piperidines and gacyclidine

The central nervous system of adult mammals has been classically considered as structurally rigid, tightly wired, and unable to be repaired. We have shown that there exists a rather considerable degree of intrinsic plasticity due to the neurons themselves, but merely to glial cells and to multipotent stem cells. The spinal cord constitutes a good model on which we could demonstrate, with vascular and traumatic animal paradigms, that an early pharmacologic intervention could reduce significantly the extent of lesions and the subsequent functional deficit. Moreover, we showed that regeneration of severed central axons could occur, provided that the astrocytes' component of the glial scar was modified. Finally, transplants of embryonic neurons were shown to repair the axonal circuitry below a sectioned cord, and to restore reflex functions. All these data point to unprecedented perspectives of efficient therapies in acute and chronic neurological diseases.

Topics: Animals; Axons; Cyclohexanes; Cyclohexenes; Fetal Tissue Transplantation; Male; Mammals; Mice; Mice, Knockout; Models, Animal; Nerve Regeneration; Neuronal Plasticity; Neurons; Neuroprotective Agents; Piperidines; Rats; Spinal Cord Injuries

Gacyclidine is a new phencyclidine derivative with neuroprotective properties. Tritiated gacyclidine and its enantiomers bind to NMDA receptors with binding parameters similar to those of other non-competitive NMDA receptor antagonists. The (-)enantiomer, (-)GK11, exhibits an affinity (2.5 nM) similar to that of dizocilpine (MK-801), while the (+)enantiomer, (+)GK11, has a 10 times lower affinity. When its interaction with NMDA receptors is prevented, gacyclidine binds also to "non-NMDA" binding sites which are mainly located in the molecular layer of the cerebellum on the dendritic tree of Purkinje cells. These binding sites do not appear to be related to any known neurotransmitters. In primary cortical cultures, gacyclidine and its enantiomers, at 0.1 to 5.0 microM, prevent glutamate-induced neuronal death. In rats, in vivo neurotoxicity of gacyclidine is far low than that of MK-801. No necrotic neurons were detected in animals sacrificed at 18 or 96 h after treatment with gacyclidine (1, 5, 10 or 20 mg/kg i.v.). At the highest (20 mg/kg) but not the lower doses (1-100 mg/kg) electron microscopy revealed the presence of few cytoplasmic or intramitochondrial vacuoles. In soman-treated monkeys gacyclidine enhanced neuroprotective activity of "three drugs cocktail" (atropine + diazepam + pralidoxime). Moreover, in rats, gacyclidine exerts a dose- and time-dependent neuroprotection in three models of spinal cord lesion. Beneficial effects of gacyclidine include reduction of lesion size and improvement of functional parameters after injury. In traumatic brain injury models gacyclidine improves also behavioral parameters and neuronal survival. Optimal protection is obtained when gacyclidine is administered at 0 to 30 min after injury. It is, therefore, concluded that gacyclidine exhibits neuroprotective effects similar to those of other NMDA receptor antagonists, with the advantage of being substantially less neurotoxic maybe due to its interaction with "non-NMDA" binding sites.

Topics: Animals; Binding Sites; Cells, Cultured; Cyclohexanes; Cyclohexenes; Humans; Neuroprotective Agents; Piperidines; Receptors, N-Methyl-D-Aspartate; Stereoisomerism

Beaufour-Ipsen is developing gacyclidine (GK-11) for the potential treatment of traumatic brain injury. Phase II clinical trials of the compound for this indication had been completed as of October 1999 and the company is looking for a partnership before undertaking further clinical development for this and, possibly, other indications [344879], [346265], [386763]. Phase II trials for acute spinal cord injury gave disappointing results and development for this indication has been discontinued [344879].

Topics: Animals; Clinical Trials as Topic; Contraindications; Cyclohexanes; Cyclohexenes; Humans; Neuroprotective Agents; Piperidines; Receptors, N-Methyl-D-Aspartate; Structure-Activity Relationship

