CURRENT THERAPEUTIC RESEARCH VOL. 56, NO. 5, MAY 1995

EFFECTS OF CEREBRAL METABOLIC ENHANCERS ON FUNCTION IN RODENTS

KOICHIRO TAKAHASHI,l MINORU YAMAMOTO,’ MASANORI SUZUKI,’ YUKIKO OZAWA,’ TAKASHI YAMAGUCHI,l HIROFUMI ANDOH, AND KOUICHI ISHIKAWA2

‘Department of Pharmacology, Clinical Pharmacology Research Laboratory, Yamunouchi Pharmaceutical Co. Ltd., and ‘Department of Pharmacology, School of Medicine, Nihon University, Tokyo, Japan

AFWI’RACT

The effects of cerebral metabolic enhancers (, bi- femelane, idebenone, and ) on -induced hypother- mia, the immobility period in forced swimming tests, and passive avoidance learning behavior were compared with the effects of ami- triptyline in rodents. Indeloxazine, , and antagonized hypothermia in mice given reserpine. Indeloxaxine and amitriptyline decreased the immobility period in mice in the forced swimming test in a dose-dependent manner. The latency of step- through in the passive avoidance test in rats was prolonged by ad- ministration of indeloxazine but shortened by administration of amitriptyline. Neither idebenone nor nicergoline displayed any phar- macologic action in these tests. The results suggest that indeloxaxine possesses an activity similar to that of amitriptyline but differs from amitriptyline in its anticholinergic properties and its ability to ameliorate impaired brain function such as that of learning behavior. In addition, indeloxazine exhibited broader effects on brain functions than either bifemelane, idebenone, or nicergoline.

INTRODUCTION

Cerebral metabolic enhancers (drugs that enhance energy metabolism) including brain glucose and ATP levels such as indeloxazine,1*2 bi- femelane, 3*4idebenone?6 and nicergoline,7>8 are currently used for the treatment of patients with various psychiatric symptoms. These symptoms include reduced spontaneity and emotional disturbance in patients with cerebral vascular disease. Clinical trials of these drugs for the treatment of in aged populations are now in progress. However, the antide- pressant actions of these drugs have not been compared under the same experimental conditions. The present study compared the effects of indel- oxaxine, bifemelane, idebenone, and nicergoline with those of amitripty-

Address correspondence to: M. Yamamoto, Department of Pharmacology, Clinical Pharmacology Research Laboratory, Yamanouchi Pharmaceutical Co. Ltd., l-l-8, Azusawa, Itabashi-ku, Tokyo 174, Japan. Received forpublication on February 16,1995. Printed in the U.S.A. Reproduction in whole or part is not permitted.

478 oo11-393x196/~3.50 K. TAKAHASH ET AL..

line on reserpine-induced hypothermia, forced swimming, and passive avoidance learning behavior in rodents. Amitriptyline, a typical antide- pressant currently used for the treatment of depressed patients, was cho- sen as the positive control drug to evaluate the antidepressive activity of the other study drugs.

MATERIALSANDMETHODS

The experiments were carried out using male ICR strain mice (approxi- mately 30 g each) and male Wistar rats (approximately 300 g each). The animals were group-housed in cages under a l&hour light-dark cycle (lights turned on between 7:30 AM and 830 PM) with laboratory chow and water given ad libitum. Test drugs were administered orally to the mice 18 hours after the subcutaneous administration of reserpine (2 mg/kg). Rectal temperature was measured immediately before and at 60, 120, and 180 minutes after drug administration.

Drugs

Indeloxazine hydrochloride and amitriptyline hydrochloride, synthe- sized at Yamanouchi Pharmaceutical Co., Ltd., Tokyo, Japan, were dis- solved in distilled water and administered orally. Bifemelane hydrochlo- ride, idebenone, and nicergoline, purchased commercially from Eisai, Tokyo, Japan, Takeda, Tokyo, Japan, and Tanabe Co. Ltd., Tokyo, Japan, respectively, were dissolved in 0.3% methylcellulose and administered orally. Distilled water and 0.3% methylcellulose were administered orally as vehicle control. Physostigmine sulfate (Tokyo Kasei Co. Ltd., Tokyo, Ja- pan), used as a positive control drug, was dissolved in saline solution. All drugs and vehicles were administered in a volume of 0.1 mLkg and 1 mL/kg to mice and rats, respectively.

