Jean-Marc Kamenka

Interaction of phencyclidines with the muscarinic and opiate receptors in the central nervous system.

Phencyclidine (N-(l-phenylcyclohexyl)piperidine)was introduced ill the late 1950s as an intravenous general anesthesic that was non-toxic, non-flammable and produced minimal cardiorespiratory depression 9,13. These clinical advantages were unfortunately offset by its prolonged duration of action and psychotomimetic effects 1,5. properties that have contributed to the emergence of phencyclidine as a major drug of abuse in the United States z°,21. During 1975 in the city of Detroit alone, phencyclidine was reported to be the single most common drug in emergency overdose screening 4. Known on the streets as angel dust . angel mist . PCP. hog and peace pills. phencyclidine may be taken orally, parenterally, or inhaled in powdered form. It is present in a wide variety of hallucinogen preparations that are falsely labeled as pure LSD, mescaline, tetrahydrocannabinol, psilocybin, cocame, 3,4-methylenediox~amphetamine or a combination of these 1°,1~,23. Phencyctidine produces long-lasting psychosis thought to resemble schizophrenia more than any produced by other psychedelics 1~. Phencyclidine exaggerates psychopathology. Schizophrenics experience severe thought disorders and behavioral problems as long as one month after a single injection. Quiet mental patients become catatonic, reactive ones become overreactive and restless 2,t6. Phencyclidine is such an emerging drug problem that a full issue of Clinical Tox ico logy has recently been devoted to it 7. This paper analyzes the possible interactions of a series of molecules of the phencyclidine family with a series of neurotransmitter receptors. Brain tissue preparations as well as binding experiments were carried out as described by Young and Snyder 25 (glycine receptor), Zukin et al. 26 (y-aminobutyric acid receptor), Roberts z2 (glutamate receptor), Burt et al.Z (dopamine receptor), Bennett and Snyder a (.serotonin receptor), Yamamura and Snyder 24 (muscarimc cholinergic receptor), Bylund and Snyder 6 (fl-adrenergic receptor), and Pert and Snyded 9 (opiate receptor). Radioactively labelled effectors were obtained as follows:

[3H]TCP: a new tool with high affinity for the PCP receptor in rat brain.

PCP binding sites have previously been demonstrated in the central nervous system with [3H]PCP. We now describe the binding properties to rat brain membranes of [3H]TCP, a PCP derivative. It is very advantageous to use [3H]TCP instead of [3H]PCP for the 3 following reasons: (i) it has a better affinity (Kd = 7.4 nM) for PCP binding sites than PCP itself; (ii) it dissociates slowly from its binding sites (t 1/2 = 20 min); (iii) the non-specific binding component obtained with [3H]TCP is much lower than that found with [3H]PCP.

[3H]N-[1-(2-Benzo(b)thiophenyl) cycohexyl]piperidine ([3H]BTCP): a new phencyclidine analog selective for the dopamine uptake complex

A benzothiophenyl group instead of a phenyl ring on phencyclidine (PCP) yields a molecule N-[1-(2-benzo(b)thiophenyl)cyclohexyl]piperidine (BTCP), which is one of the more potent known dopamine (DA) uptake inhibitors (IC50 = 7 nM). This compound also has low affinity for the PCP receptor (K0.5 = 6 microM). The sodium-dependent [3H]BTCP binding to rat striatal membranes was investigated. [3H]BTCP bound to two different sites: one with very high affinity (Kd1 = 0.9 nM, Bmax1 = 3.5 pmol/mg protein) which paralleled the distribution of dopaminergic nerve endings and a second with lower affinity (Kd2 = 20 nM, Bmax2 = 7.5 pmol/mg protein). There was a good correlation between the abilities of drugs specific for the DA uptake complex and of PCP analogs to inhibit high affinity [3H]BTCP binding and [3H]DA synaptosomal uptake. This study also demonstrated that PCP interacts with the DA uptake site since it is a competitive inhibitor of high affinity [3H]BTCP binding. This site, however, is not the PCP receptor, which has a different pharmacological selectivity.

