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Clozapine: Selective Labeling of Sites Resembling 5HT6 Serotonin Receptors May Reflect Psychoactive Profile
Molecular Medicine volume 1, pages 398–406 (1995)
Clozapine, the classic atypical neuroleptic, exerts therapeutic actions in schizophrenic patients unresponsive to most neuroleptics. Clozapine interacts with numerous neurotransmitter receptors, and selective actions at novel subtypes of dopamine and serotonin receptors have been proposed to explain clozapine’s unique psychotropic effects. To identify sites with which clozapine preferentially interacts in a therapeutic setting, we have characterized clozapine binding to brain membranes.
Materials and Methods
[3H]Clozapine binding was examined in rat brain membranes as well as cloned-expressed 5-HT6 serotonin receptors.
[3H]Clozapine binds with low nanomolar affinity to two distinct sites. One reflects muscarinic receptors consistent with the drug’s anticholinergic actions. The drug competition profile of the second site most closely resembles 5HT6 serotonin receptors, though serotonin itself displays low affinity. [3H]Clozapine binding levels are similar in all brain regions examined with no concentration in the corpus striatum.
Besides muscarinic receptors, clozapine primarily labels sites with properties resembling 5HT6 serotonin receptors. If this is also the site with which clozapine principally interacts in intact human brain, it may account for the unique beneficial actions of clozapine and other atypical neuroleptics, and provide a molecular target for developing new, safer, and more effective agents.
Clozapine is an important therapeutic agent in treating schizophrenia. Though developed as a neuroleptic, it is unique in its therapeutic profile and may provide major benefits to patients who are resistant to other neuroleptics (1,2). Even in patients who respond to conventional neuroleptics, clozapine may be more efficacious (2). Clozapine appears to relieve negative symptoms, including apathy and emotional withdrawal, that resist conventional neuroleptics and displays a very low incidence of extrapyramidal side effects (EPS). Diminished EPS and greater therapeutic efficacy are also manifested by more recently developed atypical neuroleptics, including risperidone, olanzepine, seroquel, and sertindole (2).
Therapeutic actions of conventional neuroleptics correlate closely with their potencies in blocking dopamine D2 receptors, which presumably explains their therapeutic and EPS actions (3–5). Imaging D2 receptors in humans by positron emission tomography reveals less occupancy of D2 receptors by atypical than conventional neuroleptics, when administered at clinically effective doses (6). The lesser incidence of EPS associated with the atypical drugs may also reflect their greater anticholinergic potencies, as muscarinic anticholinergic drugs are well known to relieve Parkinsonian, EPS symptoms (7,8).
To explain the greater antischizophrenic actions of atypical neuroleptics, researchers have evaluated effects of these drugs at novel sites. Thus, clozapine displays uniquely high affinity for dopamine D4 receptors (9). Serotonin 5HT2 receptors have also been implicated. Ritanserin, a selective 5HT2 antagonist, decreases EPS elicited by haloperidol (10). Mianserin, also a 5HT2 antagonist, relieves negative symptoms in schizophrenics receiving conventional neuroleptics (11). Compared with conventional neuroleptics, atypical drugs tend to have higher affinities for 5HT2 than D2 receptors (12). However, there are exceptions such as chlorpromazine and amoxapine, both more potent at 5HT2 than D2 sites (13).
To assess clozapine actions at various receptors, most studies have evaluated clozapine’s potency in competing for the binding of radioligands. However, ligands exert “induced fit”, altering receptor conformation so that the potency of a drug in competing for binding of a radioligand may not faithfully reflect that drug’s propensity to bind to the receptor in vivo. Ideally, one would like to specify the receptors to which clozapine binds when encountering the human brain in vivo.
To evaluate sites to which clozapine binds preferentially, we have examined the binding of [3H]clozapine itself to rat brain membranes. We report labeling of muscarinic cholinergic receptors as well as sites resembling 5HT6 serotonin receptors.
Materials and Methods
All unlabeled drugs were obtained from Research Biochemicals International (Natick, MA, U.S.A.). [3H]Clozapine was generously provided by Dr. S. Hurt NEN-Dupont (Boston, MA, U.S.A.).