Forced Swimming-Induced Immobility in Mice

The apparatus consisted of Plexiglass cylinders (24.5 cm x 6 cm x 18 cm; WW-3002, Ohara Co., Tokyo, Japan) with a waterwheel in the center. The waterwheel consisted of a Plexiglass shaft (5 cm long and 8 cm in diameter) on which 24 paddles (0.1 cm wide and 3.6 cm long) were attached at constant intervals. The waterwheel would move when application of a load greater than 5 g was applied to one of the paddles. The number of rotations of the waterwheel through an arc of 120” was counted via a photointerrupter attached to the shaft during the final 4 minutes of a 6-minute test. The tank contained water at a temperature of about 23 “C to a height of 12 cm; half of the paddles were resting in the water.

479 EFFECT OF INDELOWNE ON BRAIN FUNCTION

Tests were conducted according to the method of Nomura et al.’ When a mouse was dropped into the water, it attempted to climb onto the wheel to escape from the water but could not do so due to rotation of the water- wheel. However, when the animal’s attempts to escape were finally aban- doned, the waterwheel would stop turning. The number of wheel rotations before the animal abandoned its attempts to escape were counted. Each mouse was placed in the water tank 1 hour after receiving a single oral (PO) administration of the drug. The number of rotations of the wheel was counted every 2 minutes for 6 minutes. The wheel counts during the final 4 minutes of the g-minute test for each mouse receiving the drug were compared with those of a control group of mice which had received distilled water and were exposed to a comparable situation.

Passive Avoidance Learning in Rata

Training was conducted according to the step-through procedure de- scribed by Jarvik and Kopp. lo The apparatus (Ohara Co.) consisted of two compartments (with clear plastic tops): an illuminated compartment (40 cm x 25 cm x 25 cm) containing a 60-watt lamp 25 cm above the top of the compartment (50 cm above the floor) and a dark compartment (25 cm x 15 cm x 25 cm). The two compartments were separated by a closed guillotine door (10 cm x 7 cm). Before training of passive avoidance learning, each rat underwent a single pretraining trial using the following procedure. The rat was placed in the illuminated compartment and the opaque guillotine door was raised 10 seconds later. After the rat entered the dark compartment, it was al- lowed to remain there for 10 seconds. In order to conduct the training, the rat was placed in the illuminated safe compartment. A door allowed the rat to enter the dark compartment, which contained a grid on the floor. Once the rat’s four paws were on the grid, a foot shock (60 V, 50 Hz) was deliv- ered to the floor grid for 2 seconds. The rat could escape the shock only by stepping back into the illuminated compartment. The rat was then re- turned to its home cage. The test trial was conducted 24 hours after the training session. The rat was again placed in the illuminated compartment of the same appa- ratus used in training, and the response latency before entering into the dark compartment was measured. The results were recorded as the aver- age step-through latency for each experimental group of animals. The rats were given a single po drug dose 60 minutes before the training session, in accordance with the methods of Yamamoto and Shimizu.” The step- through latency was measured both during the acquisition trial and the test trial (24 hours after the acquisition trial). The maximum observation period for monitoring behavior in this test was set at 10 minutes.

480 K. T- ET Al..

Values are expressed as the mean + SE. Results for reserpine- hypothermia, forced swimming-induced immobility, and passive avoid- ance learning behavior were analyzed using Student’s t test and Mann- Whitney U test, respectively. P < 0.05and P < 0.01 were considered significant.

RESULTS

Reaerpine-Induced Hypothermia in Mice

Oral administration of indeloxaxine (1 and 3 mgkg), bifemelane (30 mg/kg po), and amitriptyline (1 and 3 mg/kg) antagonized the reserpine- induced hypothermia in a dose-dependent manner (Table I). In contrast, oral administration of idebenone (10 and 30 mg/kg) and nicergoline (1 to 10 mg/kg) exerted no effect on the reserpine-induced hypothermia.

Forced Swimming-Induced Immobility in Mice

Both indeloxaxine (50 mg/kg po) and amitriptyline (30 mg/kg po) in- creased the number of rotations by mice in a dose-dependent manner (Ta-

Table I. Effects of indeloxazine, bifemelane, idebenone, nicergoline, and amitriptyline on reserpine-induced hypothermia in mice. Each value representa the mean for 6 to 7 mice.

Temperature Change (2). Dose Tnatment (mo/lro 0W) 1 hour 2 hours 3houn

Control f 0.7 f 0.4 ” 0.7 Control E : x.: 4.5 ” 0.6 Bifemelane 4:3 r 0:3 ;.; I ;.;$ 2:2 : 0’6 1.1 + 0:3 Control Nicergoline ?A I E 2:3 I 013 I!! i:3 f 0.4 1.6 2 0.6 Control 2.9 f 0.5 3.3 !?0.6 Amitriptyline ?: r :.;t 4.4 k 0.6 z.45 ;.;$ 3:9 z 0:7* 5.9 * 0.6$ . +f .

