Vol. 226, No. 2

Printed in U.S.A.

ABBREVIATiONS: THD, thioridazine; HLP, Haloperidol; CLZ, clozapine; CNS, central nervous system; CAR, conditioned avoidance responding;CL, confidence limits; MED, minimal effective dose; NIAMDD, National Institute of Arthritis, Metabolism and Digestive Diseases; [3HJQNB, [�I-1Jquinuclidinyl benzilate.

462

0022-3565/83/2262-0462$02.00/OTHE JOURNAL OF PHARMACOLOGYAND EXPERIMENTAL THERAPEUtICS

Copyright 0 1983 by The American Society for Pharmacology and Experimental Therapeutics

SCH 23390, a Potential Benzazepine Antipsychotic with UniqueInteractions on Dopaminergic Systems

LOUIS C. IORIO, ALLEN BARNETT, FREDERICK H. LEITZ, VINCENT P. HOUSER and CHRYZANTA A. KORDUBA

Research OWision, Schering-Plough Corporation, Bloomfield, New Jersey

Accepted for pu�ation May 10,1983

ABSTRACT

SCH 23390 [R�+)-8-chIoro-2,3,4,5-tetrahydro-3-methyI-5-phe-nyl-1 H-3-benzazepine-7-ol) possesses pharmacologic effectssimilar to standard antipsychotics, including selective supressionof conditioned avoidance responding in rats and squirrel mon-keys, blockade of apomorphine-induced stereotypy in rats andblockade of methamphetamine-induced lethality in aggregatedmice. At effective doses in these tests, no changes in grossbehavior, neurological or autonomic function were observed. Incontrast to the standards tested, SCH 23390 blocked dopamine-stimulated adenylate cyclase at concentrations (lCso 0.01 �M)

about 2000 times lower than those needed to block spiperonebinding (ICso 24 SM). This suggests specific D1-receptor an-tagonism. Inability of SCH 23390 to cause hyperprolactinemia,considered to be a D�-receptor effect, is consistent with thishypothesis. SCH 23390 showed lower increases in dopamineturnover suggesting that the blockade of SCH 23390 may bemore specific for post- than presynaptic sites. Additional evi-dence for the selectivity of SCH 23390 among putative postsyn-aptic dopamine sites includes its lack of effect on apomorphine-induced hypothermia or emesis. Based on these results, it ispostulated that SCH 23390 is a selective D1-receptor antagonist.

Commonly used antipsychotic drugs such as the phenothia-

zines chlorpromazine and THD, the butyprophenone HLP and

the piperazinyldiazepine CLZ have been hypothesized to act by

blocking dopaminergic function in the CNS (Clement-Cormier

et at., 1974; Snyder et at., 1974; Bunney and Aghajanian, 1975;

Seeman and Lee, 1975; Berger et at., 1978; Barchas et at., 1978).

The major CNS side effects seen in the clinic with most of

these drugs, including acute extrapyramidal symptoms (tremor,

akathisia and rigidity) and tardive dyskinesia (involuntary

muscle movement primarily in the buccolingual area seen afterprolonged treatment), have also been attributed in part toantidopaminergic action (e.g., Clement-Cormier et at., 1974;

Snyder et al., 1974; Gerlack, 1977). CLZ appears to have a low

liability for producing antidopaminergic side effects in man

(Sayers and Amsler, 1977), a finding purported to be due to its

anticholinergic activity (Snyder et at., 1974) because anticho-

linergic treatment attenuates acute extrapyramidal symptoms

induced in man by antipsychotic drugs (Byck, 1975).The relationships cited above led us to search for potential

antipsychotic drugs that either do not act by an antidopami-

nergic action or have a more selective action. To accomplishthis, we used blockade of CAR as the primary and necessary

requisite for further interest, then conducted studies to evaluate

Received for publication December 17, 1982.

possible antidopaminergic effects. With this approach, we dis-

covered the anti-CAR effects of SCH 23390 [(R)-(+)-8-chloro-

2,3,4,5 - tetrahydro - 3 - methyl-5-phenyl-1H-3-benzazepin-7-ol]

(fig. 1). We then found that SCH 23390 had in vivo and in vitro

antidopaminergic effects markedly different compared with thestandard antipsychotic drugs which we tested. This report

summarizes the pharmacology of SCH 23390, showing that it

has antipsychotic potential with a unique profile of interactionwith various dopaminergic systems, consistent with the concept

that SCH 23390 is a specific D1-receptor antagonist.A preliminary account of portions of this work has been given

previously (Iorio et at., 1981).

Methods

Animals and treatments. Male CF1 mice, 20 to 25 g, and male

Sprague-Dawley CD rats, 130 to 250 g, obtained from Charles River

Breeding Laboratories, Inc. (Wilmington, MA), squirrel monkeys (750-

900 g) obtained from Primate Imports Corp. (Long Island, NY) and

beagles bred at Schering Laboratories (Lafayette, NJ) were used inthese studies. For oral and parenteral administration to all speciesexcept dogs, SCH 23390 as the hemimaleate was suspended in 0.4%

aqueous methylcellulose. Volumes (milliliters per kilogram) adminis-

tered orally were 20 for mice, 5 for rats and 2 for squirrel monkeys and

volumes for parenteral administration were 10 for mice and 1 for all

other species. In dogs, drugs were placed in capsules for oral adniinis-tration. Doses of SCH 23390 and all other drugs used in this study are

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

CI

Fig. 1. SCH 23390.

1983 Unique Profile of SCH 23390 on Dopamine Systems 463

expressed in terms of weight of the free base per kilogram of bodyweight.