Today, organophosphorus nerve agents are still considered as potential threats in both military or terrorism situations. These agents act as potent irreversible inhibitors of acetylcholinesterase in both central and peripheral nervous systems. Conventional treatment of organophosphate poisoning includes the combined administration of a cholinesterase reactivator (an oxime), a muscarinic cholinergic receptor antagonist (atropine) and a benzodiazepine anticonvulsant (diazepam). However, numerous studies have demonstrated that the excitatory amino acid glutamate also plays a prominent role in the maintenance of organophosphate-induced seizures and in the subsequent neuropathology especially through an overactivation of the N-methyl-D-aspartate (NMDA) receptor subtype. Contrary to other non-competitive NMDA antagonists successfully tested in rodents exposed to organophosphate, gacyclidine is a novel antiNMDA compound which is in the process of approval for human use in France for neurotraumatology. This review summarizes the therapeutic value of gacyclidine as a complement to the available emergency treatment against severe organophosphate poisoning. Previous data obtained from experiments on primates in several scenarios mimicking military or terrorist attacks, using soman as the nerve agent, were used. Primates pretreated with pyridostigmine and receiving conventional emergency therapy at the first signs of poisoning survive. However, only gacyclidine is able to ensure complete management of nerve agent poisoning for rapid normalization of EEG activity, clinical recovery and neuroprotection. Gacyclidine also ensures optimal management of severe nerve agent poisoning in animals neither pretreated nor receiving emergency therapy likewise during an unexpected exposure. However, this beneficial effect is obtained provided that medical intervention is conducted rapidly after intoxication. Globally, the current lack of any other NMDA receptor antagonist suitable for human use reinforces the therapeutic value of gacyclidine as a central nervous system protective agent for the treatment of OP poisoning.

Topics: Animals; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Haplorhini; Humans; N-Methylaspartate; Organophosphorus Compounds; Piperidines; Receptors, N-Methyl-D-Aspartate; Seizures; Soman; Time Factors

The aim of this study was to assess the safety and efficacy of intravenous (IV) injections of gacyclidine, a novel NMDA receptor antagonist, for neurological and functional recovery following acute traumatic brain injury. This multicenter, prospective, randomized, placebo-controlled, double-blind study compared four parallel groups. Two IV doses were administrated (placebo, 2x0.005mg/kg, 2x0.001mg/kg, 2x0.02mg/kg): the first dose was given within 2 hours following the trauma, and the second dose 4 hours after the first. Fifty-one patients were enrolled and 48 studied between March 1995 and June 1997 in France. Evaluation criteria for safety were physical examination, cardiovascular parameters, blood chemistry, hematology, ECG, and neuropsychological changes monitored after medication. Primary evaluation criteria for efficacy was the Glasgow coma scale complemented by the initial CT-scan and Glasgow outcome scale, motor deficiencies, neuropsychological changes, and functional indenpendence at D90 and D365 or endpoint. Intracranial pressure (ICP) monitoring was not taken into account because all the clinical centers participating in this study did not use this technique in daily practice during the inclusion period. Twelve patients died during the follow-up period, none of these deaths being related to the drug. Serious adverse events (181) were reported by most of the patients with no significant differences between groups. Only 10 of these adverse events were considered to be drug-related. Safety-related laboratory tests did not show any relevant changes. Concerning efficacy, the predefined prognostic factors (initial CT-scan score, initial Glasgow Coma Scale and occurrence of low systolic blood pressure during the first 24 hours) largely determinated the patient's outcome. When the prognostic factors were taken into account together with the dose level in a logistic regression model, gacyclidine showed a beneficial long-term effect and a best dose-result in the 0.04mg/kg treated group. Data obtained in this clinical trial appeared sufficient to warrant a European multicenter study on gacyclidine using the same evaluation criteria and ICP monitoring.

Topics: Adolescent; Adult; Aged; Blood Pressure; Brain Injuries; Cyclohexanes; Cyclohexenes; Double-Blind Method; Electrocardiography; Endpoint Determination; Female; Glasgow Coma Scale; Humans; Intracranial Pressure; Male; Middle Aged; Neuroprotective Agents; Neuropsychological Tests; Physical Examination; Pilot Projects; Piperidines; Prospective Studies; Tomography, X-Ray Computed; Treatment Outcome