* The difference in rectal temperature measured at l-hour intervals after treatment. The rectal temperature immediately before administration of the test drugs ran ed from 28.0 to 28.5 “C. Si nificantly different from the control group: tf < 0.0‘5 , SP c 0.01 (Student’s t test). Ni! = not done.

481 EFFECT OF INDELOXAZINB ON BRAIN FUNCTION

ble II). However, bifemelane (10 to 100 mg/kg PO), idebenone (10 to 100 mg/kg PO), and nicergoline (1 to 30 mg/kg po) had little effect on this variable.

Passive Avoidance Learning in Rats

Indeloxazine (10 and 20 mg/kg po) prolonged the step-through latency in rats (Table III). The facilitator-y actions of both indeloxaxine and phy- sostigmine showed bell-shaped responses. In contrast, bifemelane (3 to 30 mg/kg PO),idebenone (3 to 30 mg/kg PO),and nicergoline (1 to 10 mg/kg po) had no effect on passive avoidance behavior. Amitriptyline (4 mg/kg in- traperitoneally) shortened latency in this test.

DISCUSSION AND CONCLUSION

Forced swimming is a standard pharmacologic technique for evaluating the antidepressant activity of drugs. l2 The present study demonstrated that indeloxazine and amitriptyline caused a dose-dependent decrease in the forced swimming-induced immobility period of mice. The finding that both indeloxazine and amitriptyline antagonized reserpine-induced hypo-

Table II. Effects of indeloxazine, bifemelane, idebenone, nicergoline, and amitriptyline on forced swimming-induced immobility in mice. Each value represents the mean f SE for 10 mice.

Dose Treatment (mgkg orally) Rotations”

Control lndeloxazine

Control Bifemelane

Control ldebenone

Control Nicergoline

Control Amitriptyline

* Number of times in which mice rolled one third of the wheel in forced swimming apparatus during the final 4 minutes of a 6-minute test. Significantly different from control group: tf < 0.05; $f < 0.01 (Mann-Whitney U statistic test). K. TAKAHAMI ET AL.

Table III. Effects of indeloxazine, bifemelane, idebenone, nicergoline, amitriptyline, and phy- sostigmine on passive avoidance learning step-through latency in rats. Each value represents the mean f SE for eight rate.

Sisp-Tllroughlatency’ Tmrbnent (seconds)

Control lndeloxazine 3p0

;gi 27i z 52’ Control Bifemelane 3p0 10 po 30 po Control ldebenone &lo 10 po 30 po Control Nicergoline ,s 10 no Control Physostigmine

Control 217 * 54 Amitriptyline 0.5lP 1 IP 1931491:: ::: 2 IP 4 IP 56 r ll$ po = orally; IP = intraperitoneally. l Ste -through latency before rats enter dark compartment in passive avoidance learning test. SigniP icantly different from the control group: tP < 0.05, $P < 0.01 (Mann-Whitney U statistic test). thermia in mice indicates a facilitatory effect of both drugs on the central system, including the noradrenergic and serotoninergic systems. The antidepressant effects of indeloxazine and amitriptyline ob- served in the forced swimming test may thus be ascribable to facilitation of the central monoaminergic system. l3 In contrast, acquisition of passive learning behavior in rats was accelerated by the administration of indel- oxazine but disrupted by amitriptyline, due, in part, to its anticholinergic property. The mechanism of such facilitation in passive learning behavior is not clear, but may involve facilitatory effects of indeloxazine on the central serotoninergic and systems. Indeloxazine has been shown to increase release from the cerebral cortex in rats using a microdialysis method14 and to decrease the number of reserpine- induced pontogeniculo-occipital waves in cats.” Indeloxazine has been re- ported to ameliorate the impairment of passive avoidance behavior in mid- dle cerebral artery-occluded rats16 and senescence-accelerated mice.17 These results suggest that indeloxazine can exert antidepressant and

483 learning enhancement effects and may be useful in the treatment of de- pressive symptoms in patients with organic brain syndrome. Unlike indeloxazine and amitriptyline, bifemelane, idebenone, and nicergoline displayed little effect on immobility in the forced swimming test. Bifemelane has been reported to decrease the immobility period in a forced swimming test but no dose dependency was observed.” Bifemelane, but not idebenone or nicergoline, antagonized reserpine-induced hypother- mia, suggesting that bifemelane exerts facilitatory effects on the central noradrenergic system. This agrees with the finding of Tobe et a1.3 Fur- thermore, neither bifemelane, idebenone, nor nicergoline changed passive learning behavior in rats. In previous studies, however, bifemelane,3 ide- benone,lg and nicergoline7 have been shown to ameliorate or facilitate learning behavior. The discrepancy between the present results and pre- vious reports may be related to study conditions such as dosage and ex- perimental procedures. Amitriptyline is currently used for the treatment of endogenous de- pression in young patients. Its use in treating depression in aged popula- tions or in patients with organic brain syndrome may be limited due to the associated deterioration of brain function and other side effects (eg, anti- depressants with anticholinergic properties frequently induce cardiotoxic- ity and dry mouth). 2o This study has shown that in addition to having antidepressant properties, indeloxazine facilitates learning behavior, partly through its effects on the central monoaminergic system.