Methamphetamine-induced aggregate toxicity in mice. Meth-amphetamine was used to produce lethality in groups of 10 mice housedin 11 x 26 x 13 cm plastic chambers. Test drugs were administered 30mm before i.p. injection of methamphetamine at 15 mg/kg, a dose that

typically killed at least 90% of the mice within 4 hr. The number of

deaths in each group was recorded 4 hr after methamphetamine admin-

istration. The ED�o (95% CL) of each test compound was determinedusing probit analysis (Litchfield and Wilcoxon, 1949).

Apomorphine-induced stereotypy in rats. A modification of the

method of Costall and Naylor (1975) was used. Groups of five rats, 225

to 275 g, were injected with apomorphine, 2 mg/kg s.c., 30 mm after

P.O. administration of test drug. Stereotypy was scored every 15 mmafter administration of apomorphine for a total of 75 mm (five read-

ings). The scoring system used was: 0, no stereotypy; 1, discontinuoussniffing 2, continuous sniffing; 3, continuous sniffing, discontinuous

biting, gnawing or licking-, and 4, continuous biting, gnawing or licking.In control groups, apomorphine-induced stereotypy usually peaked 15to 30 mm after dosing (scores were 3 to 4 in virtually all rats) and wascompletely absent 90 mm after dosing. In rats that received bothapomorphine and test drug, blockade of apomorphine effects wasconsidered present when a score of less than 3 was recorded at any of

the five readings. The percentage of rats showing blockade at each dosewas used to determine ED� (CL) using probit analysis (Litchfield and

Wilcoxon, 1949).CAR suppression in rats and squirrel monkeys. Rats were

required to jump onto a platform located 6�/4 inches above the grid floor

of an experimental chamber in response to a 5-sec tone to avoid a 10-second foot shock (0.6 mA). Each experimental session consisted of 20such trials presented at 30-sec intervals. A correct CAR occurredwhenever the rat jumped onto the platform during the tone (before

foot shock). An escape response occurred whenever the ratjumped ontothe platform during shock. A response failure is defined as the lack of

jumping to the platform during the 5-sec tone period and 10-sec shockperiod.

Groups of six to eight rats were trained on two consecutive days

(total of 40 trials). Rats that reached criterion on day 2 (correct CARS

on 16 or more of the 20 trials) were treated with either a test drug orvehicle on day 3. Suppression of CAR was analyzed statistically using

Student’s t test comparing the performance of drug-treated to vehicle-

treated rats. The MED for each drug is defmed as the lowest dosetested that significantly (P < .05) reduced avoidance responding.

For squirrel monkeys, a nondiscriminated-discriminated paradigmwas used. The squirrel monkeys could move freely on a grid floor thatcould be electrified and they could press a lever to avoid or escape

shock. The session began with a 20-sec period with no warning stimulus

(nondiscriminated period), followed by a 10-sec period with a tone(discriminated period) and a 2-sec shock period. A lever press made by

the animal during any segment of this schedule reset the interval tothe beginning ofthe 20-sec warning-free period. All animals were tested

5 days a week, receiving vehicle on Wednesdays and drugs on Thurs-days. Session length was 50 mm, beginning 30 mm after drug admin-

istration. Statistical comparisons were made between control and drug

sessions using Student’s t test. The MED for each drug, defined as thelowest dose tested that significantly (P < .05) reduced avoidanceresponding, was used to estimate relative potencies.

Adenylate cyclase studies. Rat corpus striata were homogenizedin 30 volumes of 2mM Tris-2mM ethylene glycol bis (fl-aminoethylether)N, N’-tetraacetic acid buffer. Fifty-microliter aliquots of thiswere added to 450 Ml of 80 mM Tris-6 mM MgC12 buffer containing 10mM theophylline, 0.2 mM ethylene glycol bis(fi-aminoethyl ether)N,N’-tetraacetic acid and appropriate concentrations of drugs. After a

15-mm incubation at 10#{176}C,ATP was added to a final concentration of1.5 mM, the mixture incubated for 30 mm at 37’C, then placed in

boiling water for 5 mm to stop the reaction. After centrifugation at2000 x g for 20 mis, an aliquot of the supernatant was analyzed forcyclic AMP content with the aid ofa cyclic AMP binding protein assay

kit (Diagnostic Products Corp., Los Angeles, CA) and a second aliquotwas analyzed for protein levels according to the method of Miller

(1959).

I3HjSpiperone binding. A modification of the method of Burt etat. (1975) was used. Tissues were homogenized in 50 volumes of cold

Tris buffer (0.05 M), at 50,000 X g in the cold. The resulting pellet was

rehomogenized in 10 volumes of Tris buffer. One hundred-microliteraliquots of this homogenate (containing the equivalent of 10 mg oftissue) were added to incubation tubes with appropriate amounts of

drugs in a final volume of 1 ml containing (in millimolars): NaCl, 120;

KCI, 5; CaCl2, 2; M�Cl2, 1; and 10 sM pargyline and 0.1% ascorbicacid. The tubes were incubated at 37’C for 10 mis, then placed on ice,when a b-gil aliquot of [3HJ-spiperone stock solution was added toeach tube to give a final concentration of 0.8 nM. The tubes were thenincubated for 10 mm at 3TC and the contents were filtered on GF/B

glass filters (Millipore, Inc., Bedford, MA) and quickly rinsed with two

4-ml aliquots of the incubation buffer. The filters were placed in aliquid scintillation counting vial with 10 ml of scintillation fluid (Scm-tisol, Isolab, Inc., Akron, OH) and the radioactivity content determinedin a liquid scintillation counter with internal standardization.

