PCP Receptor Research Articles

Tris inhibits (+)-[ 3H]SKF-10,047 binding to σ receptors

Tris-HCl is the most commonly used buffer in studies of radioligand binding to σ receptors, with concentrations as high as 50 or 100 mM often used. We report here that these concentrations of Tris substantially inhibit (+)-[ 3H]SKF-10,047 binding to σ receptors. The well-established inhibitory effect of Tris-HCl on ligand binding to PCP receptors did not contribute to the presently reported inhibition of (+)-[ 3H]SKF-10,047 binding. The IC 50 of Tris, determined in the presence of 10 mM potassium phosphate buffer, was 15.4±1.2 mM ( n=3, pH 8.0, 25°C, 1 nM radioligand). Equilibrium saturation studies revealed an apparent competitive inhibition of binding.

Effects of phencyclidine-like drugs on responding under muitiple fixed-ratio, fixed-interval schedules

The effects of phencyclidine and a series of related drugs were studied on rates and patterns of responding by rats under a multiple fixed-interval 300s, fixed-ratio 30-response schedule of food presentation. Dizocilipine produced large increases in the rate of FI responding and smaller increases in the rate of FR responding. TCP slightly increased the rate of FI responding and PCE slightly increased the rate of FR responding. Higher doses of all drugs decreased rates of responding under both schedule components with the potency order of dizocilpine > PCE > PCP > TCP > ketamine. This relative potency for decreasing rates of FI and FR responding correlated highly with the affinity of these drugs for PCP receptors, suggesting that the rate decreasing effects of these drugs are mediated through these receptors. An analysis of local rates of responding within the FI suggested that dizocilpine was different from the arylcyclohexylamines in that it was the only drug that increased rates of responding late in the FI component. Previous reports have shown that the ability of arylcyclohexylamines to increase punished responding and to act as discriminative stimuli are correlated highly with binding to PCP receptors. The present experiments suggest that decreases in rates of responding under multiple schedule control are also mediated by PCP receptors.

Diazepam antagonizes effects on dopamine metabolism produced by pcp receptor agonists

1. 1. Male rats were injected with either saline, diazepam, MK-801, or diazepam plus MK-801. 2. 2. In previous work with phencyclidine (PCP), diazepam significantly reduced the increase in homovanillic acid (HVA) in olfactory tubercle and prefrontal cortex. 3. 3. Diazepam also lowered the HVA increase following MK-801 in caudate, olfactory tubercle, and prefrontal cortex. 4. 4. Benzodiazepine receptors may modify dopaminergic function at PCP receptors that affect dopamine neurotransmission.

Characterization of specific binding sites for [3H]-1,3-di-o-tolyl-guanidine (DTG) in the rat glioma cell line C6-BU-1.

The aim of the present study was to find out if a cell line of glial origin possesses sigma and/or phencyclidine (PCP) binding sites. Binding of [3H]1,3-di-o-tolyl-guanidine (DTG), a highly selective ligand for sigma binding sites, and of [3H]N-[1-(2-thienyl)cyclohexyl] piperidine ([3H]TCP), a radioligand specific for PCP receptors, to C6-BU-1 glioma cells was investigated. Binding of [3H]DTG to C6-BU-1 cell membranes was reversible, saturable (Bmax = 10.5 pmol/mg protein), and of high affinity (KD = 26 nM). C6-BU-1 cells do not possess PCP receptors as indicated by negligible specific binding of [3H]TCP to C6-BU-1 cell membranes. Specific binding of [3H]DTG was reduced in the presence of Ca2+ and to a lesser extent by Mg2+. The rank order of potency of various PCP and sigma ligands was DTG > (+)3-[(3-hydroxy-phenyl)-N-n-propyl-piperidine] [(+)3-PPP] > haloperidol > pentazocine > (-)3-PPP > PCP > metaphit > dextromethorphan > (-)butaclamol > (+)butaclamol > (-)N-allylnormetazocine [(-)SKF 10,047] > MK801 > (+)SKF 10,047 > ketamine. The drug specificity, confirmed by a reversed stereoselectivity for the benzomorphan opiate SKF 10,047, indicated that these sites correspond to a subtype of sigma binding sites, the so-called sigma 2 binding site. Thus, the C6-BU-1 cell line is the first glial cell line demonstrated to have sigma 2 binding sites.

