TCP Binding Research Articles

Discovery of 6,11-ethano-12,12-diaryl-6,11-dihydrobenzo[b]quinolizinium cations, a novel class of N-methyl-D-aspartate antagonists.

6,11-Ethano-12,12-diaryl-6,11-dihydrobenzo[b]quinolizinium cations 8, a novel class of N-methyl-D-aspartate (NMDA) antagonists acting at the phencyclidine site, have been identified. Structure-activity relationship studies around the lead compound 8a led to the identification of 12g (WIN 67870-2), one of the most potent compounds in this series. Compound 12g has a Ki = 1.8 +/- 0.2 nM vs [3H]TCP binding, has 700-fold selectivity for binding to the open state of the NMDA receptor-ionophore, and was devoid of MK-801- and PCP-like behavioral effects in rats. Compound 12g was neuroprotective in cultured mouse cortical neurons and exhibited antiischemic activity in a rat middle cerebral artery occlusion/reperfusion model of focal ischemia.

NE-100, a novel sigma ligand: Effects on [ 3H]TCP binding to intact primary cultured neuronal cells

N,N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride (NE-100), a potent and highly selective sigma ligand, haloperidol, (+)pentazocine, 1-(cyclopropylmethyl)-4-(2′-(4″-fluorophenyl)-2′-oxoethyl) piperidine HBr (DuP 734) and 4-(2′-(4″-cyanophenyl)-2′-oxoethyl)1-(cyclopropylmethyl)piperidine (XJ 448) inhibited carbachol-induced inositol 1,4,5-triphosphate (IP 3) formation in a dose-dependent manner. The rank order of potency of the tested drugs for inhibition was: haloperidol ≥ (+) pentazocine = NE−100 > DuP 734 = XJ 448 . In addition, the effects of NE-100, DuP 734 and XJ 448 upon [ 3H]TCP binding were examined using primary cultured neuronal cells derived from the fetal rat telencephalon. These drugs inhibited [ 3H]TCP binding to intact cells. The ability of the test drugs to inhibit [ 3H]TCP binding to primary cultured neuronal cells was in the order: NE-100 > DuP 734 > XJ 448 . Thes observations suggest that NE-100 indirectly modulates the N-methyl-D-aspartate (NMDA)/phencyclidine (PCP) receptor ion channel complex (NMDA receptor-ion channel), presumably through sigma-1 sites.

Density of NMDA‐Coupled and Uncoupled 1‐[1‐(2‐[3H]Thienyl)cyclohexyl]piperidine Recognition Sites in the Brain and Spinal Cord: Differential Effects of NMDA Agonists and Antagonists

Binding of 1-[1-(2-[3H]thienyl)cyclohexyl]piperidine ([3H]TCP) to mouse brain and spinal cord membranes was studied using compounds selective for the NMDA-coupled 1-(1-phenylcyclohexyl)piperidine (PCP) and/or sigma recognition sites. In both tissues, [3H]TCP labeled two populations of binding sites. Density of the low-affinity sites was approximately the same in both tissues, but the population of the high-affinity [3H]TCP sites was three times bigger in the brain than in the spinal cord. Self- and cross-displacement studies showed that the high-affinity [3H]TCP binding sites could be identical with NMDA receptor-coupled PCP sites, whereas the low-affinity [3H]TCP sites may be associated with sigma binding sites in both tissues. The NMDA-coupled PCP sites labeled in the presence of 6.25 nM [3H]TCP constituted a much higher percentage of the total binding in the brain (75%) than in the spinal cord (44%). Consistent with this, reintroduction of glycine and glutamate significantly increased, but DA antagonists significantly inhibited [3H]TCP binding in the brain but not in the spinal cord. Together, these data suggest that a large component of [3H]TCP-labeled binding sites in the spinal cord may be associated with sigma but not the NMDA receptor-coupled PCP sites.