Dopamine synthesis studies. Dopamine synthesis rates were de-

termined as previously described by Bariletto et aL (1975). Rats were

injected i.v. with 100 �Ci [3,5-�HJtymsine (30-50 Ci/mmol), sacrificed15 mm later to obtain corpus striatum and olfactory tubercule tissues,

which were then homogenized in 0.4 M perchloric acid. Labeled catech-ols were isolated by adsorption and elution from short alumina columnsand the radioactivity measured in a liquid scintillation counter. Syn-thesis rates were calculated using the fraction of tritium present ascatechol compounds (disintegrations per minute per gram of catecholsdivided by disintegrations per minute per gram of total tritium) mul-

tiplied by 2 because one tritium atom is lost on conversion of [3,5-3Hjtyrosine to 3H-labeled catechol compounds. The resulting first order

rate constant was then multiplied by the tyrosine concentration (157

nmol/g in the corpus striatum and 171 nmol/g in the olfactory tuber-

cule, as determine by the method of Waakes and Udenfriend, 1957).

To correct for nonenzymatic formation of 3H-labeled catechols, ratswere given a-methyl-tyrosine methylester HC1 at 200 mg/kg i.p., a dose

that completely blocked tyrosine hydroxylase, 30 mm before [3,5-3H1

tyrosine and the amount of labeled catechols in the tissues of rats was

subtracted from all values before calculating synthesis rates.

Serum prolactin level studies. Rats were administered drugsorally and sacrificed by decapitation at selected times after dosing.Blood was collected and centrifuged to obtain serum. Levels of prolactin

in serum were determined by double antibody radioimmunoassay

method, using NIAMDD-Anti-Rat Prolactin serum and NIAMDD-RatProlactin RP-1 with a biological potency of 11 lU/mg as the standard,obtained from the NIAMDD. The minimum detectable quantity of

serum prolactin was 0.1 ng/ml. All data are expressed as nanogram per

milliliter of NIAMDD-Rat Prolactin RP-1 standard.Hypothermia studies in mice. Apomorphine-induced hypother-

mis in mice is blocked by standard neuroleptics (Schelkunov, 1968;Barnett et al., 1972), suggesting a dopaminergic mechanism. Studieswere conducted to determine whether SCH 23390 could antagonize thiseffect. The method of Barnett et a!. (1972) was used. Groups of five

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

464 lorlo et al. Vol. 226

vate serum prolactin levels.

male CF1 mice were housed in plastic cages (30 x 15 x 13 cm) in a

temperature-controlled room at 19.5-21.5’C for at least 1 hr before

taking body temperatures. Drugs were injected i.p. in a volume of 10ml/kg b.w. Body temperatures were taken with an Ellab Temperatureprobe inserted into the esophagus. A base-line temperature reading was

taken for each animal followed by i.p. treatment with vehicle, HLP(0.25 mg/kg) or SCH 23390 (1, 3 or 10 mg/kg). Thirty minutes later,vehicle or apomorphine was administered to all animals and a finaltemperature reading was taken 20 mm later.

Emesis studies in dogs. Apomorphine-induced emesis in dogs is

blocked by neuroleptics such as HLP. Studies were conducted in beagledogs (9-13 kg) to determine whether SCH 23390 had this activity. Dogswere injected with apomorphine at 0.8 mg/kg s.c., which causes emesisin all dogs within 5 mm. To evaluate blockade, vehicle (0.4% aqueous

methylcellulose suspension), HLP at 0.01, 0.3 and 1 mg/kg, or SCH23390 at 3 and 10 mg/kg was administered 30 mm before apomorphine,and the dogs were observed for emesis for 1 hr after apomorphine.

Oxotremorine-reduced tremors and salivation in mice. Amodification ofthe method of Everett (1964) was used. Test drugs wereadministered to groups of five mice 30 mm before administration of

oxotremorine at 0.5 mg/kg s.c., a dose that causes tremors and saliva-

tion in virtually all treated mice. Blockade ofthese effects was evaluated

15 mm after oxotremorine administration. Percent blockade of each

treated group was then used to determine the ED�o (95% CL) by probitanalysis (Litchfield and Wilcoxon, 1949).

[3H]QNB binding. Rat brains, less cerebellum, were homogenized

in 10 volumes of cold 0.32 M sucrose in a smooth glass homogenizer

fitted with a Teflon pestle. The homogenate was centrifuged at 1000 x

g for 10 mm, the supernatant diluted with an equal volume of 0.32 Msucrose and the resulting solution homogenized with a polytron (Brink-mann Instruments, Inc., Westbury, NY) in the cold. The incubationmedium was made up to contain 0.86 ml of 0.05 M sodium-potassiumphosphate buffer (pH = 7.4), 50 �d of tissue homogenate, 100 z1 of

phosphate buffer containing test drug and 60 �l of [3H]QNB stock

solution to produce a final concentration of0.6 nM. This was incubated

for 60 mm at 37#{176}C.Then 3 ml ofcold phosphate buffer were added andthe contents quickly filtered on GF/B Millipore glass fiber filters. Theincubation tubes were washed twice more with 3-ml aliquots of thephosphate buffer, each of which was also passed over the filters. Thefilters were placed in scintillation counting vials and 10 ml of 950 Ascintillator (New England Nuclear, Boston, MA) was added. Theradioactivity was determined in a liquid scintillation counter withinternal standardization ([3Hjhexadecane, New England Nuclear).

