f i c a n t l y decreased by both i . v . and i . c . v , administration. The results suggest that N3benzylated compounds penetrate into the brain in s u f f i c i e n t amounts to induce
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Hypnotic Activity of N3-ByTd
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CNS a c t i v i t y . In conclusion, we found that a new hypnotic substance, NS-ByTd, exerts hypnotic action and other CNS depressant e f f e c t s .
References I. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13.
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