Changes in muscarinic (M 1 and M 2 subtypes) and phencyclidine receptor density in the rat brain following trimethyltin intoxication

The main purpose of this study was to determine whether one or both of the muscarinic receptor subtypes (M 1 and M 2) contributed to the total cholinergic receptor loss found in trimethyltin (TMT) treated rats and to assess the effect of TMT of phencyclidine (PCP) receptor density in several regions of the rat brain. The distribution and changes in muscarinic (M 1 and M 2) receptor and PCP receptor sites were analysed by means of quantitative autoradiography using [ 3H]quinuclidinyl benzilate (QNB) and [ 3 H]N -(1-(2-thienyl) cyclohexyl) 3,4-piperidine (TCP) respectively. The results demonstrate a TMT induced decrease in [ 3H]QNB binding in a large number of brain regions particularly the hippocampal formation, for both M 1 and M 2 muscarinic receptor subtypes. There is also a decrease in [ 3H]TCP binding in several brain regions. The effects of TMT on PCP receptors suggest that TMT induced damage is not restricted to the cholinergic system and that N- methyl- d-aspartate (NMDA) receptors are also affected.

An electrophilic affinity ligand based on (+)-MK801 distinguishes PCP site 1 from PCP site 2

The electrophilic affinity ligand, (+)-3-isothiocyanato-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cycl ohepten-5,10 - imine hydrochloride [(+)-MK801-NCS] was characterized for its ability to acrylate phencyclidine (PCP) and sigma binding sites in vivo. Initial studies, conducted with mouse brain membranes, characterized the binding sites labeled by [3H]1-[1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP). The Kd values of [3H]TCP for PCP site 1 (MK801-sensitive) and PCP site 2 (MK801-insensitive) were 12 nM and 68 nM, with Bmax values of 1442 and 734 fmol/mg protein, respectively. Mice were sacrificed 18-24 hours following intracerebroventricular administration of the acylator. The administration of (+)-MK801-NCS increased [3H]TCP binding to site 2, but not to site 1. Although (+)-MK801-NCS decreased [3H](+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d; ccyclohepten-5,10-imine maleate ([3H](+)-MK801) binding to site 1, it had no effect on [3H]TCP binding to site 1. Viewed collectively with other published data, these data support the hypothesis that PCP sites 1 and 2 are distinct binding sites, and that [3H]TCP and [3H](+)-MK801 label different domains of the PCP binding site associated with the NMDA receptor.

Blockade of NMDA-induced potentiatiation of [ 3H]TCP binding to rat brain membranes by histogranin

Histogranin (H-Met-Asn-Tyr-Ala-Leu-Lys-Gly-Gln-Gly-ArB-Thr-Leu-Tyr-Gly-Phe-COOH) was first isolated from bovine adrenal medulla (1) and shown to antagonize «in vitro» N-methyl-D-aspartate (NMDA)-induced locus coeruleus cell depolarization (unpublished) and «in vivo» NMDA-induced mouse convulsions. Histogranin possesses its own high affinity saturable binding site in rat brain membranes insensitive to the presence of NMDA (2). On the other hand, histogranin is a potent non-competitive inhibitor of the binding of the NMDA receptor ligand, [ 3 H]CGP 39653 (1)

Effects of Kamikihito, a Traditional Chinese Medicine, on Neurotransmitter Receptor Binding in the Aged Rat Brain Determined by In Vitro Autoradiography (1): Changes in the [3H]QNB Binding

Using in vitro autoradiography, we investigated the effects of Kamikihito (KKT), a traditional Chinese medicine, on the specific binding of [3H]quinuclidinyl benzilate (QNB) and [3H]N-(1-[2-thienyl]cyclohexyl)-3,4-piperidine (TCP) in the rat brain. The Bmax but not the Kd values for [3H]QNB binding to the caudate/putamen and accumbens in aged rats were lower than those in young rats. The [3H]TCP binding was also decreased in aged rats compared with that in young rats. Long-term administration of KKT modulated the [3H]QNB binding in young but not aged rats.

Phencyclidine-binding sites in mouse cerebral cortex during development and ageing: effects of inhibitory amino acids