Results

Studies on antipsychotic potential. Effects of SCH 23390and selected standard antipsychotics in four procedures thought

to reflect antipsychotic activity are summarized in tables 1 and2. Administered orally to rats, SCH 23390 caused dose-related

suppression of CAR (table 1). Suppression was specific in thatthe rats responded to shock by escaping and only at high doses

and/or early time points (1 hr after 10 mg/kg, 0.5 hr after 20mg/kg) were any failures to escape noted. Similar effects wereseen with CLZ except that SCH 23390 was about 4 to 8 timesmore potent when the two drugs were compared at 2 to 3 hrafter dosing. SCH 23390 and CLZ had comparable time courses:activity peaked within 1 hr after oral administration and lasted2 to 3 hr at threshold doses. Significant activity of SCH 23390was still evident 6 hr after administration of 20 mg/kg p.o., adose 4 times the MED. This dose-duration profile differed from

those seen after HLP, THD and perphenazine, which exhibitedpeak activities at least 4 hr after oral administration. As shownin table 1, the 4-hr MEDs for these drugs are lower than the 1-

hr MEDs. Figure 2 shows that SCH 23390 administered s.c.caused specific dose-related suppression of CAR at doses as low

as 0.009 mg/kg s.c. This compares to the MED after oraladministration of 5 mg/kg p.o.

In squirrel monkeys, SCH 23390, like HLP and CLZ, shifted

responding from the nondiscriminated period to the discrimi-

nated period and/or shock period as indicated by increases in

escape responses or number of shocks taken. The MEDs forthese three drugs in squirrel monkeys were similar.

SCH 23390 was also active in antagonizing apomorphine-

induced stereotyped behavior in rats. HLP was significantlymore potent (table 2), whereas THD and CLZ did not blockapomorphine-induced stereotypy even at the relatively high

dose of 100 mg/kg p.o.SCH 23390, like standard antipsychotics, caused dose-related

blockade of methamphetamine-induced lethality of grouped

mice. It was more potent than CLZ, equipotent to THD andchlorpromazine and less potent than HLP or perphenazine inthis test (table 2).

Effects on dopamine-stimulated adenylate cyclase.Tested for ability to block dopamme-stimulated adenylate cy-clase in homogenates of rat corpus striatum, SCH 23390 was

found to be the most potent drug tested, about 10 times more

potent than perphenazine and 100 times more potent than CLZ

or THD (table 3). The results with the standard antipsychoticdrugs are consistent with data published by Clement-Cormier

etat. (1974).

Effects on [3lljspiperone binding to rat striatal mem-branes. Antipsychotics are potent inhibitors of radiolabeledHLP binding (Seeman et at., 1976). Tested for this property,

but using radiolabeled spiperone instead of HLP because of itsgreater specificity (Lauduron et at., 1977), SCH 23390 had onlyweak inhibitory effects (table 3), about �4io to Moo the potencies

of the weakest reference standards, CLZ and THD. Moreover

the ICso for SCH 23390 in the test was more than 2000 times

its ICre for inhibiting dopamine-stimulated adenylate cyclase.All of the other antipsychotics tested were equiactive or more

potent in displacing spiperone than in inhibiting adenylate

cyclase.Effects on dopamine synthesis rate. Carlson and Lind-

quist (1963) proposed that antipsychotics act by blocking cen-tral dopamine receptors and that this leads via feedback to anincreased dopamine synthesis rate. In these studies, SCH

23390, like CLZ, THD and HLP, caused dose-related increasesin dopamine synthesis rates in the rat corpus striatum and

olfactory tubercle (table 4). SCH 23390, CLZ and THD wereapproximately equipotent and had about #{189}oto l’4oo the potency

of HLP. However, synthesis rates after the higher doses ofCLZ, THD and HLP were increased about 200% in the striatumand 80 to 100% in olfactory tubercle, whereas SCH 23390increased rates less than 60% in both of these tissues.

Effects on rat serum prolactin levels. Drugs that block

dopamine receptors in the anterior pituitary are known to

elevate serum prolactin levels in man as well as in the rat(Meltzer et at., 1978). SCH 23390 was compared with CLZ,THD and HLP for this activity (table 5). Our data confirm the

earlier report (Meltzer et at., 1978) that these three antipsycho-

tics significantly elevate serum prolactin levels in rats. These

effects occurred at doses that were either at (CLZ) or below(HLP and THD) the minimal effective dose of each drug for

suppression of CAR in rats. In contrast, SCH 23390 at a widerange of doses (up to 6 times its MED for CAR suppression)

and times (30-240 mm after dosing) did not significantly ele-

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

Comparison of SCH 23390 and SeleCted antipsychotic drugs for block of CAR in rats and squirrel monkeys,lethalfty in grouped mice (METh-Tox) and apomorphlne’ � � . stereotypy in rats (APO-stereotypy)

DrugCAR, Rats MED’ CAR, Monkeys

�

*P0�motypy

�b�%% �)METH40x

�C(95% Q)1 Pr 4 tr

mg/kgp.o. ir#{231}/kqp.o. rr#{231}/kgp.o. irvj/kgp.o.

SCH 23390 5 20.0 1 .3 2.1 (1 .0-4.2)” 2.5(1.6-3.8)

HLPPerphenazineTHDGil

2.51 0

>16040

0.62.5

4080

1

5

0.6 (0.4-0.9r0.9 (0.4-1 .3�

>�oof�

>100’�

0.1 (0.05-0.2)’0.3(0.05-0.5)’5.0(3.2-7.6)

17.0(10.1-28.7)’

a The lowest dose used that signfficantly (P < .05) suppressed CAR determined at 1 and 4 hr after treatment in at least three groups of five rats each and 30 to 60mm after drug treatment in at least six squirrel monkeys and three rhesus monkeys.

b That dose (95% CL) that blocks apOmOrphine-StereOtypy in 50% of treated rats, determined 4 hr after oral adminiStratiOn of test drugs. At least four groups of liverats each were used to obtain dose-response curves for each drug.