Preparation of Rat Brain Membranes
Sprague-Dawley rats (200–300 g) were killed by decapitation. Brains were rapidly removed and specific brain regions dissected. Tissue was homogenized by Polytron in 50 mM Tris HCl, pH 7.4, at 4°C. Homogenates were centrifuged at 48,000 × g for 10 min. Pellets were resuspended and rehomogenized in the same buffer and centrifuged a second time at 48,000 × g for 10 min. Pellets were resuspended in 50 mM Tris HCl, pH 7.4, to a concentration of 15 mg/ml. Except in regional dissection studies, membranes from whole rat brain minus cerebellum were employed for binding assays.
Preparation of Membranes from 5HT6 Receptor Expressing Cells
Human embryonic kidney 293 cells stably expressing rat 5HT6 receptors were grown in D-modified Eagle’s medium with 10% FBS, 1 mM sodium pyruvate, and G418 at 300 µg/ml. Cells were grown to confluence, rinsed with 5 ml 0.5 mM EDTA in phosphate buffered saline (PBS). Cells were then washed for 5 min with an additional 5 ml EDTA/PBS. Cells were released by this treatment and treated as above for brain membranes. Final membrane concentration was ∼10.0 mg/ml protein. These cells express approximately 800 fmol/mg protein of 5HT6 receptor binding activity (14).
[3H]Clozapine Binding Assays
Binding assays were performed in a final volume of 500 µl Tris HCl, pH 7.4. Membrane preparation (0.25 ml) was added to each tube. Compounds for competition were added in a 50-µl volume. Fifty microliters of a 2% bovine serum albumin (BSA) solution was added to reduce nonspecific filter binding. [3H]Clozapine (specific activity 51.3–89.1 Ci/mmol) in 50 µl was added to give a final concentration of 1 nM. For all experiments performed in the presence of scopolamine, 100 µl was added to give a final concentration of 10 nM. For the initial experiments without scopolamine, 100 µl of distilled water were used. Nonspecific binding was determined by addition of 1 µM (final concentration) unlabeled clozapine. Tubes were incubated for 20 min at 37°C. The incubation was terminated by rapid filtration over 0.5% polyethyleneimine-soaked filters (GF/B) and washed 2× 3 ml with ice-cold 50 mM NaCl.
As an initial screen, we evaluated inhibition of [3H]clozapine binding by agents acting at sites where clozapine is thought to exert effects (Table 1). Scopolamine and atropine are uniquely potent with IC50 values of 0.8 nM and 1.0 nM, respectively. Scopolamine maximally inhibits about 60% of [3H]clozapine binding at 5 nM (Fig. 1). Accordingly, in all subsequent experiments we include 10 nM scopolamine so that hereafter [3H]clozapine binding will refer to binding measured in the presence of scopolamine.
[3H]Clozapine binding is saturable with a Kd of 4.5 nM and Bmax of 380 fmol/mg protein (Fig. 2a). Scatchard analysis of [3H]clozapine binding as well as of a displacing curve of unlabeled clozapine competing for [3H]clozapine binding reveals a single binding site with a Kd of 6.7 nM and a Bmax of 240 fmol/mg protein (Fig. 2b). Since dopamine receptors are most highly concentrated in the corpus striatum, we compared [3H]clozapine binding in the striatum, hippocampus, and cerebral cortex (Table 2). The Kd’s are similar in all areas, suggesting similar binding sites, and the Bmax values are roughly equal, suggesting a regional distribution of [3H]clozapine binding that does not fit with a classical dopamine D1 or D2 receptor.
The relative potencies of numerous drugs in inhibiting [3H]clozapine binding provide suggestions about the types of receptors that are most likely involved (Table 1). Triprolidine, a classic histamine H1 antagonist, displays an IC50 of 1 µM, almost 1000 times higher than its affinity for histamine H1 receptors. The potent α-1 adrenoceptor antagonist prazosin has an IC50 greater than 10 mM. Similarly, agents active at sigma, GABA, and glutamate receptors display low affinity for [3H]clozapine sites.
Clozapine is well known to influence dopamine receptors with an IC50 for D1 and D2 receptors of 172 nM and 182 nM, respectively (3). Accordingly, we compared the potencies of various dopamine-related agents in competing for [3H]clozapine binding with their affinities for cloned and expressed D1, D2, D3, and D4 receptors (Table 3). [3H]Clozapine does not seem to bind to D1a or D1b receptors (D1b being the same as D5). For instance, (+)butaclamol has a Ki value in the low nanomolar range for the D1 receptor but a Ki of 1 µM for [3H]clozapine binding. D2, D3, and D4 receptors have a fairly similar pharmacologic profile. One major exception is that D4 receptors display a low affinity for most neuroleptics but relatively high affinity for clozapine (9). Thus, the potency of clozapine at D4 receptors is consistent with its potency at [3H]clozapine binding sites. However, there are other major differences. For instance, spiperone displays subnanomolar affinity for D2, D3, and D4 receptors but has a Ki of 30 nM at [3H]clozapine sites. Further, raclopride displays low nanomolar potency at D2, D3, and D4 sites, but, at 10 µM concentration, it fails to affect [3H]clozapine sites. Finally, the Ki of dopamine for [3H]clozapine sites is 5 mM, more than 1000 times greater than its Ki for D2, D3, and D4 receptors.