Patent Evaluation: Novel N-cycloalkylamines as Sigma Agonists

SummaryNovelty: N-Cycloalkylamines are disclosed with activity as a receptor agonists. These compounds are potentially useful as antipsychotic, anticonvulsant and anti-ulcer agents.Biology: Competitive binding data are presented showing that the compounds have selective affinity for the sigma receptor in comparison to the, μ, δ and PCP receptors. Psychotropic activity is demonstrated by inhibition of picrotoxin-induced convulsion (ED50 values about 50 mg/kg, ip, and 100 mg/kg, po). Anti-ulcer activity is shown by inhibition of cysteamine induced gastrointestinal ulceration.Chemistry: Syntheses of the compounds are outlined in two schemes, forming a subsidiary part of the claim with ten examples. 1-[2-(N-Cycloproyimethyl-N-methyl)amino-2-phenyl-butyl]-2-(2-thienyl)cyclopropane is one of two specifically claimed compounds.

Recent advances in the phencyclidine model of schizophrenia

Phencyclidine (PCP, "angel dust") induces a psychotomimetic state that closely resembles schizophrenia. As opposed to amphetamine-induced psychosis, PCP-induced psychosis incorporates both positive (e.g., hallucinations, paranoia) and negative (e.g., emotional withdrawal, motor retardation) schizophrenic symptoms. PCP-induced psychosis also uniquely incorporates the formal thought disorder and neuropsychological deficits associated with schizophrenia. The purpose of the present paper is to review recent advances in the study of the molecular mechanisms of PCP action and to describe their implications for the understanding of schizophrenic pathophysiology. Twenty-five papers were identified that described the clinical dose and serum and CSF levels at which PCP induces its psychotomimetic effects. The dose range of PCP-induced effects were compared to the dose range at which PCP interacts with specific molecular targets and affects neurotransmission. It was found that PCP-induced psychotomimetic effects are associated with submicromolar serum concentrations of PCP. At these concentrations PCP interacts selectively with a specific binding site (PCP receptor) that is associated with the N-methyl-D-aspartate (NMDA)-type excitatory amino acid receptor. Occupation of its receptor by PCP induces noncompetitive inhibition of NMDA receptor-mediated neurotransmission. Other NMDA antagonists such as the dissociative anesthetic ketamine induce PCP-like neurobehavioral effects in proportion to their potency in binding to the PCP receptor and inducing NMDA receptor inhibition. These findings suggest that endogenous dysfunction of NMDA receptor-mediated neurotransmission might contribute to the pathogenesis of schizophrenia. The relative implications of the PCP and amphetamine models of schizophrenia are discussed in relationship to the diagnosis and etiology of schizophrenia.

Synthesis and receptor binding of enantiomeric N-substituted cis-N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl)cyclohexylamines as high-affinity sigma receptor ligands.

N-Alkyl-substituted derivatives of (+)- and (-)-cis-N-[2-(3,4-dichlorophenyl)ethyl]-2-(1-pyrrolidinyl)cyclohexylamin e have been synthesized in nine steps in a stereospecific manner starting from cyclohexene oxide. The key step in the reaction sequence involved catalytic hydrogenation of oxime 8 in the presence of PtO2 and AcOH to give the cis diamine (+/-)-7. Most of the compounds in this series exhibited very high affinity at sigma receptors when tested against [3H]-(+)-3-PPP, and in general it was observed that the 1R,2S enantiomers bound more potently to sigma receptors than their corresponding 1S,2R enantiomers. The most potent sigma ligand found in this class was the unsubstituted derivative (1R,2S)-(-)-4, which exhibited an affinity constant of 0.49 nM. This compound was also found to be very selective for sigma receptors. It exhibited little or no affinity for kappa opioid, PCP, and dopamine-D2 receptors. It was also demonstrated that the cis configuration as opposed to the trans configuration of (+)- and (-)-5 was necessary for a higher sigma receptor affinity.

