ELSEVIER Atypicality of Several Antipsychotics on the Basis of In Vivo Dopamine-D2 and Serotonin-SHT2 Receptor Occupancy Tomiki Sumiyoshi, M.D., Kimiko Suzuki, M.D., Hiroshi Sakamoto, M.D., Nariyoshi Yamaguchi, M.D., Hirofumi Mori, M.D., Kazuhiro Shiba, Ph.D., and Koichi Yokogawa, Ph.D. An in vivo receptor binding technique was used to were found for clozapine, RMI-81512, and tiospirone evaluate the binding profiles of typical and atypical compared to haloperiodol and pimozide. Zotepine, antipsychotic drugs to striatal dopamine-D2 and frontal mosapramine, and clocapramine produced ratios that fall serotonin-S-HT2 receptors in a rat brain using more between these two groups. Chlorpromazine was specific ligands than those previously employed. [3HJ­ exceptional as a typical antispychotic by these criteria. YM-09151-2 or {3H]-ketanserin was injected into the tail Relatively strong antagonism of S-HT2 receptors by vein 10 minutes after administration of test drugs. One atypical antipsychotics was confirmed by this in vivo hour after the ligand injection, radioactivities in the measure of receptor binding using more selective ligands striatum, frontal cortex, and cerebellum were counted to than those used in previous studies. obtain receptor occupancies by the test drugs. Higher [Neuropsychopharmacology 12:57-64, 1995] ratios of potency in occupying 5-HT2 versus D2 receptors KEY WORDS: Atypical antipsychotic drugs; D2 receptors; likely to produce disturbances of EPS or elevation of S-HT2 receptors; Receptor binding study; In vivo serum prolactin (Young and Meltzer 1980; Porsolt and Jafre 1981; Neale et al. 1983), and demonstrate favor­ positive and negative The mode of action of antipsychotic drugs has been dis­ able clinical efficacy for both are resistant to treatment cussed in relation to their potency to block central symptoms of patients who et al. 1988). dopamine-D2 receptors (Creese et al. 1976; Seeman et with typical antipsychotics (Kane for serotoninz (5-HT2) al. 1976). The so-called "typical" antipsychotic drugs Since a relatively high affinity drugs was postulated have been shown to produce catalepsy in rats (Moore receptors of atypical antipsychotic Meltzer et al. (1989a, b), a number of studies to test and Gershon 1989) and often, extrapyramidal side by in laboratory effects (EPS) in patients (Sovner and DiMascio 1978), this hypothesis have been performed et al. 1992; Stock­ by blocking the nigrostriatal dopamine system. "Atyp­ animals (Leysen et al. 1992; Matsubara PET measurement ical" antipsychotic drugs, on the other hand, are less meier et al. 1993) and in human (Nordstrom et al. 1993; Nyberg et al. 1993). We have previously demonstrated that the atypical antipsychotic From the Department of Neuropsychiatry (TS, KS, HS, NY) Ra­ drugs clozapine, RMI-81582, and risperidone show rel­ dioisotope Center (HM, KS) and Hospitals Pharmacy (KY) of Kanazawa, University School of Medicine, Kanazawa 920, Japan atively higher 5-HT2 to D2 ratios of in vivo receptor Address correspondence to: Tomiki Sumiyoshi, M.D., Department affinities than haloperidol, measuring time courses of of Psychiatry, University Hospitals of Cleveland, Hanna Pavilion, a receptor occupancy by single doses of these drugs 11100 Euclid Ave, Cleveland, OH 44106. Received February 16, 1994; revised July 8, 1994; accepted July 19, (Sumiyoshi et al. 1993, 1994a, b). 1994. In the present study, we used [3H]-YM-09151-2 and NEUROPSYCHOPHARMACOLOGY 1995-VOL. 12, NO. 1 © 1995 American College of Neuropsychopharmacology Published by Elsevier Science Inc. 0893-133X/95/$9 .50 655 Avenue of the Americas, New York, NY 10010 SSDI 0893-133X(94)00064-7 58 T. Sumiyoshi et al. NEUROPSYCHOPHARMACOLOGY 1995-VOL. 12, NO. 1 [3H]-ketaserin, selective ligands for D2 and 5-HT2 re­ Xno)/(Xs-Xns] x 100 (%), where each abbreviation ceptors, respectively (Laduron et al. 1982; Niznik et al. represents the radioactivity (%dose/g tissue) of Xo, 1985; Terai et al. 1989), to measure the in vivo affinities striatum, or frontal cortex of drug-treated rats; Xno, of several antipsychotic drugs to these receptors by cerebellum of drug-treated rats; Xs, striatum or fron­ analyzing receptor occupancy. The test compounds in­ tal cortex of vehicle-treated rats; and Xns, cerebellum clude the new antipsychotic drug Y-516 (mosapramine) of vehicle-treated rats. andclocapramine,iminodibenzylderivativesthathave been shown to exert favorable clinical effect on schizo­ Data Analysis phrenic patients (Yamagami et al. 1988; Asano et al. 1992). The "atypicality" of these iminodibenzyls is also Dose-response competition curves were analyzed with determined on the basis of in vivo D2 and 5-HT2 recep­ the minimal-square method (Paalzow and Edlund 1979) tor occupancy in comparison with traditional typical and DEso (mg/kg) values reflecting binding potencies and atypical antipsychotics. of the test drugs were calculated. The EDso values were then transformed into (-)log of the EDso (mol/kg) (pEDso) to compare the receptor binding potencies of MATERIALS AND METHODS the test drugs in the present study with those of Stock­ meier et al. (1993). In Vivo Receptor Binding Male Wistar rats (210 to 240 g; Sankyo Laboratory, Drugs Toyama, Japan) were housed in a temperature-con­ trolled room with a 12-hour dark/light cycle (lights on [3H]-YM-09151-2 (specific activity 3.22 TBq/mmol) and at 8:30) and had free access to food and water. In vivo [3H]-ketanserin (specific activity 2.22 TBq/mmol) were binding of [3H]-YM-09151-2 (specific activity 3.22 purchased from DuPont New England Nuclear Corpo­ TBq/mmol) to D2 receptors or [3H]-ketanserin (specific ration (Boston, MA). The following compounds were activity 2.22 TBq/mmol) to 5-HT2 receptors was mea­ generously supplied by the manufacturers: haloperidol sured according to a method previously reported (DainipponPharmaceutical Co., Osaka, Japan); cloza­ (Sumiyoshi et al. 1993, 1994a, b). For the kinetic study, pine (Sandoz Ltd., Basel, Switzerland); RMI-81582 [3H]-YM-09151-2 or [3H]-ketanserin (1540 to 1680 kBq/ (Marion Merrill Dow Pharmaceuticals, Cincinnati, OH); kg body weight) were injected into the lateral tail vein tiospirone (Bristol-Myers-Squibb, Wallingford, CT); of rats that had received an intraperitoneal injection of chlorpromazine, mosapramine, clocapramine (Yoshi­ a vehicle (dimethyl-sulfoxide, Wako Chemical Co., tomi Pharmaceutical Ltd., Osaka, Japan); zotepine, Oaska, Japan, 1 ml/kg), 10 minutes before. The rats were pimozide (Fujisawa Pharmaceutical Co., Osaka, Japan). sacrificed by decapitation at 15, 30, 45, 60, 120, or 240 minutes after the injections of ligands. For competition RESULTS studies, rats were pretreated with an intraperitoneal in­ jection of varying doses of antipsychotic drugs or the The values of radioactivity in each brain region at the same volume (0.21 to O.24 ml) of the corresponding ve­ different time intervals after the injection of [3H]-YM- hicle (dimethyl-sulfoxide), 10 minutes prior to the in­ 09151-2 are shown in Figure 1. A relatively large amount jection of [3H]-YM-09151-2 or [3H]-ketanserin. Based of [·3H]-YM-09151-2 accumulates in the striatum as on results of the kinetic study, the rats were decapi­ compared to the cerebellum. For [3H]-ketanserin­ tated 60 minutes after the ligand injection. The brains treated rats, the values in the frontal cortex continued were rapidly removed and dissected into cerebellum, to be higher than those in the cerebellum, as shown striatum, frontal cortex, and the rest of the brain. After in Figure 2. In the following drug competition studies, weighing, each region of the brain was solubilized with rats were sacrificed 60 minutes after the injection of a tissue solubilizer (Soluene 350, Packard Co., CN) by radioligand, based on stable receptor occupancies by incubation for 2 to 3 hours at 50°C. A scintillation cock­ drugs at this time point as reported in our previous tail (Aquasol 2, New England Nuclear) was added to studies (Sumiyoshi et al. 1993, 1994a, b). Figures 3 to the solubilized tissues adjusted to pH 7.0 with 0.5 N 10 show the occupancy of 04 and 5-HT2 receptors fol­ HO solution (for inhibition of pseudo-fluorescence). Af­ lowing various doses of typical antipsychotic drugs, ter 12 to 24 hours, the radioactivity concentrations in pimozide and chlorpromazine; atypical antipsychotic the tissues were counted with a liquid scintillation drugs, clozapine, RMI-81582 and tiospirone; other counter (LSC-1000, Aloka Co., Tokyo, Japan) and the drugs, zotepine, mosapramine and clocapramine. The values, expressed as o/odose/g tissue, were calculated. dose-occupancy curves of haloperidol were reported Receptor occupancy was determined by a modification elsewhere (Sumiyoshi et al. 1994a). Based on these oc­ of the method of former reports (Geoders and Kuhar cupancy curves, the EDso values (mg/kg) of the test 1985; Miller et al. 1987): percent occupancy <1>=[1-(Xo- compounds for the in vivo occupancy of striatal D2 and NEUROPSYCHOPHARMACOLOGY 1995-VOL. 12, NO. 1 In Vivo Receptor Occupancy of Antipsychotics 59 .......... (I) ::J (/) (/) 2.0 i 100 -~ e 'OD (I) ..........• 1.5 a Striatum >, -(/) 0 0 0 . .t.. Cerebellum -0 (/) 50 a. 1.0 ::, o*+I C 0 .._.. ro 0 >, (I) 0 ·"'= >-E 0.5 Jl -~ -ii (.) ro 0.1 0 -0 0 0 ro 30 60 120 240 Dose of Pimozide (mg/kg) 0:: Time (minutes) Figure 3. Occupancy (<I>%) of striatal D2 and frontal 5-HT2 receptors in rat brain with various doses of pimozide. [3H]­ Figure 1. Kinetics of [3H]-YM-09151-2 binding in vivo in rat YM-09151-2 or [3H]-ketanserin (1540 to 1680 kBq/kg body brain. Rats were injected with [3H]-YM-09151-2 (1540 to 1680 weight) was injected into a tail vein at 10 minutes after ad­ kBq/kg body weight) at time zero and killed at various times ministration (IP) with various doses of pimozide. Sixty after injection. Values represent the mean± SD of five rats minutes after the ligand injection, radioactivity in the stria­ at each time point.