The binding of N-[1-(2- thienyl)cyclohexyl]-[ 3H]piperidine ([ 3H]TCP) to the phencyclidine-binding sites in the N-methyl-D-aspartate (NMDA) receptor complex-associated ion channel was characterized in cerebral cortical membranes from 3-day-old to 24-month-old mice. The binding was saturable, exhibiting only one binding component during the whole life-span studied. The maximal binding capacity B max, calculated per protein content, decreased during postnatal development until 3 months of age, remaining thereafter constant in ageing mice, thus indicating the greatest availability of phencyclidine-binding sites in the immature cerebral cortex. The binding contant K D increased during the first postnatal week, remained thereafter unchanged and increased again during the second year of life, indicating a decreased affinity of these receptor sites for the ligand. The general properties of the binding; potentiation by glutamate and NMDA, as well as by glycine in a strychnine-insensitive manner, prevailed during development and ageing, certain of the effects being however less pronounced in the immature brain. Taurine and β-alanine stimulated TCP binding, acting probably at the glycine modulatory site. The actions of these inhibitory amino acids were weak and inconsistent when compared to that of glycine. Since NMDA receptors have been suggested to be involved in neuronal plasticity and learning and memory processes, these modifications in the properties of cortical phencyclidine-binding sites might be of importance in the regulation of excitatory amino acid functions during development and ageing.

Paradoxical effects of 2‐amino‐4‐phosphonobutanoic acid and other 2‐amino‐ω‐phosphonoalkanoic acid on the n‐methyl‐d‐aspartate receptor ion channel

AbstractAn extensively washed membrane preparation was used to measure the noncompetitive binding site of the N‐methyl‐D‐aspartate (NMDA) receptor complex with [3H]‐1‐[1‐(2‐thienyl) cyclo‐hexyl] piperidine ([3H]‐TCP). Addition of glutamic acid (Glu) or Glu + glycine (Gly) increased the binding of [3H]‐TCP, reflecting that the NMDA receptor ion channel was opened by these compounds. [3H]‐TCP binding was also increased by various 2‐amino‐ω‐phosphonoalkanoic (AP) compounds, of which 2‐amino‐4‐phosphonobutanoic acid (AP4) was the most effective. The effects of AP compounds were also observed in the presence of Gul. However, in the presence of Gly, or in the presence of Glu + Gly, 2‐amino‐5‐phosphonopentanoic acid (AP5) and 2‐amino‐7‐phosphonoheptanoic acid (AP7) inhibited the binding. The enhancing effect of AP4 was also inhibited by these compounds. Our conclusion is that AP compounds exert complex effects: they bind to and effect the NMDA receptor complex ion channel not only via the competitive binding site but may also interact with the potentiating Gly binding site. © Wiley‐Liss, Inc.

Potent quinoxaline-spaced phosphono alpha-amino acids of the AP-6 type as competitive NMDA antagonists: synthesis and biological evaluation.

A series of alpha-amino-3-(phosphonoalkyl)-2-quinoxalinepropanoic acids was synthesized and evaluated for NMDA receptor affinity using a [3H] CPP binding assay. Functional antagonism of the NMDA receptor complex was evaluated in vitro using a stimulated [3H]TCP binding assay and in vivo by employing an NMDA-induced seizure model. Some analogues also were evaluated in the [3H]-glycine binding assay. Several compounds of the AP-6 type show potent and selective NMDA antagonistic activity both in vitro and in vivo. In particular alpha-amino-7-chloro-3-(phosphonomethyl)-2-quinoxalinepropanoic acid (1) displayed an ED50 of 1.1 mg/kg ip in the NMDA lethality model. Noteworthy is alpha-amino-6,7-dichloro-3-(phosphonomethyl)-2-quinoxalinepropanoic++ + acid (3) with a unique dual activity, displaying in the NMDA receptor binding assay an IC50 of 3.4 nM and in the glycine binding assay an IC50 of 0.61 microM.

Neomycin is an agonist at a polyamine site on the N-methyl-D-aspartate receptor.