[3H]Thienyl-phencyclidine ([3H]TCP) binds to two different sites in rat brain. Localization by autoradiographic and biochemical techniques

A high affinity [3H]thienyl-phencyclidine ([3H]TCP) binding and its similarity to that of [3H]phencyclidine ([3H]PCP) have been demonstrated on whole rat brain homogenates. We now describe the regional distribution of the [3H]TCP binding sites in the rat brain with fixed sections and frozen slide-mounted sections visualized by autoradiography and with homogenates of 12 regions by direct binding experiments. The 3 techniques give a similar pattern for the [3H]TCP binding distribution and the biochemical study reveals that two distinct binding sites for [3H]TCP exist: one of high affinity (5-10 nM) in the forebrain, which should be responsible for the psychotropic effects and a second one of lower affinity (50-80 nM) in the hindbrain and the spinal cord, which should be involved in the extrapyramidal behavior induced by PCP and congeneers. Competition experiments have shown that muscarinic compounds interact only with the hindbrain receptor possibly in two different sites, although morphine interacts with a very low affinity with the forebrains high affinity receptor. Results obtained with SKF-10,047 (N-allylnormetazocine) seem to indicate that TCP and sigma-receptors are different.

Neuroprotective effects of a novel NMDA antagonist, Gacyclidine, after experimental contusive spinal cord injury in adult rats

The aim of this study was to analyze the optimal time-window for neuroprotection by a novel NMDA antagonist, Gacyclidine, after experimental spinal cord injury, in terms of its functional, histopathological and electrophysiological effects. This molecule has already demonstrated its capacity for reducing the extent of an ischemic lesion and is currently experimented in a clinical trial of spinal cord injury. In this study, the spinal cord of rats was damaged by a contusive method and the animals were treated by saline or 1 mg/kg of Gacyclidine i.v., 10, 30, 60 and 120 min after injury. The time-course of the motor score was evaluated on days 1, 7 and 18 after injury, and somatosensory evoked potentials were determined on day 20. The animals were then killed and the cross-sectional area of the spinal cord (at the epicenter of the injury, above and below the injury), was measured. Walking recovery was better (P<0.0125) in the group treated 10 min after injury than in the untreated injured animals after 18 days of injury. Motor performances were related to the preservation of a larger undamaged area of spinal cord at the level of the injury (P<0.0125). Somatosensory evoked potential amplitudes were also higher in this group. These results confirm that Gacyclidine attenuates spinal cord damage after an experimental spinal cord lesion. Recovery was better within the group treated 10 min after injury compared with the other groups, which certainly confirms that the acute time-course of glutamate release requires rapid pharmacological intervention to achieve good results.

Effects of thienylphencyclidine (TCP) on seizure activity and brain damage produced by soman in guinea-pigs : ECoG correlates of neurotoxicity

The capacity of thienylcyclohexylpiperidine (TCP), a non-competitive blocker of the N-methyl-D-aspartate (NMDA) receptor, to counteract the convulsant, lethal, and neuropathological effects of 2 x LD50 of soman (an irreversible inhibitor of cholinesterase) was investigated in guinea-pigs treated by pyridostigmine and atropine sulphate. The effects of a weak dose of TCP (1 mg/kg) used in the present study globally reproduced those previously obtained with a higher dose (2.5 mg/kg; [Neurotoxicology 15 (1994) 837]): TCP was again most protective when given curatively within the first hour of soman-induced seizures. In this condition, (a) paroxysmal activity ceased in 10-20 min, (b) all the animals survived, (c) the majority of them recovered remarkably well and did not show any brain damage 24 h after the intoxication, and (d) the minimal duration of seizure activity normally required for producing soman-induced brain damage in other pharmacological environments was increased from 10 to 40 min to 80 min. Strikingly, when TCP was given 120 min after seizure onset, it failed to show any anticonvulsant activity but still provided neuroprotection in the hippocampus. The present study also gives additional evidence (see [Neurotoxicology 21 (4) (2000) 521]) that in soman poisoning, (a) the development of brain damage depends on the occurrence of ECoG seizures, (b) the topographical distribution of lesions depends on seizure duration, and (c) an increase of the relative power in the lowest (delta) frequency band might be a reliable marker of neuronal degradation. All these findings confirm that (a) glutamatergic NMDA receptors are involved in the mechanisms of soman-induced seizures and brain damage, (b) non-competitive antagonists of NMDA receptors might be promising candidates for post-treatment of soman poisoning, and (c) ECoG parameters from ECoG tracings and power spectrum might serve as useful external predictors for soman-induced neuropathological changes.