A substantial number of serotonin receptor subtypes have been differentiated by ligand binding studies. Molecular cloning has led to the demonstration of an even greater heterogeneity so that many of the sites labeled in brain membranes by ligand binding actually represent a mixture of 5HT receptor subtypes. To compare drug potencies at 5HT receptor subtypes with [3H]clozapine binding sites, we have summarized reports from the literature on molecularly cloned and expressed receptors (Table 4). [3H]Clozapine sites in brain are not likely to involve 5HT1C receptors, as spiperone is almost fifty times more potent at clozapine binding sites than at 5HT1C receptors. In addition, mesulergine is more than 300 times as potent at 5HT1C sites than at [3H]clozapine sites. The 5HT1E site is also excluded, as ergotamine has a 60-fold higher affinity for [3H]clozapine sites than for the cloned 5HT1E receptor. The 5HT2 receptor can likewise be excluded, since it has a dramatically higher affinity for ritanserin than [3H]clozapine binding sites. The 5HT7 receptor does not fit, because mesulergine’s affinity for 5HT7 receptors is 50 times greater than for [3H]clozapine sites.
The 5HT6 receptor displays high affinity for a variety of psychotropic drugs (14, 35). We utilized cell lines expressing molecularly cloned rat 5HT6 receptors and in our laboratory compared drug affinities for [3H]clozapine, [3H]5HT, and [3H]LSD binding to the expressed 5HT6 receptors with [3H]clozapine binding in rat cerebral cortex membranes (Table 5). The similarities in the pharmacology of the 5HT6 receptor and [3H]clozapine sites are striking. Tricyclic structures, including tricyclic antidepressants and phenothiazines, display nanomolar potencies at 5HT6 receptors and [3H]clozapine sites. Some structures related to ergots, such as methiothepin, are extremely potent at 5HT6 receptors and clozapine sites, whereas mesulergine, which is in the same structural class, is about 100 times less potent at both sites. For some drugs, affinities vary depending on whether the ligand is [3H]clozapine, [3H]5HT, or [3H]LSD. However, none of these differences are marked. One notable exception is 5HT itself whose Ki at [3H]clozapine sites is 0.1–0.2 mM (data not shown) whereas it displays a 0.15 µM Ki at 5HT6 receptors.
The relatively low potency of 5HT itself at [3H]clozapine binding sites remains a puzzle. Potencies of neurotransmitters and other agonists in competing for [3H] antagonists are often relatively weak and vary considerably depending on the ligand employed. Antagonist ligands such as [3H]clozapine may fix the conformation of the receptor in an antagonist preferring conformation that diminishes affinity of the agonist. Alternatively, sites labeled by [3H]clozapine may not be 5HT6 receptors but instead a distinct receptor with similar drug specificity.
The main finding of this study is abundant, high-affinity binding of [3H]clozapine to rat brain membranes. Our results are in agreement with other studies of [3H]clozapine binding showing two high-affinity binding sites (34, 35). The first is a muscarinic cholinergic receptor which has low nanomolar affinity and represents the majority ∼60% of [3H]clozapine binding in the brain. The second site demonstrates a pharmacology similar to that we have seen for the cloned 5HT6 receptor. It is possible that other sites, such as the D4 receptor, may be labeled by [3H]clozapine but would escape detection because of low abundance in brain or because of overlapping pharmacology as occurs with multiple serotonin receptors.
One notable feature of [3H]clozapine binding is its equal distribution in the cortex, hippocampus, and striatum. In one published study of cloned 5HT6 receptors, mRNA levels were roughly equal in these three regions (33). While Sibley and collaborators (14) initially described high levels of 5HT6 mRNA in the corpus stiatum with negligible levels in cortex and hippocampus, reexamination using in situ hybridization indicates substantially higher levels of 5HT6 receptor mRNA in cortex and hippocampus than first reported (D. R. Sibley and J. E. Lachowitz, unpublished observations). Thus, both on the basis of drug specificity and regional distribution, the [3H]clozapine binding sites not involving muscarinic receptors resemble the 5HT6 receptor.