Neomycin appears as a full agonist and spermidine as a partial agonist at the site where polyamines enhance 1-[1-(2-thienyl)cyclohexyl][3H]piperidine ([3H]TCP) binding on the N-methyl-D-aspartate (NMDA) receptor. Other aminoglycosides also enhance [3H]TCP binding with efficacies roughly proportional to the number of primary amine groups. The polyamine antagonists ifenprodil and arcaine inhibit enhancement of [3H]TCP binding by spermidine or neomycin. The inhibition of [3H]TCP binding by arcaine is apparently competitively reduced by neomycin and spermidine, supporting a common site. Diethylenetriamine (previously described as a polyamine antagonist) may be a partial agonist. Enhancement by neomycin or spermidine is not additive to that of Mg2+, consistent with competition of Mg2+ and spermidine or neomycin at the site where these compounds enhance [3H]TCP binding. Polyamines also enhance the binding of the competitive antagonist 2-(2-carboxypiperazin-4-yl)[3H]propyl-1-phosphonic acid ([3H]CPP). Neomycin, which does not enhance [3H]CPP binding, inhibits the enhancement by spermidine. That this site is distinct from the site where spermidine and neomycin increase [3H]TCP binding is supported by different pharmacology. Arcaine and diethylenetriamine do not inhibit spermidine enhancement of [3H]CPP binding. Mg2+ also does not compete with the spermidine enhancement of [3H]CPP binding. Ifenprodil inhibits the spermidine enhancement of [3H]CPP binding. The data suggest two or more polyamine sites, with arcaine selective for the site that enhances [3H]TCP binding. Neomycin is an agonist at one polyamine site and antagonist to the second.

Thiokynurenates prevent excitotoxic neuronal death in vitro and in vivo by acting as glycine antagonists and as inhibitors of lipid peroxidation

Several derivatives of kynurenic and thiokynurenic acids were synthesized and tested for their ability to protect primary cultures of cerebellar granule cells against excitotoxic damage, and to affect the binding of [ 3H]glycine ([ 3H]Gly), [ 3H]α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid ([ 3H]AMPA), [ 3H]3-(2-carboxypiperazine-4-yl-)propyl-1-phosphonic acid ([ 3H]CPP), [ 3H]kainic acid and [ 3H]N-[1-(2-thienyl)cyclohexyl]-3,4-piperidine ([ 3H]TCP) to rat cortical membranes. Kynurenic and thiokynurenic acid derivatives with one or two halogens in position 5 or 7 were selective glycine antagonists, failing to affect N-methyl-D-aspartate (NMDA), kainate or AMPA sites at micromolar concentrations. 7-Cl-kynurenic, 7-Cl-thiokynurenic, 5,7-diCl-kynurenic and 5,7-diCl-thiokynurenic acids had similar IC 50s for displacing [ 3H]Gly from its strychnine-insensitive site and for reducing the stimulated (0.5 μM NMDA and 1 μM glycine) [ 3H]TCP binding to cortical membranes. However, 7-Cl-thiokynurenic acid was particularly potent to prevent excitotoxic neuronal death in cultured cerebellar granule cells. This action may be ascribed to inhibition of lipid peroxidation, a property which was demonstrated for the 5- or 7-Cl derivatives of thiokynurenic acid. Futhermore, 7-Cl-thiokynurenic acid reduced excitotoxic damage caused by the injection of quinolinic acid in the rat striatum. Thus, 7-Cl-thiokynurenic acid appears to be a new compound with interesting antiexcitotoxic properties both in vitro and in vivo.

125I]Thienylphencyclidine, a novel ligand for the NMDA receptor

We have monitored the binding of [ 125I]thienylphencyclidine ([ 125I]TCP), a novel high affinity radioiodinated ligand that specifically recognizes the NMDA (N-methyl-D-aspartate) receptor in rat brain membranes. [ 125I]TCP binds with an affinity of about 30 nM, and recognizes a similar number of binding sites to previously employed ligands for this receptor. [ 125I]TCP binding is characterized by slow association and dissociation rates, and the latter can be modified by the addition of Mg 2+ or Zn 2+, as previously described for [ 3H]dizocilpine ([ 3MK801). Other phencylidine-like ligands displaced [ 125I]TCP binding with the order of potency dizocilpine > thienylphencyclidine > ITCP > phencyclidine > ketamine. The binding of [ 125I]TCP was also increased by NMDA and glycine-site agonists and inhibited by antagonists of these sites. Surprisingly, however, the polyamines spermidine and spermine did not increase [ 125I]TCP, even though the polyamine antagonist arcaine was an effective inhibitor of binding. These results show that [ 125I]TCP is a useful ligand for the NMDA receptor complex that binds to the receptor in a manner that is qualitatively distinct from previously described ligands.