Polyamine effects upon N-methyl- D-aspartate receptor functioning: differential alteration by glutamate and glycine site antagonists

Polyamines such as spermidine potentiate activation of the N-methyl- D-aspartate (NMDA)-type excitatory amino acid receptor. The goal of the present study was to investigate interactions between the putative polyamine binding site and previously described sites for glutamate and glycine. Binding of the high-potency PCP receptor ligand [ 3H]MK-801 to well-washed rat brain membranes was used as an in vitro probe of NMDA receptor activation. Spermidine concentration-response studies were performed in the absence and presence of both glutamate and glycine, with and without D-(−)-2-amino-5-phosphonovaleric acid ( D(−)AP-5) or 7-chlorokynurenic acid (7Cl-KYN). Incubation in the presence of spermidine alone induced a 20.4-fold increase in [ 3H]MK-801 binding with an EC 50 value of 13.3 μM. The mean concentration of spermidine which induced maximal stimulation of binding was 130 μM ( n = 10,S.E.M.= 24.66,range= 25–250 μM). Glutamate (10 μM) decreased the EC 50 value for spermidine-induced stimulation of [ 3H]MK-801 binding to 3.4 μM. Glycine (10 μM) did not significantly alter either maximum spermidine-induced [ 3H]MK-801 binding or the EC 50 value for spermidine-induced stimulation of [ 3H]MK-801 binding. Incubation in the presence of the specific glutamate antagonist D(−)AP-5 attenuated [ 3H]MK-801 binding in a glutamate-reversible fashion. The competitive glycine antagonist 7Cl-KYN decreased maximum spermidine-induced [ 3H]MK-801 binding in a glycine-reversible fashion. In addition, 7Cl-KYN increased the EC 50 value for spermidine-induced stimulation of [ 3H]MK-801 binding while D(−)AP-5 was without effect. These findings suggest that glutamate and glycine regulate the polyamine binding site differentially. PCP-like agents induce a psychotomimetic state closely resembling schizophrenia by inhibiting NMDA receptor-mediated neurotransmission. The ability of polyamines to modulate NMDA receptor functioning suggests a potential site for pharmacological intervention.

Characteristics of muscarinic cholinergic, γ-aminobutyric acid A and phencyclidine receptors in spontaneously epileptic rats; in vitro quantitative autoradiographic analysis

Characteristics of muscarinic cholinergic (mACh), γ-aminobutyric acid A (GABA A) and phencyclidine (PCP) receptors in the spontaneously epileptic rats (SER), which exhibit both absence-like seizures and tonic convulsion, were examined using in vitro quantitative autoradiography. Computer analysis using autoradiographic technique revealed that the amount of the specific binding of [ 3H]quinuclidinyl benzilate (QNB) to mACh receptors in the striatum of SER was more than that of zitter rats, not exhibiting both seizures and convulsion. However, the specific bindings of [ 3H]muscimol and [ 3H]N-(2-thienyl)cyclohexyl)3,4-piperidine (TCP) to GABA A and PCP receptors, respectively, of SER were not different from those of zitter rats in various regions tested. These results suggest that hyperfunction of mACh receptors in the striatum is involved in the appearance of absence-like seizures and tonic convulsion of SER.

Sigma receptors. Putative links between nervous, endocrine and immune systems.

The sigma receptor is a neuronal substrate that binds several psychoactive compounds. These include cocaine, some steroids, dextromethorphan, phencyclidine (PCP), and benzomorphans such as pentazocine and N-allyl-normatezocine (SKF-10047). Many newer atypical antipsychotic drugs also bind to the sigma receptor. The sigma receptor, however, is not the PCP receptor. The sigma receptor exists in the central nervous system, endocrine, immune and certain peripheral tissues. Progesterone and certain steroids have been shown to represent endogenous ligands for the sigma receptor. The sigma receptor resides likely in the nonsynaptic region of the plasma membrane. The sigma receptor exists in two forms: high-affinity and low-affinity. The solubilized sigma receptor retains all of the pharmacological characteristics of a membrane-bound receptor. A major physiological role of the sigma receptor may involve the modulation of a tonic potassium channel.