N-[1-(2-thienyl)cyclohexyl]-piperidine (TCP) does not block kainic acid-induced status epilepticus but reduces secondary hippocampal damage.

Distant damage, localized in the CA3 and CA1 areas, was observed in the hippocampus of rats as a consequence of status epilepticus (SE) induced by the injection of 2.5 nmol of kainic acid (KA) into the amygdala. In animals pretreated with an intraperitoneal injection of the non-competitive antagonist of the N-methyl-D-aspartate receptor, N-[1-(2-thienyl)cyclohexyl]-piperidine (TCP) (20 mg/kg), distant neuronal damage was reduced (CA1 neurons were always spared) whereas the rats still developed SE with an earlier onset. These results demonstrate the protective effect of TCP and confirm that epileptic activity and brain damage may be dissociated by NMDA receptor antagonists.

Localization of dopamine carriers by BTCP, a dopamine uptake inhibitor, on nigral cells cultured in vitro

BTCP, N-[1-(2-benzo(b)thiopenyl)cyclohexyl]piperidine, a derivative of phencyclidine, acted as a potent dopamine (DA) uptake blocking agent on primary cultures of dopaminergic neurons obtained from substantia nigra (IC50 = 70 nM). This value was closely related to IC50 determined for reference DA uptake inhibitors such as nomifensine (70 nM) or benztropine (50 nM), showing the specificity of BTCP towards the DA carrier. Thus, we used BTCP as a tool to visualize the DA uptake complexes on cultures, a model which preserves the integrity of the neurons. The [3H]BTCP binding sites directly visualized by radioautographical (RAG) labelling seemed to follow the fibres (axons or dendrites) of neurons in culture whereas the cell bodies were not labelled. The [3H]DA uptake visualized by RAG labelling, was inhibited either partially by BTCP at a concentration near its IC50 or totally by a high concentration of BTCP, all over the dopaminergic neurons (neurites and somas) immunostained with an anti-DA antiserum. Thus, the distribution of DA carriers can be investigated by a suitable tool, BTCP, a powerful and selective DA uptake blocker. These carriers have been visualized by radioautography with tritiated BTCP along the neurites, and the uptake can be totally blocked by a high concentration of BTCP all over DA neurons in vitro.

Homochiral structures derived from 1-[1-(2-thienyl)cyclohexyl]piperidine (TCP) are potent non-competitive antagonists of glutamate at NMDA receptor sites

Abstract The racemates of cis (pip/Me) 1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine and cis (pip/Me) 1-[1-(2-furanyl)-2-methylcyclohexyl]piperidine, 2 derivatives of the noncompetitive N -methyl- d -aspartate (NMDA) antagonist 1-[1-(2-thienyl)cyclohexyl]piperidine (TCP) have been synthesized. The corresponding isomers were resolved by a crystallization procedure, via the diastereoisomeric salts formed with (+)- and (-)-di- O,O ′-4-toluoyltartaric acid. The absolute configuration was determined by single-crystal X-ray analysis and by comparison with previously published data in related series. The 4 homochiral structures obtained were tested for their affinity for the PCP receptor sites labelled with [ 3 H]TCP. Their neuroprotective potency was assessed in primary cultured neuronal cells against neurotoxicity induced by glutamate. The 2 (-)-1 S , 2 R -isomers displayed the highest affinity for the PCP receptor sites and the highest efficacy for neuronal protection in comparison with the (+)-1 R , 2 S -isomers.

N-[1-(2-Benzo(tb)thiophenyl)cyclohexyl]piperidine (BTCP) and cocaine induce similar effects on striatal dopamine: a microdialysis study in freely moving rats

N-[1-(2-Benzo(b)thiophenyl)cyclohexyl]piperidine (BTCP) and cocaine inhibit dopamine (DA) uptake but bind to different sites on the transporter. Their dose-dependent effects (i.p. administration) on extracellular DA levels in the rat striatum were measured by in vivo microdialysis. Both drugs dose-dependently increased DA levels with a maximum effect 60 min post injection. BTCP (20 mg/kg) had a greater peak effect than cocaine (40 mg/kg). For doses inducing similar behavioral effects (cocaine, 20mg/kg; BTCP, 10 mg/kg) similar DA increases were observed in the striatum and the nucleus accumbens. Although both drugs bind on the DA transporter on different sites and induce different behavioral effects when administered chronically, their acute administration increased striatal DA level in a similar way.