Effects of glycine antagonists on Mg 2+- and glycine-induced [formula omitted] binding

We investigated the effects of glycine antagonists, 3-amino-1-hydroxy-2-pyrrolidone (HA-966), 7-chlorokynurenic acid (7-Cl-KYNA), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), 6,7-dichloro-3-hydroxy-2-quinoxalinecarboxylic acid (DHQXC), 6,7-dichloroquinoxaline-2,3-dione (DCQX), and 5-chloro-indole-2-carboxylic acid (5-Cl-I2CA), on Mg 2+- and glycine-induced [ 3 H]N-(1-[2- thienyl]cyclohexyl)-3,4- piperidine([ 3 H]TCP) binding to well-washed rat cortical membranes. Except for 5-Cl-I2CA, all the glycine antagonists completely inhibited not only glycine- but also Mg 2+-induced [ 3H]TCP binding in a concentration-dependent manner. Out of all the glycine antogonists examined DHQXC most selectively inhibited Mg 2+-induced [ 3H]TCP binding, while DCQX was the most selective for inhibiting glycine-induced [ 3H]TCP binding.

MPTP lesions of the nigrostriatal dopaminergic projection decrease [3H]1-[1-(2-thienyl)cyclohexyl]piperidine binding to PCP site 2: further evidence that PCP site 2 is associated with the biogenic amine reuptake complex.

Our previous studies have demonstrated that, using membranes of guinea pig brain, [3H]1-[1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP) labels not only the phencyclidine binding site associated with the NMDA receptor (PCP site 1), but also a second high affinity binding site which is associated with the biogenic amine reuptake carrier (termed PCP site 2). To test this hypothesis, the binding of [3H]GBR12935 to the dopamine transporter, and [3H]TCP binding to PCP sites 1 and 2 were measured in caudates harvested from control, MPTP-treated and reserpine-treated dogs. MPTP treatment decreased dopamine levels by over 99%, decreased [3H]GBR12935 binding by over 90%, decreased [3H]TCP binding to PCP site 2 by about 50%, and had no significant effect on [3H]TCP binding to PCP site 1. These data are consistent with the hypothesis that a portion of PCP site 2 is associated with dopaminergic nerve terminals in dog caudate.

Long-term potentiation is associated with increased [ 3H]AMPA binding in rat hippocampus

The location and nature of the changes underlying long-term potentiation (LTP) remain controversial issues. In this study, we tested the possibility that changes in binding properties of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/quisqualate andN-methyl- d-aspartate (NMDA) subtype of glutamate receptors are associated with LTP. LTP was elcited in vivo by stimulation of the perforant pathway in anesthetized rats. One hour following stimulation the animals were sacrificed. We performed quantitative ligand binding autoradiography on frozen brain sections using [ 3H]AMPA and 3H]N-(1-(2-thienyl)cyclohexyl)-3,4-piperidine ([ 3H]TCP) to label the AMPA/quisqualate and the NMDA receptors, respectively. No changes in [ 3H]TCP binding were detected in any of the treatment groups. However, increases in [ 3H]AMPA binding were observed only in animals that exhibited LTP. These increases were bilateral and present in several subfields of the hippocampus and cortical areas. Administration of the NMDA receptor antagonist, ketamine, prior to tetanic stimulation prevented both the increase in binding and the induction of LTP. These results suggest that changes in the characteristics of AMPA/quisqualate receptors are a biochemical correlate of LTP.

The quaking mouse: an epileptic mutant with alterations affecting the modulatory mechanisms of the NMDA receptor complex

The binding of [ 3H]glutamate and of [ 3H]1-(1-(2-thienyl)cyclohexyl)piperidine ([ 3H]TCP) has been examined in the genetically epileptic mutant mouse, quaking. The density of [ 3H]glutamate binding sites did not differ between the quaking mice and their controls of the same strain. In the absence of exogenous glutamate or glycine, the density of [ 3H]TCP binding sites was also similar in the two strains. In both the mutants and their controls, exogenously added glutamate, glycine and glutamate plus glycine dose-dependently increased the binding of [ 3H]TCP. In the 3 conditions, the modulation of [ 3H]TCP binding was significantly more efficient in the quaking mice than in the controls. Furthermore, in the presence of glutamate (10 −5 M), the increase of the affinity of the ligand for the ion channel binding site was higher in the mutants than in the controls. These results suggest that the modulatory mechanisms of the N- methyl- d-aspartate/ionophore receptor complex might be altered in these mutants. These alterations might be related to the previously observed anticonvulsant properties of NMDA receptor antagonists in the quaking mouse model of inherited epilepsy.