C That dose (95% CL) that blocks methamphetamlne-induced lethality in 50% of grouped mice. Methamphetamine was adn*ustered 30 mm after test drugs to at leastfour groups of 10 mice for each drug and deaths were counted 4 hr later.

d Determined 1 hr after drug treatment.a Determined 4 hr after drug treatment., Significantly (P < .05) less potent in this test than SCH 23390.a ietermined both 1 and 4 hr after drug treatment.S Not tested.

I Significantly (P < .05) more potent in this test than SCH 23390.

1983 Unique Profile of SCH 23390 on Dopamine Systems 465

TABLE 1

Dose-response and dose-duration relationship. for SCH 23390 and CLZ in blocking CAR In rats

SCH � Cu

Dose line sterv� % CAR % Esc�es % F�Iures Dose Tine sterv� % CAR % Esc�es % F�Iures

mg/kgp.o. hr mg/kgp.o. Ii

5.0 0.5123

39.0

53.4e

71 .681.5

57#{149}5b

378L�

27219.8

3.5

8.8

1 .20.7

20 123

86.371 .5’76.5’

13.723.5b23.5b

05.00

10 0.5123

21.631 .4’68.884.4

67.5b42.9k’27.5b15.6

10.925.7

3.70

40 1234

6

63.1

63.162.580.082.8

36.2”

33�7b2OAY’20.017.1

0

3.217.5

00.1

20 0.5

1246

20.027.5k40.754.558.6

41 7b60.9k’53.6b43.lb4�7b

3S.3z�11.6

5.71.80.7

12346

25.638.564.353.157.1

45.6b44235�6b

34�3L�40.7b

28.8k’17.3

0�112.6

2.2

. Statistically significantly decrease (P < .05, Student’s t test)oompared with vehicle-treated rats (CARs ranged from 86-92% Of total responses). Data were obtainedusing different groups of six to eight rats for each dose at each time point. Data obtained with SCH 23390 at 2.5 mg/kg p.o. and CLZ at 10 mg/kg p.o. are notsignificantly different (P < .05) from controls and hence not presented here.

b Statistically significant increase (P < .05, Student’s t test) compared with vehicle-treated rats (escapes ranged from 8-14% and failures ranged from 0-3%).

TABLE 2

�inaucea

Effects on apomorphine-induced hypothermia in mice.As previously reported (Barnett et at. 1972), apomorphine, 2

mg/kg i.p., significantly (P < .05) reduced body temperature of

mice, an effect which was completely blocked by HLP, 0.25

mg/kg. In contrast, SCH 23390 did not significantly reduceapomorphine-induced hypothermia at any of the doses tested

(1-10 mg/kg i.p.). Thus, SCH 23390 did not block apomorphineat doses at least 40 times greater than those of HLP (table 6).

Effects on apomorphine-induced emesis in beagles.SCH 23390 did not block apomorphine-induced emesis in bea-

gles at 3 or 10 mg/kg p.o. (table 7). At these doses, the dogs

manifested moderate CNS depression including decreased

alertness, decreased spontaneous motor activity, increased pas-

sivity and ataxia. In contrast, HLP completely blocked emesisat doses as low as 0.3 mg/kg p.o., which also caused CNS

depression similar to that seen with SCH 23390.

Tests for anticholinergic activity. SCH 23390 at doses

up to 30 mg/kg p.o., 12 times its ED� for blockade of meth-amphetamine-induced lethality in mice (table 2), did not have

anticholinergic activity in mice as measured by block of oxotre-morine-induced tremors or salivation. At the dose of 30 mg/kgP.O., a significant reduction in spontaneous motor activity wasobserved, confirming adequate absorption in this species. Inthis test, scopolamine had ED� values (95% CL) of 2.9 (1.6-5.1) and 2.8 (1.9-4.1) mg/kg p.o., respectively, against tremorsand salivation.

SCH 23390, like HLP, exhibited very weak displacement of[3HIQNB binding to both corpus striatum [IC�o (95% CL)95.2 (57.7-179.7 SM)] and olfactory tubercule [IC� (95% CL)= 68.2 (22.3-298.3)] membranes of rats. The standards THD,CLZ and chlorpromazine were about 200 to 300 times morepotent as displacers. The IC� values for these three drugs inboth tissues ranged from 0.1 to 1.3 �M. There were no signifi-cant differences in potency between [3H]QNB displacement forthe corpus striatum vs. olfactory tubercule for any of thesedrugs.

DiscussionSCH 23390 possesses a variety of pharmacologic effects sim-

ilar to those seen with standard antipsychotics, including selec-

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

CONTROL AVOIDANCESTABLE 4

80

60 -

*

C/,

U,z00.U,w

zwC.,

l�J0.

RATS

40 -

DANCES

20 -

FAILURES � a

� � ___#{149}...-�+._____#{149}#{149} I

0.005 0.009 0.02 0.04

DOSE. mg/kg s.c.

0.08

a vaiues are means ± SE. of three to four observations per point. Data wereanalyzed by comparing treated groups with corresponding vehicle-treated groupswith an analysis of variance using Duncan’s multiple range statistic.

a Statistically significant(P < .05) increase compared with vehicle-treated group.

tive suppression of avoidance responding in rats and squirrel

monkeys, dose-related blockade of methamphetamine-induced

lethality in aggregated mice and blockade of apomorphine-

induced stereotypy in rats. There were no changes in gross

behavior, neurologic or autonomic functions at effective dosesin any of the three species used in these procedures.