Kinetic mechanisms of glycine requirement for N-methyl- d-aspartate channel activation

Glycine potentiates N-methyl- d-aspartate (NMDA) receptor-mediated responses via its interaction with a strychnine-insensitive glycine recognition site. We have previously shown that the potent glycine receptor antagonist 7-chlorokynurenic acid (7C1-KYN) dose-dependently inhibits [ 3H]MK-801 binding to the PCP receptor and that this effect is reversed by glycine. [ 3H]MK-801 binding to the PCP receptor within the NMDA receptor-gated ion channel is a measure of channel activation. Association of PCP receptor ligands is biexponential with the fast component of binding serving as a marker of activated NMDA channels. In the present study we utilize 7C1-KYN as a probe of the kinetic mechanism of the glycine effect upon NMDA receptor functioning. In the presence of l-glutamate, incubation with 7Cl-KYN completely abolished the fast component of [ 3H]MK-801 association in 4 out of 5 experiments. In the fifth experiment where the fast component was detected, it accounted for less than half of that seen in the presence of l-glutamate alone. 7Cl-KYN-induced inhibition of the fast component of [ 3H]MK-801 association was reversed by the addition of glycine. Since the fast component represents ligand binding to the PCP receptor via the open NMDA channel, selective reduction of this component by 7Cl-KYN indicates that glycine receptor antagonists reduce the probability of channel opening, and also that the selective reduction in the component of [ 3H]MK-801 binding that manifest fast kinetics can serve as a marker for glycine antagonists.

Effects of sigma receptor ligands on schedule-controlled behavior of rats: relation to sigma and PCP receptor binding affinity.

Eleven drugs were examined for their ability to inhibit sigma and phencyclidine (PCP) receptor binding, as labelled by (+)[3H]-R-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine ((+)-3-PPP), [3H]ditolylguanidine (DTG), (+)[3H]N-allylnormetazocine (NANM) and [3H]1-(1-(2-thienyl)cyclohexyl)piperidine (TCP), in membrane preparations from whole rat brain. The same drugs were studied for their effects under a fixed-ratio (FR) schedule of food reinforcement in rats. The relative potency order of the drugs for decreasing FR responding was: haloperidol greater than (+)-3-PPP greater than (-)NANM greater than BMY 14802 greater than PCP greater than (+)NANM greater than DTG greater than rimcazole greater than JO 1783 greater than JO1784 greater than (-)butaclamol. The binding affinities of all 11 drugs for either the [3H]DTG, (+)[3H]-3-PPP, (+)[3H]NANM or [3H]TCP site did not correlate significantly with the potencies of the same drugs for decreasing FR behavior. Rimcazole, (+)-3-PPP and haloperidol, at behaviorally inactive doses, were studied for their effects as antagonists of the rate-decreasing effects of JO 1784, DTG and (+)NANM: rimcazole attenuated the effects of DTG and (+)NANM but not JO 1784; (+)-3-PPP attenuated the effects of (+)NANM but not JO 1784 and DTG; and haloperidol was devoid of antagonistic actions. Moreover, BMY 14802 did not attenuate the rate-decreasing effects of (+)-3-PPP. These results further indicate that it is difficult to distinguish between purported sigma agonist and antagonist drugs.