Alterations in phencyclidine and sigma binding sites in schizophrenic brains: Effects of disease process and neuroleptic medication

The specific binding of [ 3H]TCP and [ 3H](+)3-PPP, radioligands which respectively label PCP-NMDA and sigma binding sites was measured in tissue homogenates prepared from dissected areas of control and schizophrenic postmortem brains. [ 3H]TCP binding was bilaterally increased in orbital frontal cortex (Brodmann area 11) of schizophrenic brains. This finding may be due to an increased glutamatergic innervation of orbital frontal cortex since it parallels our findings of increased [ 3H]kainate and [ 3H] D- aspartate binding in this area. In contrast, [ 3H](+)3-PPP binding was reduced in each of the four brain regions examined. The reductions were greatest in brains from the schizophrenic subjects receiving neuroleptics at the tine of death. Neuroleptics remaining in the brains of these subjects may compete in vitro with [ 3H](+)3-PPP for binding to the sigma site.

Effect of amygdaloid kindling on the high- and low-affinity [ 3H]TCP binding sites of the rat CNS

The regulation of the binding sites of [ 3H]TCP, a non-competitive ligand of the N- methyl- d-aspartate (NMDA) receptor, was studied on membranes prepared from different CNS regions of amygdaloid-kindled rats. The high-affinity binding sites ( K d H = 4.2–7.4 nM), identified as the NMDA-gated ion channels, were not affected by kindling or by a daily injection of TCP (5 mg/kg before each electrical stimulation) which prevented kindling. These results suggest that the NMDA receptors participate to the establishment and not to the permanence of kindling. Kindling increases the number of low affinity [ 3H]TCP binding sites in the hippocampus (+21%, P < 0.01) without change of the affinity ( K d L = 340 nM). In the striatum both K d L and B max L were increased (3.3–4.4 fold, P < 0.001) in animals pretreated with TCP before each electrical stimulation for 20 days. These last results argue in favour of a function of the low-affinity [ 3H]TCP binding sites, the nature of which remains to be determined.

3H]1‐[2‐(2‐thienyl)cyclohexyl]piperidine labels two high‐affinity binding sites in human cortex: Further evidence for phencyclidine binding sites associated with the biogenic amine reuptake complex

Previous work demonstrated two high-affinity PCP binding sites in guinea pig brain labeled by [3H]TCP (1-(1-[2-thienyl]cyclohexyl)piperidine): site 1 (N-methyl-D-aspartate [NMDA]-associated) and site 2 (dopamine-reuptake complex associated). The present study examined brain membranes prepared from various species, including human, for the presence of site 2, defined as binding in the presence of (+)-5-methyl-10,11-dihydro-5H-dibenzo [a, d]cyclohepten-5,10-imine maleate ((+)-MK801) minus binding in the presence of 10 microM TCP (nonspecific binding). Studies were conducted in absence of sodium which was found to be inhibitory to [3H]TCP binding. The results demonstrated detectable levels of site 2 in brain membranes of guinea pig, rabbit, pig, mouse, sheep, and human but not in the rat or chicken. Using human cortical membranes, site 2 was the predominant binding site. Detailed studies conducted with human cortical tissue showed that high-affinity dopamine (1-[2- [bis(4-fluorophenyl)-methoxy]ethyl]-4-(3-phenylpropyl)piperazine (GBR12909)], [1,2]benzo(b)thiophenylcyclo-hexylpiperidine (BTCP), and serotonin (fluoxetine) uptake inhibitors produced a wash-resistant inhibition of [3H]TCP binding to site 2, but not site 1. Preincubation of guinea pig brain membranes with BTCP was shown to produce an increase in the dissociation rate of [3H]TCP from PCP site 2. Structure activity studies with various uptake inhibitors showed that GBR12909, benztropine, fluoxetine, and BTCP have higher affinity for site 2 than for site 1. (+)-MK801, ketamine, and tiletamine were very selective for site 1, whereas dexoxadrol and TCP were moderately selective for site 1. These results suggest that human cortex possesses high-affinity PCP binding sites associated with biogenic reuptake binding sites, and that guinea pig brain, but not rat brain, may be an appropriate animal model for studying PCP site 2 in human brain.