Over the past decade, there has been a sharp increase in the number of newly identified synthetic drugs. These new drugs are often derivatives of previously abused substances but have unpredictable toxicity. One of these drugs is gacyclidine, a derivative of phencyclidine (PCP). Gacyclidine has been studied as a neuroprotective agent in trauma and as a therapy of soman toxicity. There are no previous reports of its use as a drug of abuse.. During a two-month period in the summer of 2013, a series of patients with severe agitation and end-organ injury were identified in an urban academic Emergency Department (ED). A urine drug of abuse screen was performed on all patients, and serum samples were sent for comprehensive toxicology analysis. A total of five patients were identified as having agitation, rhabdomyolysis, and elevated troponin (Table 1). Three of the five patients reported use of methamphetamine, and all five patients had urine drug screens positive for amphetamine. Comprehensive serum analysis identified methamphetamine in three cases, cocaine metabolites in one case, and a potential untargeted match for gacyclidine in all five cases. No other drugs of abuse were identified.. This is the first series of cases describing possible gacyclidine intoxication. The possible source of the gacyclidine is unknown but it may have been an adulterant in methamphetamine as all patients who were questioned reported methamphetamine use. These cases highlight the importance of screening for new drugs of abuse when patients present with atypical or severe symptoms. Gacyclidine has the potential to become a drug of abuse both by itself and in conjunction with other agents and toxicity from gacyclidine can be severe. It is the role of the medical toxicology field to identify new agents such as gacyclidine early and to attempt to educate the community on the dangers of these new drugs of abuse.

Topics: Academic Medical Centers; Adult; Combined Modality Therapy; Cyclohexenes; Designer Drugs; Drug Users; Emergency Service, Hospital; Fatal Outcome; Female; Humans; Illicit Drugs; Male; Middle Aged; Multiple Organ Failure; Neurotoxicity Syndromes; Piperidines; Psychomotor Agitation; Psychotropic Drugs; Rhabdomyolysis; Toxicokinetics; Treatment Outcome; Treatment Refusal

The NMDAR glutamate receptor subtype mediates various vital physiological neuronal functions. However, its excessive activation contributes to neuronal damage in a large variety of acute and chronic neurological disorders. NMDAR antagonists thus represent promising therapeutic tools that can counteract NMDARs' overactivation. Channel blockers are of special interest since they are use-dependent, thus being more potent at continuously activated NMDARs, as may be the case in pathological conditions. Nevertheless, it has been established that NMDAR antagonists, such as MK801, also have unacceptable neurotoxic effects. Presently only Memantine is considered a safe NMDAR antagonist and is used clinically. It has recently been speculated that antagonists that preferentially target extrasynaptic NMDARs would be less toxic. We previously demonstrated that the phencyclidine derivative GK11 preferentially inhibits extrasynaptic NMDARs. We thus anticipated that this compound would be safer than other known NMDAR antagonists. In this study we used whole-genome profiling of the rat cingulate cortex, a brain area that is particularly sensitive to NMDAR antagonists, to compare the potential adverse effects of GK11 and MK801. Our results showed that in contrast to GK11, the transcriptional profile of MK801 is characterized by a significant upregulation of inflammatory and stress-response genes, consistent with its high neurotoxicity. In addition, behavioural and immunohistochemical analyses confirmed marked inflammatory reactions (including astrogliosis and microglial activation) in MK801-treated, but not GK11-treated rats. Interestingly, we also showed that GK11 elicited less inflammation and neuronal damage, even when compared to Memantine, which like GK11, preferentially inhibits extrasynaptic NMDAR. As a whole, our study suggests that GK11 may be a more attractive therapeutic alternative in the treatment of CNS disorders characterized by the overactivation of glutamate receptors.

Topics: Animals; Astrocytes; Cyclohexenes; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Gyrus Cinguli; Inflammation; Injections, Intraperitoneal; Memantine; Microglia; Neurons; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Gacyclidine, a non-competitive NMDA receptor antagonist, is a phencyclidine derivative with neuroprotective properties. It has been previously safely administered intravenously to acute traumatic brain-injured patients. Experiments in guinea pigs have shown that local administration of gacyclidine to the cochlea can suppress salicylate-induced tinnitus. Thus, we thought that patients with therapy-resistant sensorineural tinnitus might benefit from a local therapy with gacyclidine. As a compassionate treatment, we administered aqueous gacyclidine solution via a Durect RWmuCath(TM) into the round window niche in six patients with unilateral deafness associated with tinnitus. The response of each patient to the drug treatment was given a numerical value by the use of a visual analogue scale (VAS) on a scale of 0-10 for tinnitus intensity, where 0 represented no tinnitus and 10 represented unbearable tinnitus-intensity or -annoyance (subjective). After constant perfusion of gacyclidine for 40-63 h, four out of six patients experienced a temporary relief from their tinnitus. No serious side effects were recorded in any of the cases. Gacyclidine might present a potent drug for the suppression of sensorineural tinnitus in humans and therefore should be considered for future double-blinded, placebo-controlled clinical trials. For lasting effective treatment, controlled intracochlear and long-term delivery of the drug seems to be necessary. Further studies investigating the toxicological effects of gacyclidine intracochlear perfusion as well as different dosages and therapy durations are under way to ensure the safety of the drug for long-term human use and warrant clinical trials.