The most novel aspect of the pharmacology of SCH 23390 is

that it differs markedly from the antipsychotic drugs CLZ,THD and HLP with respect to interaction with dopaminergicsystems. SCH 23390 is approximately 2000 times more potentin blocking dopamine-stimulated adenylate cyclase than dis-

placing [3Hjspiperone binding, whereas the reference standards

are approximately equipotent in these tests. These results

466 lorio et al. Vol. 226

Fig. 2. Dose-response relationship of SCH 23390 on CAR in ratsmeasured 30 mm after s.c. administration. Each point represents themean number of avoidance/failures for a group of at least six rats.* Significant differences (P < .05, Student’s t test) from control values.

TABLE 3Comparison of SCH 23390 and selected antipsychotics for block ofdopamine-stimulated adenylate cyclase (DA-cyclase) and [‘H]spiperone binding (SPIP-binding)

DA-cyclase SP?P.bindlngDrug �

,�M

SCH 23390 0.01 1 (0.004-0.030) 23.79 (7.16-76.13)Perphenazine 0.115c(0.014�1.01) -.

CLZ 1 �59C (0.286-7.95) 0.79�’ (0.57-1.08)

THD 1 .88’ (0.632-6.24) 0.23d (0.10-0.55)HLP 7.17c (1.06-59.09) 0.lld (0.07-0.18)

Spiperone -. 0.02” (0.01-0.03)

a �eterm�nei by linear regression analysis, that concentration (95% CL) neces-

sary to block by 50% DA.cyclase. The DA concentration used was 100 �M. Theapparent Km for dopamine in stimulathg the adenylate cyclase was 1 .16 � andin the presence of 0.1 ,�M SCH 23390 was 177.2 ,iM; this gave a dlesocia�onconstant (l�) for SCH 23390 for inhibiting cyclase of 0.66 nM.

b Determined by linear regression analysis. that concentration (95% CL) neces-sary to block by 50% SPIP-binding. The dissociation constant (Ks,) for spiperonewas 0.499 nM and the concentration used was 0.8 nM.

C Significanfly (P < .05) higher lC�, compared with that of SCH 23390.d � (P < .05) lower K� compared with that of SCH 23390.

a Not tested.

Effects of SCH 23390, ThD, CLZ and HIP on doparate In rat corpus striatum and olfactory tUberCie

mine synthesis

Mean Synthesis RateTreatment Dose

+ S.E.

Corpus st#{241}atum 0�ory tuberde

mg/kgp.o. nmol/g/hr

Vehicle 39.4±2.6 41.6± 5.4SCH23390 10 45.1 ±0.5b 50.3±2.2

30 60.7 ± 1 5b 66.9 ± 3.9w100 61.7 ± 5�9b 61.5 ± 55b

Vehicle 29.1 ± 3.2 35.8 ± 1.6

CLZ 20 56.1 ± 8.0” 53.8 ± 1.3#{176}40 66.9±9.8#{176} 67.6±7.4#{176}

60 84.0 ± 3.4#{176} 64.5 ± 4.6#{176}Vehicle 35.0 ± 2.3 46.7 ± 2.8THD 10 67.1 ± 7.9 68.8 ± 7.8#{176}

30 96.4 ± 4.9#{176} 73.9 ± 6.9#{176}

100 106.4±1.6#{176} 95.0±6.0#{176}Vehicle 31.9±3.3 44.2±2.5

HLP 0.1 45.2 ± 1.4#{176} 43.1 ± 7.90.3 83.2 ± 4.8#{176} 68.4 ± 2.91.0 114.2 ± 1.6#{176} 84.2 ± 8.4

TABLE 5Effects of SCH 23390 and selected antipsychotic drugs on ratserum prolactin levels

Drug 0r� Dose Mean ± S.E.

mg/kg mm ng/mI serum

(Vehicle) (30-240) (2-12)CLZ 20.0

40.080.0

303030

7.7 ± 2.823.3 ± 4.8#{176}27.8 ± 4.5#{176}

THD 20.040.080.0

606060

14.0±2.0#{176}

32.0±6.1#{176}28.3 ±4.0#{176}

HLP 0.61.252.50

303030

7.4±1.349.2±12.5#{176}80.1 ± 12.9#{176}

SCH 23390 10.030.0

2.55.0

10.030.010.030.010.030.0

30306060

6060

120120240240

18.4 ± 4.59.5 ± 2.8

16.8±4.511.5±3.112.1±2.610.3 ± 2.2

3.1±1.23.1 ± 0.54.6 ± 2.0

5.6 ± 2.8a Vaiues were obtained in groups of six rats for each determination. Results in

this table are combined from several experiments. The range of serum prolactinlevels of Vehicle-treated groups at any time point or in any experiment was 2 to 12ng/ml of serum.

a Significant increase (P < .05, Student’s t test) obtained by companng datawith appropriate vehicle control data Obtained on the same day in the sameexperiment, e.g., the mean (±S.E.) concentration of prolactin in the serum 30 mmafter those experiments in which CLZ was evaluated was 10.3 (3.0) ng/ml ofserum.

suggest that SCH 23390 might be a relatively specific antago-nist at dopamine receptors associated with adenylate cyclase,termed D1-receptors (Kebabian and Calne, 1979), with weakactivity at D2-receptors for which butyrophenones like spipe-rone and HLP (Seeman et at., 1976; Creese et a!., 1975) have ahigh affinity. This hypothesis is directly supported by therecent work ofDr. Hyttel and colleagues at Lundbeck Company