Compared with conventional neuroleptics, clozapine and other atypical neuroleptics display two unique properties, a low incidence of EPS and greater therapeutic efficacy for negative and possibly even for positive symptoms of schizophrenics. This enhanced efficacy was first shown in patients resistant to conventional neuroleptics (1), but may hold for nonresistant patients as well (1, 36). Properties unique to the atypical drugs may thus clarify neural mechanisms mediating negative symptoms as well as modulating EPS. Influences on 5HT receptors appear relevant to clozapine actions, as clozapine displays very great potency at most 5HT receptors, and 5HT receptors modulate dopamine systems involved in EPS. Thus, chronic administration of conventional neuroleptics decreases firing rates of the A10 dopamine cells that project to the limbic system and the A9 cells of the substantia nigra that project to the corpus striatum. Presumably, the slowed firing rate of the dopamine cells accentuates the effects of direct dopamine receptor blockade. The observations that clozapine (37) and other atypical neuroleptics (13,38,39) slow A10 but not A9 cells could explain the lowered EPS incidence associated with clozapine. Metabolic measures showing greater dopamine release in the corpus striatum following atypical than conventional neuroleptics support the neurophysiologic data (36,40). A link to 5HT2 receptors comes from observations that combinations of the 5HT2 selective drug ritanserin and the D2 antagonist haloperidol elicit striatal dopamine metabolic patterns resembling clozapine (41). Similarly, administration of ritanserin alleviates EPS provoked by haloperidol (10). The similarity of 5HT6 and 5HT2 receptors suggests that pharmacologic data implicating 5HT2 receptors in actions of atypical neuroleptics would apply also to 5HT6 receptors. Interestingly, ritanserin’s affinity for 5HT6 receptors varies depending on the ligand employed with Ki values of 16 nM with [3H]5HT and 1000 with [3H]clozapine. At [3H]clozapine binding sites in cortical membranes, ritanserin has a Ki of 150 nM.
Notably, while clozapine has low nanomolar affinity for numerous serotonin as well as other receptors, the predominant receptors labeled by [3H]clozapine in the present study were muscarinic cholinergic and 5HT6-like. In part, this may reflect receptor density in that the very low density of D4 receptors in rat brain (9) may have precluded their labeling. Alternatively, conformational alterations in receptors elicited by a radioligand may influence a drug’s apparent affinity. Accordingly, the best indication of sites with which the drug will interact in vivo will come from properties of a radiolabeled drug’s binding in the intact human brain. Whether the same sites are labeled by [3H]clozapine in rat brain membranes is unclear. Nonetheless, sites labeled by [3H]clozapine, whether reflecting 5HT6 or related receptors, may provide a model system for identifying candidate atypical neuroleptics and clarifying their unique psychoactive properties.
Kane J, Honigfeld G, Singer J, Meltzer H. (1988) Clozapine for the treatment-resistant schizophrenic. Arch. Gen. Psych. 45: 789–796.
Meltzer HY. (1988) Clozapine: Clinical advantages and biologic mechanisms. In: Schulz C, Tamminga C, Chase TN, Christensen AV, Gerlach J (eds). Schizophrenia: A Scientific Focus. Oxford University Press, New York, pp 302–309.
Peroutka SJ, Snyder SH. (1980) Relationship of neuroleptic drug effects at brain dopamine, serotonin, alpha-adrenergic, and histamine receptors to clinical potency. Am. J. Psychiat. 137: 1518–1522.
Richelson E. (1984) Neuroleptic affinities for human brain receptors and their use in predicting adverse effects. J. Clin. Psychiat. 45: 331–336.
Seeman P. (1992) Dopamine receptor sequences. Therapeutic levels of neuroleptics occupy D2 receptors, clozapine occupies D4. Neuropsychopharmacology 7: 261–284.
Fardé L, Nordström A-L, Wiesel FA, Pauli S, Halldin C, Sedvall G. (1992) Positron emission tomographic analysis of central D1 and D2 dopamine receptor occupancy in patients treated with classical neuroleptics and clozapine. Arch. Gen. Psych. 49: 538–544.