Descending systems activated by morphine (ICV) inhibit kainic acid (IT)-induced behavior

Modulation of spinal systems activated by N-methyl-D-aspartate (NMDA) and substance P administered IT have been an area of interest in several laboratories. In the present investigations, behavior induced by the excitatory amino acid kainic acid, but not quisqualate, is demonstrated to be modulated in a manner similar to that previously observed for NMDA. Biting, scratching and licking behavior was induced by IT injections of excitatory amino acids or substance P in mice. Behavior induced by kainic acid (IT) injection was inhibited in a dose-dependent manner by coadministration of morphine (ICV), norepinephrine (IT), N-ethyl carboxamidoadenosine (NECA) (IT) and agonists interacting at PCP receptors (IT). Kainic acid and NMDA differed, however, in that a dopaminergic agonist, apomorphine, inhibited kainic acid-, but not NMDA-induced behavior and a selective NMDA receptor antagonist inhibits NMDA-, but not kainic acid-induced behavior. Behavior induced by quisqualate (IT) was not inhibited by any treatment and may have nonspecific actions in this type of assay. Our observations support independent spinal sites of action for behavior induced by kainic acid and NMDA, but several similarities were observed in the modulation of spinal systems activated by these agents.

Dextromethorphan and phencyclidine receptor ligands: differential effects on K(+)- and NMDA-evoked increases in cytosolic free Ca2+ concentration.

The ability of dextromethorphan (DXM) and phencyclidine (PCP) receptor ligands to attenuate increases in cytosolic free Ca2+ concentration ([Ca2+]i) evoked by N-methyl-D-aspartate (NMDA) and high extracellular [K+] was examined using the fluorescent dye Fura 2 in cultured rat hippocampal pyramidal neurons. The DXM receptor ligand caramiphen (40 microM) reduced K(+)-evoked rises in [Ca2+]i to a greater extent than NMDA-evoked rises; the reverse was true for the PCP receptor ligands ketamine (10-40 microM) and dextrorphan (10 microM). DXM itself, which has affinity for both DXM and PCP receptors, reduced both K(+)- and NMDA-evoked increases in [Ca2+]i in a concentration-dependent manner. The results suggest that DXM receptor ligands may at least in part exert their known anticonvulsant and neuroprotective effects by reducing Ca2+ influx through voltage-activated Ca2+ channels.

A rapid method for evaluating the behavioral effects of phencyclidine-like dissociative anesthetics in mice.

A simple and rapid method for detecting the behavioral effects of phencyclidine and related dissociative anesthetics is described. Dissociative anesthetics such as phencyclidine (PCP) and dizolcipine, which bind with high affinities at N-methyl-D-aspartate (NMDA) coupled cation channels ("PCP receptors"), produced a dose-related increase in the percentage of mice that fell from a 1.5 cm deep circular arena mounted on a 60 cm platform. A similar behavior was not manifest by other classes of compounds examined including competitive NMDA antagonists, an antagonist at strychnine-insensitive glycine receptors, and sigma-receptor ligands with moderate to low affinities for PCP receptors. Pretreatment of mice with glycine reduced in a dose-dependent manner the percentage of falls elicited by a maximally effective dose of dizolcipine. This simple procedure may prove useful for both the rapid detection of dissociative anesthetics and evaluation of putative PCP antagonists.

Chronic treatment of rats with the specific σ ligand D-pentazocine fails to modulate dopamine D 2 and σ binding in brain

Drugs that bind at the σ site have been proposed to have psychotomimetic and/or antipsychotic activity that is independent from that mediated by the phencyclidine (PCP) receptor. Some evidence suggests that σ ligands may produce their effects by modulation of dopamine neurotransmission. The present study examined potential interactions between the σ and dopamine system following chronic treatment with d-pentazocine, a specific σ ligand that has negligible affinity for either PCP or dopamine receptors. Rats treated for one month with either saline or 10 mg/kg per day d-pentazocine revealed no significant differences in either the K D or B max of [ 3H]haloperidol-labeled σ or [ 3H]YM-09151-2-labeled dopamine D 2 receptors across multiple brain regions. These results are discussed in relation to the pharmacology of putative σ-receptor subtypes.

Differential interaction of phencyclidine-like drugs with the dopamine uptake complex in vivo.