(personal communication) showing that SCH 23390 is a more

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

1983 Unique Profile of ScH 23390 on Dopamine Systems 467

TABLE 6Effects of S�H 23390 and HLP on apomorphine

hypothermia In mice

Treatment r Dose Treatment 2’ Dose

mg/kgi.p. mg/kg i.�

Vehicle Vehicle 37.2 ± 0.8Vehicle Apomorphine 2 33.9±1.1#{176}

HLP 0.25 Vehicle 37.4 ± 0.9HIP 0.25 Apomorphine 2 36.8 ± 0.9c

SCH23390 10 Vehicle 36.3±0.6SCH 23390SCH23390

SCH 23390

13

1 0

ApomorphineApomorphineApomorphine

222

35.0±0.8#{176}35.2±1.8#{176}34.0 ± 1 �7d

a Each treatment group contained five mice. Treatment 2 was administered 30

rdfl after treatment 1 and measurements were made 1 hr later.a Statistically significant decrease (P < .05. Student’s t test) compared with

vehide-vehide controls.C StatiStiCally significant increase (P < .05. Student’s t test) compared with

vehicle-apomorphine treated group.d Not significantly different (P > .05, Student’s t test) from vehide-apomorphine

treated group.

TABLE 7

Failure of SCH 23390 to block apomorphine-Induced emesis indogs

Treatment’ Dose No. Treated No. with Emes�

mg/kgp.o.

Vehicle 6 6HLP 0.01

0.03#{176}0.1#{176}

444

400

SCH23390 3.0#{176}10.0#{176}

46

45

a ���phine at 0.8 mg/kg s.c. was used to evoke emesis. Treatments were

given 30 miii before and dogs were observed for 1 hr after apomorphine adminis-tration.

b Moderate CNS depression was observed at these doses.

potent displacer binding of [cis-3H]-(Z)-piflutixol, a specific D1-receptor ligand (Hyttel, 1981), than [3Hjspiperone binding torat striatal membranes. Our data showing that SCH 23390 does

not cause hyperprolactinemia is also consistent with this hy-pothesis as prolactin secretion is thought to be mediated

through D2-receptors which are not linked to adenylate cyclase

(Kebabian and Calne, 1979).Because SCH 23390 is a relatively good blocker of apomor-

phine-induced stereotypy, but is less potent and causes smaller

increases in dopamine turnover than standard antipsychotics

tested, it seems likely that it is more specific for postsynaptic

than presynaptic receptors. This suggests that postsynapticsites are of the D1 type, although blockade at D2 receptor sitescannot be ruled out because SCH 23390 probably also has weakantagonistic activity at these sites. Similarly, of one assumes

that apomorphine hypothermia and emesis are mediated

through postsynaptic dopamine receptors, these receptorsshould be either D2 receptors or a D1-receptor subtype as SCH

23390 does not block these measures at doses higher than those

that block methamphetamine-induced aggregate toxicity andapomorphine-induced stereotypy.

From the standpoint of side-effect potential, it is not known

to what extent a drug like SCH 23390, which manifests selectiveD1-receptor antagonism with lower increases in dopamine turn-

over, might cause extrapyramidal side effects in man. However,this drug at doses up to 300 mg/kg p.o. in rats did not causecatalepsy (data not shown), a behavior usually found with

existing antipsychotic drugs and considered to reflect extrapyr-

amidal liability. If the lack of catalepsy in rats is due to lack ofreceptor blockade at sites other than D1-receptors, it may well

be that SCH 23390 will have less potential to cause extrapyr-amidal side effects. In addition, SCH 23390, unlike other anti-psychotics, should not cause hyperprolactinemia. Its inabilityto block [3H]QNB binding or oxotremorine-induced tremors/salivation and the absence of mydrasis in rodents at doses upto 300 mg/kg p.o. (data not shown) suggest that side effectssuch as dry mouth are unlikely.

In summary, this work shows that SCH 23390 has selectiveactivity in procedures predictive of antipsychotic potential,with a low liability for hyperprolactinemia or anticholinergicside effects. Studies showing its differential blockade of dopa-mine-stimulated adenylate cyclase over [3Hjspiperone binding

suggest that, unlike standard antipsychotics, it has selectiveaffinity for D1 us. D2 receptors. This might lead to a low liabilityfor extrapyramidal effects in humans.

Acknowledgments

The authors wish to thank Dr. Joel Berger and Mr. Wei Chang for thesynthesis of SCH 23390, James Flynn, Steven Bariletto and Dr. John Veals fortheir expert technical assistance; and Vera Brown for her secretarial skills inpreparation of this manuscript.

References

BARCHAS, J. D., BanGER, P. A., MATFHY5SE, S. AND WYA’I’r, R. J.: Thebiochemistry ofaffective dimMers and schizophrenia. In Principles of Psycho-pharmacology, ed. by W. G. Clark and J. del Guidice, 2nd ed., pp. 105-131,Academic Press, New York, 1978.

BAmLrrro, S., DOLLAR, E. AND LEITZ, F.: Effects of ainantadine on the rate ofdopamine synthesis in rat corpus striatum. J. Neurochem. 25: 139-142, 1975.

BiutNz’rr, A., GOLDSTEIN, J. AND TAsna, R. I.: Apomorphine-induced hypother-mis in mice: A possible dopaminergic effect. Arch. mt. Pharmacodyn. Ther.198: 242-247, 1972.

BERGEn, P. A., ELLIOTF, G. R. AND BARCHAS, J. D.: Neuroregulators andschizophrenia. In Psychopharmacology, a Generation of Progress, ed. by M. A.Lipton, A. DiMascio and K. F. Killam, pp. 1071-1082, Raven Press, New York,1978.

BUNNEY, B. S. AND AGHAJANIAN, G. K.: Antipsychotic drugs and central dopa-minergic neurons: A model for predicting therapeutic efficacy and incidence ofextrapyramidal side effects. In Predictability in Psychopharmacology Preclin-ic:al and Clinical Correlations, ad. by A. Sudilovaky, S. Gershon and B. Beer,pp. 225-245, Raven Press, New York, 1975.