Miller RJ, Hiley CR. (1974) Anti-muscarinic properties of neuroleptics and drug-induced Parkinsonism. Nature 248: 596–597.
Snyder SH, Greenberg D, Yamamura HI. (1974) Antischizophrenic drugs and brain cholinergic receptors. Arch. Gen. Psych. 31: 58–61.
Van Tol HHM, Bunzow JR, Guan H, et al. (1991) Cloning of the gene for human dopamine D4 receptor with high affinity for the antipsychotic clozapine. Nature 350: 610–614.
Bersani G, Grispini A, Marini S, Pasini A, Valducci M, Ciani N. (1986) Neuroleptic-induced extrapyramidal side effects: Clinical perspectives with ritanserin (R35667), a new selective 5-HT2 receptor blocking agent. Curr. Ther. Res. 40: 492–499.
Rogue A, Rogue P. (1992) Mianserin in the management of schizophrenia. In: Schizophrenia Intl. Conf. Abstr. Vancouver, British Columbia, p. 135.
Meltzer HY, Shigehiro M, Lee J. (1989) Classification of typical and atypical antipsychotic drugs on the basis of dopamine D1, D2 and serotonin 2 pKi values. J. Pharmacol. Exp. Ther. 251: 238–246.
Stockton ME, Rasmussen K. (1993) A comparison of olanzepine and clozapine effects on dopamine neuronal activity: An electrophysiological study. Neurosci. Abstr. 19: 383.
Monsma FJ, Shen Y, Ward RP, Hamblin MW, Sibley DR. (1993) Cloning and expression of a novel serotonin receptor with high affinity for tricyclic psychotropic drugs. Mol. Pharmacol. 43: 320–327.
Cheng YC, Prusoff WH. (1973) Relationship between the inhibition constant (K ini) and the concentration of inhibitor which causes 50 percent inhibition (IC50) of an enzymatic reaction. Biochem. Pharm. 22: 3099–3108.
Dearry A, Gingrich JA, Falardeau P, Fremeau T, Bates MD, Caron MG. (1990) Molecular cloning and expression of the gene for a human D1 dopamine receptor. Nature 347: 72–76.
Zhou Q, Grandy DK, Thambi L, Kushner JA, Van Tol HHM, Cone R, Pribnow D, Salon J, Bunzow JR, Civelli, O. (1990) Cloning and expression of human and rat D1 dopamine receptors. Nature 347: 76–80.
Sokoloff P, Giros B, Martres M, Bouthenet M, Schwartz J. (1990) Molecular cloning and characterization of a novel dopamine receptor (D3) as a target for neuroleptics. Nature 347: 146–151.
Fargin A, Raymond JR, Lohse MJ, Kobilka BK, Caron MG, Lefkowitz, RJ. (1988) The genomic clone G-21 which resembles the β-adrenergic receptor sequence encodes the 5HT-1A receptor. Nature 335: 358–360.
Albert PR, Zhou Q, Van Tol HHM, Bunzow JR, Civelli O. (1990) Cloning, functional expression, and mRNA tissue distribution of the rat 5-hydroxytryptamine 1A receptor gene. J. Biol. Chem. 265: 5825–5832.
Maroteaux L, Saudo F, Amlaiky N, Boschert U, Plassat JL, Hen R. (1992) Mouse 5HT1B serotonin receptor: Cloning, functional expression, and localization in motor control centers. Proc. Natl. Acad. Sci. U.S.A. 89: 3020–3024.
Adham N, Romanienko P, Hartig P, Weinshank RL, Branchek T. (1991) The rat 5-hybroxytryptamine 1B receptor is the species homolog of the human 5-hydroxytryptamine 1Dβ receptor. Mol. Pharmacol. 41: 1–7.
Voigt MM, Laurie DJ, Seeburg PH, Bach A. (1991) Molecular cloning and characterization of a rat brain cDNA encoding a 5-hydroxytryptamine 1B receptor. EMBO J. 10: 4017–4023.
Roth BL, Ciaranello RD, Meltzer HY. (1992) Binding of typical and atypical antipsychotic agents to transiently expressed 5-HT 1C receptors. JPET. 260: 1361–1365.
Zgombick JM, Schecter LE, Macchi M, Hartig P, Branchek TA, Weishank RL. (1992) Human gene S31 encodes the pharmacologically defined serotonin 5-hydroxytryptamine 1E receptor. Mol. Pharmacol. 42: 180–185.