References:

1. Ohtomo E. Clinical evaluation of YM-08064 (indeloxazine) in the treatment of cerebro- vascular disorder. ZgcskuNo Ayumi. 1986136535-555. In Japanese. 2. Yamamoto M. Pharmacological and biochemical properties of indeloxaxine hydrochlo- ride, a new cerebral activator. Brain Dysfinct. 1990;3:130-147. 3. Tobe A, Egawa M, Nagi R. Effect of 4(o-benxylphenoxyl-N-methylbutylamine hydrochlo- ride (MCI-2016) on the -induced deficit of spontaneous alteration behavior in rata. Jpn J Pharmucol. 1983;33:775-784. 4. Tazaki Y, Kutsuzawa T, Tohgi H, et al. Utility of E-0687 in cerebral vascular disease. Zgaku No Ayumi. 1986;137:647-670. In Japanese. 5. Nagaoka N, Shino A, Kakihana M, Iwatsuka H. Inhibitory effect of idebenone 6X-26191, a novel compound, on vascular lesions in hypertensive rata Jpn J Pharmncol. 1984,36: 291-299. 6. Ohtomo E, Abe H, Araki G, et al. Studies of utility of CV-2619 in cerebral vascular disease. Ther Res. 1985;3:117-136. In Japanese. 7. Moretti A. Metabolic and neurochemical effects of nicergoline on the central nervous system: A review of the experimental studies. An&m-Forsch Drug Res. 1979;29:1213- 1223. 8. Ohtomo E, Hirai K, Hasegawa K, et al. Clinical evaluation of TA-079 (nicergoline) in the treatment of cerebrovascular disorders. Clin EL&. 198614575-602. In Japanese. K.T-El’AL.

9. Nomura S, Shimixu J, Kinjo M, et al. A new behavioral test for antidepreeeant druge. Eur J Pharmacol. 1982;83:171-175. 10. Jarvik ME, Kopp A. An improved one-trial paeeive avoidance learning situation. Psych& Rep. 1967;21:221-224. 11. Yamamoto M, Shin&u M. Effecta of a new TRH analogue, YM-14673, on a paeeive avoidance teat ae a poeeible criterion of improvement in cognitive dieturbance in rodents. Naunyn-Schmiedeberg’s Anzh Pharmacol. 1988;338:262-267. 12. Poreolt BD, LePichon M, Jalfre M. Depreeeion: A new animal model sensitive to antide- pressant treatment. Nature. 1977;266:730-732. 13. Harada M, Maeno H. Biochemical characteristics of a potential antidepreeeant, 2-(7- indenyloxymethyl)morpholine hydrochloride (YM-08054-l), and ite derivativee with po- tential antidepressant properties. Biochem Pharmaco 1.1979;2&2645-2651. 14. Yamamoto M, Goyama M, Gzawa Y, et al. Effects of indeloxaxine hydrochloride, a cere- bral activator, on passive avoidance learning impaired by dieruption of & trans- mission in rate. Neurvpharmucology. 1993;32:696-701. 15. Yemamoto M, Shimizu M. Electroencephalographic etudiee on a new cerebral activator, indeloxexine hydrochloride. Curr Ther Rec. 1988;43:92-106. 16. Shimixu-Saeamata M, Yamamoto M, Okada M, et al. Effecta of indeloxazine hydrochlo- ride on behavioral and biochemical changes in the chronic phase of focal cerebral iech- emia in rate. Arch Znt Pharmacodyn Ther. 1991;314:74-89. 17. Yamamoto M, Shimizu M, Kawabata S, Iwai A. Effecta of indeloxaxine hydrochloride on learned behavior in senescence accelerated mouee. Eur J Pharmacol. 1989;166:345-348. 18. Moryl E, Danyez W, Quaack G. Potential antidepressive properties of , and bifemelane. Phurmacol Toxicol. 1993;72:394-397. 19. Yamazaki N, Take Y, Nagaoka A, Nagawa Y. Beneficial effect of idebenone on cerebral &hernia-induced amnesia in rate. Jpn J Pharmacol. 1984,36:349-356. 20. Williams GO. Management of depreeeion in the elderly. Primary Care. 1989;16:451-474.

485