BuRT, D. R., ENNA, S. J., Cazasa, I. AND SNYDER, S. H.: Dopamine receptorbinding in the corpus striatum of mammalian brain. Proc. Natl. Acad. Sci.U.S.A. 72: 4655-4659, 1975.

BYCK, R: Drugs and the treatment of psychiatric disorders. In The Pharmaco-logical Basis of Therapeutics, ed. by L S. Goodman and A. Gilman, pp. 152-200, New York, Macmillan Publishers Co. Inc., 1975.

CARLSON, A. AND LINDQUIST, M.: Effect of chiorpromazine or haloperidol onformation of 3-methoxytyramine and normetanephrine in mouse brain. ActsPharmacoL ToxicoL 20: 140-144, 1963.

CLEMENT-CORMIER, Y. C., KEBASIAN, J. W., PETZOLD, G. L. AND GREENGARD,P: Dopamine-sensitive adenylate cyclase in mammalian brain: A possible siteof action of antipsychotic drugs. Proc. Natl. Aced. Sci. U.S.A. 71: 1113-1117,1974.

COSTALL, B. AND NAYLOR, R. J.: Detection of the neuroleptic properties ofclozapine, sulpiride and thioridazine. Psychopharmacologia 43: 69-74, 1975.

CREasE, I., Bu�rr, D. R. AND SNYDER, S. H.: Dopainine receptor binthngDifferentiation of agonist and antagonist states with I3HIdopa’nine and (‘HIhaloperidol. Life Sci. 17: 993-1002, 1975.

EvEnz’rr, G. M.: Animal and clinical techniques for evaluating anti-parkinsonagents. In Animal and Clinical Pharmacologic Techniques in Drug Evaluation,

ed. by J. H. Nodine and P. G. Siegler, pp. 359-368, Chicago, Year Book MedicalPublishers, Inc., 1964.

Gaiu�ci, J.: The relationship between parkinsonism and tardive dyskenesia.Am. J. Psychiatry 134: 781-784, 1977.

HrrrEL, J.: Similarities between the binding of ‘H-piflutixol and ‘H-flupentixolto rat striatel dopamine receptors in vitro. Life Sci. 28: 563-569, 1981.

Ioiuo, L C., Housan, V., KoiwunA, C. A., Larrz, F. AND BARNrrr, A.: SCH23390, a benzazepine with atypical effects on dopaminergic systems. Pharina-cologist 23: 137, 1981.

KEBABIAN, J. AND CALNE, D. B.: Multiple receptors for dopamine. Nature (Lond.)277: 93-96, 1979.

LAUDURON, P. M., JAN5SEN, P. F. M. AND LEY5EN, J. E.: Spiperone: A ligandofchoice for neuroleptic receptors. Biochem. Pharmacol. 27: 317-321, 1977.

LITCHFIELD, J. T., JR. AND Wn�coxoN, F.: A simplified method of evaluatingdose-effect experiments. J. Pharmacol. Exp. Ther. 96: 99-1 13, 1949.

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

468 lorlo et al Vol. 226

MELTZER, H. Y., FANG, V. S. AND GOODE, D. J.: The effect of neuroleptics and SEaMAN, P., LEE, T., CHAU-WONG, M., TEDE8CO, J. AND WONG, K.: Dopaminealpha-methyl-para-tyrosine on serum prolactin levels in laboratory animala y#{248}�ep� in human and calf braina, using (‘H) apomorphine and an antipsy-and man. Psychopharmacol. Bull. 14: 5-7, 1978. chotic drug. Proc. Natl. Aced. Sci. U.S.A. 73: 4354-4358, 1976.

MILLER, G. L: Protein determination for large numbers of sample. Anal. Chem. Siwosn, S. H., BANERJEE, S. P., YAMAMURA, H. I. AND GREENBERG, D. A.:31: 964, 1959. Drugs, neurotransmitters and schizophrenia. Science (Wash. D.C.) 184: 1243-

SAYERS, A. C. AND AMSLER, H. A.: Clozapine. In Pharmacological and Biochem- � 1974.ical Properties of Drug Substances, ed. by M. E. Goldberg, pp. 1-31, American � i’. i. AND UDENFRIEND, S.: A fluorometric method for the estimationPharmaceutical Association Academy ofSciences, Washington, D.C., 1977. oftyrosine in plasma and tissues. J. Lab. Clin. Med. 50: 733-736, 1957.

SCHELKUNOV, E. L: Pharmacological effects of apomorphine in mice as the testfordifferentiationofantidepresaants,cholinolytics andneuroleptics. FarmakoL _______________________________________________TokaikoL 31: 559-563, 1968.

SEaMAN, P. AND LEE, T.: Antipsychotic drugs: Directcorrelationbetweenclinical �#{176}‘� reP�� requests to: Louis C. Iorio, Ph.D., Department of Pharmacology,potency and presynaptic action in dopamine neurons. Science (Wash. D.C.) SChering Corp., 60 Orange St., Bloomfield, NJ 07003.188: 1217-1218, 1975.

at ASPE

T Journals on January 5, 2015

jpet.aspetjournals.orgD

ownloaded from

本文献由“学霸图书馆-文献云下载”收集自网络，仅供学习交流使用。

学霸图书馆（www.xuebalib.com）是一个“整合众多图书馆数据库资源，

提供一站式文献检索和下载服务”的24 小时在线不限IP

图书馆。

图书馆致力于便利、促进学习与科研，提供最强文献下载服务。

图书馆导航：

图书馆首页 文献云下载 图书馆入口 外文数据库大全 疑难文献辅助工具