McAllister G, Charlesworth A, Snodin C, Beer MS, Noble AJ, Middlemiss DN. (1992) Molecular cloning of a serotonin receptor from human brain (5HT 1E): a fifth 5HT1-like sub-type. Proc. Natl. Acad. Sci. U.S.A. 89: 5517–5521.
Amlaiky N, Ramboz S, Boschert U, Plassat J, Hen R. (1992) Isolation of a mouse “5HT1E-like” serotonin receptor expressed predominantly in hippocampus. J. Biol. Chem. 267: 19761–19764.
Meltzer HY, Shigehiro M, Lee J. (1989) Classification of typical and atypical antipsychotic drugs on the basis of dopamine D-1, D-2 and serotonin 2 pKi values. JPET 251: 238–246.
Matthes H, Boschert U, Amlaiky N, Gailhe, Plassa, J, Muscatelli, Mattei M, Hen R. (1993) Mouse 5-hydroxytiyptamine 5A and 5-hydroxytryptamine 5B receptors define a new family of serotonin receptors: Cloning, functional expression, and chromosomal localization. Mol. Pharmacol. 43: 313–319.
Roth BL, Craigo SC, Choudhary MS, Uluer A, Monsma FJ, Shen Y, Meltzer HY, Sibley DR. (1994) Binding of Typical and Atypical antipsychotic Agents to 5-Hydroxytryptamine-6 and 5-Hydroxtryptamine-7 Receptors. J. Pharmacol. Exp. Ther. 268: 1403–1410.
Ruat M, Traiffort E, Leurs R, Tardivel-Lacombe J, Diaz J, Arrang J, Schwartz J. (1993) Molecular cloning, characterization, and localization of a high-affinity serotonin receptor (5HT-7) activating cAMP formation. Proc. Natl. Acad. Sci. U.S.A. 90: 8547–8551.
Shen Y, Monsma FJ, Metcalf MA, Jose PA, Hamblin MW, Sibley DR. (1993) Molecuar cloning and expression of a 5-hydroxytiyptamine 7 serotonin receptor subtype. J. Biol. Chem. 268: 18200–18204.
Ruat M, Traiffort E, Arrang J, et al. (1993) A novel rat serotonin (5-HT6) receptor: Molecular cloning, localization and stimulation of cAMP accumulation. Biochem. Biophys. Res. Commun. 193: 268–276.
Hauser D, Closse A. 3H-clozapine binding to rat brain membranes. Life Sci. 23: 557–562.
Flamez A, De Backer J, Wiczak N, Vauqelin G, De Keyser J. (1994) [3H]Clozapine is not a suitable ligand for labelling of D4 dopamine receptors in post mortem brain. Neurosci. Lett. 175: 17–20.
Meltzer HY. (1991) The mechanism of action of novel antipsychotic drugs. Schizophrenia Bull. 17: 263–287.
Chiodo LA, Bunney BS. (1985) Possible mechanisms by which repeated clozapine administration differentially affects the activity of two subpopulations of midbrain dopamine neurons. J. Neurosci. 5: 2539–2544.
Skarsfeldt T, Perregaard J. (1990) Sertindole, a new neuroleptic with extreme selectivity on A10 versus A9 dopamine neurons in the rat. Eur. J. Pharmacol. 182: 613–614.
Goldstein JM, Litwin LC, Sutton EB, Malick JB. (1993) Seroquel: Electrophysiological profile of a potential atypical antipsychotic. Psychopharmacology 112: 292–299.
Ichikawa J, Meltzer HY. (1991) Differential effects of repeated treatment with haloperidol and clozapine on dopamine release and metabolism in the striatum and the nucleus accumbens. J. Pharmacol. Exp. Ther. 256: 348–357.
Gudelsky FA, Nwajei EE, DeFife K, Nash JF. (1992) Interaction of amfonelic acid with antipsychotic drugs on dopaminergic neurons. Synapse 12: 304–311.
Supported by USPHS Grant MH-18501, Research Scientist Award DA-00074 to SHS, and a grant of the Stanley Foundation Research Awards Program. We thank H. Meltzer for advice and O. Civelli for assistance in initial studies of cloned dopamine receptors.
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Glatt, C.E., Snowman, A.M., Sibley, D.R. et al. Clozapine: Selective Labeling of Sites Resembling 5HT6 Serotonin Receptors May Reflect Psychoactive Profile. Mol Med 1, 398–406 (1995). https://doi.org/10.1007/BF03401577