The phencyclidine (PCP) derivative, [3H]N-[1-(2-benzo[b]thiophenyl)cyclohexyl]piperidine ([3H]BTCP), was used to label in vivo the dopamine uptake complex in mouse brain. The striatum accumulated the highest level of total and specific binding. Drugs which bind to the dopamine uptake site inhibited [3H]BTCP binding on an order similar to their in vitro affinities for the high-affinity [3H]BTCP site. Drugs which label selectively other monoamine uptake complexes. PCP, or sigma recognition sites were ineffective at doses up to 40 mg/kg. PCP bound to and dissociated from the dopamine uptake complex very rapidly. N-[1-(2-Thienyl)cyclohexyl]pideridine (TCP) and (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) had no effect at any time or at any dose. These results imply that the pharmacological effects of PCP are due to its simultaneous interaction with the dopamine uptake complex and the PCP receptor. Conversely, TCP and MK-801, which have the same behavioral properties as PCP, exert their action only through the interaction with the PCP receptor.

A study of cortical and hippocampal NMDA and PCP receptors following selective cortical and subcortical lesions

The neuronal localization of glutamate and phencyclidine (PCP) receptors was evaluated in the cerebral cortex and hippocampal formation of rat CNS using quantitative autoradiography. Scatchard analysis of [ 3H]glutamate binding in the cortex (layers I and II and V and VI) showed no difference in the total number of binding sites ( B max) or apparent affinity ( K d) 1 week, 1 month and 2 months following unilateral ibotenate lesions to nucleus basalis of Meynert (nbM) compared to the non-lesioned side. Quisqualic acid displacement of [ 3H]glutamate in layers I and II, 1 week following nbM destruction, revealed both high- and low-affinity binding sites (representing the quisqualate (QA) and N-methyl- d-aspartate (NMDA) sites, respectively). Compared to the control side, there was no difference in binding parameters for either of the receptors sites. In similarly lesioned animals, the NMDA receptor was specifically labelled with [ 3H]glutamate and the associated PCP receptor labelled with[ 3H] N-(1-[2-thienyl]cyclohexyl)3,4-piperidine ([ 3H]TCP) in adjacent brain sections. For both receptors, there was no change in the total number of binding sites in the cortex following destruction of nbM. On the other hand, virtually all binding to NMDA and PCP receptors was eliminated following chemical destruction of intrinsic cortical neurons. These results suggest that the NMDA/PCP receptor complex does not exist on the terminals of cortical cholinergic afferents. One week after knife cuts of the glutamatergic entorhinal pathway to the hippocampal formation only an approximate 10% reduction of NMDA and PCP receptors was seen in the dentate gyrus. Conversely, selective destruction of the dentate granule cells using colchicine caused a near identical loss of NMDA and PCP receptors (84% vs 92% respectively). It is concluded from these experiments that glutamate and PCP receptors exist almost exclusively on neurons intrinsic to the hippocampal formation and that no more than 10% of NMDA and PCP receptors exist as autoreceptors on glutamatergic terminals.

Neuropeptide Y and peptide YY do not bind to brain σ and phencyclidine binding sites

The binding of neuropeptide Y (NPY) and peptide YY (PYY) to σ and phencyclidine (PCP) receptors in brain membranes was studied. NPY and PYY at up to 500 nM did not bind to σ or PCP receptors under different binding conditions including different temperatures, membrane preparations, protease inhibitor and sources of the peptides. These findings are in contrast with the reported potent affinities of NPY and PYY for σ and PCP receptors by Roman et al. (1989, European J. Pharmacol. 174, 301).

Formula omitted] receptor complex modulation by neuropeptide Y and peptide YY in rat hippocampus in vitro

Neuropeptide Y (NPY) and peptide YY (PYY) are known to bind with high affinity to sigma (σ) and phencyclidine (PCP) binding sites in rat brain. The functional significance of these results was assessed by testing both peptides in an in vitro bioassay system used for studying the N- methyl- d-aspartate (NMDA) receptor and consisting of rat hippocampal slices preloaded with [ 3H]noradrenaline (NA) and maintained under superfusion. The addition of NMDA in the superfusion medium induced an efflux of [ 3H]NA from the slices and the presence of NPY and PYY produced an enhancement of the stimulating effect. These results suggest that NPY and PYY could have a modulatory role at the NMDA receptor complex through an interaction with the σ and/or PCP receptor.