Topics: Administration, Topical; Adolescent; Adult; Audiometry, Pure-Tone; Cochlea; Cyclohexenes; Female; Hearing Loss, Sensorineural; Humans; Male; Middle Aged; Neuroprotective Agents; Piperidines; Receptors, N-Methyl-D-Aspartate; Round Window, Ear; Tinnitus; Young Adult

Neuroprotective and repair strategies in spinal cord injuries (SCIs) have been so far largely unsuccessful. One of the prerequisites is the use of appropriate preclinical models to decipher pathophysiological mechanisms; another is the identification of optimal time windows for therapeutic interventions. The authors undertook this study to characterize early motor, sensory, autonomic, and histological outcomes after balloon compression of the spinal cord at the T8-9 level in adult rats.. A total of 91 rats were used in this study. Spinal cord balloon compression was performed at T8-9 in adult rats by inflation of a 2 Fr Fogarty catheter into the epidural space. The authors first characterized early motor, sensory, and autonomic outcomes of 2 volumes of compression (10 and 15 microl) using behavioral tests and then examined histological outcomes in the spinal cord using Luxol fast blue staining. To further substantiate the characterization of the epidural balloon-compression model, they used a noncompetitive N-methyl-D-aspartate antagonist, GK11, and demonstrated the involvement of excitotoxicity in this model.. Proportional and reproducible functional impairment resulted from compression caused by balloon inflation with either 10 or 15 microl of water and corresponded to the extent of the lesion. Indeed, during the early phase following SCI (1 week postinjury), recovery of locomotor function and bladder control correlated with the volume of inflation, whereas outcomes with respect to sensory function and reflexes were independent of compression severity. Treatment with GK11 significantly improved motor function in all groups of rats 1 week after injury and bladder voiding in the 10-microl injured rats compared to the 15-microl injured rats.. The results of this study demonstrate that spinal balloon-compression injury in the rat is a well-characterized, reproducible, and predictable model to analyze early events following SCI.

Topics: Animals; Autonomic Nervous System; Catheterization; Cyclohexenes; Disease Models, Animal; Excitatory Amino Acid Antagonists; Male; Motor Activity; N-Methylaspartate; Piperidines; Rats; Rats, Wistar; Reflex; Sensory Receptor Cells; Spinal Cord; Spinal Cord Compression; Spinal Cord Injuries; Urination

There is great interest in testing neuroprotectants which inhibit the neurodegeneration that results from excessive activation of N-methyl-D-aspartate (NMDA) receptors. As an alternative to in vivo testing in animal models, we demonstrate here the use of a complex in vitro model to compare the efficacy and toxicity of NMDA receptor inhibitors. Organotypic hippocampal slice cultures were used to compare the effectiveness of the Alzheimer's disease drug, memantine, the Parkinson's disease drug, procyclidine, and the novel neuroprotectant, gacyclidine (GK11), against NMDA-induced toxicity. All three drugs are non-competitive NMDA receptor open-channel blockers that inhibit excitotoxic injury, and their neuroprotective capacities have been extensively investigated in vivo in animal models. They have also been evaluated as potential countermeasure agents against organophosphate poisoning. Quantitative densitometric image analysis of propidium iodide uptake in hippocampal regions CA1, CA3 and DG, showed that, after exposure to 10microM NMDA for 24 hours, GK11 was the most potent of the three drugs, with an IC50 of about 50nM and complete protection at 250nM. When applied at high doses, GK11 was still the more potent neuroprotectant, and also the least cytotoxic. These findings are consistent with those from in vivo tests in rodents. We conclude that the slice culture model provides valuable pre-clinical data, and that applying the model to the screening of neuroprotectants might significantly limit the use of in vivo tests in animals.

Topics: Animals; Clinical Trials as Topic; Cyclohexenes; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Neuroprotective Agents; Piperidines; Propidium; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

To date, there has been no efficient therapeutic approach to spinal cord injuries (SCIs). This may be attributable, at least in part, to difficulties in forming predictive and accurate experimental animal models. The authors' previous studies have identified 2 relevant conditions of such a model. The first condition is the ability to compare data derived from rat models of SCI by developing mouse models of SCI that permit access to a large range of transgenic models. The second condition is that the exploration of the consequences of each mechanism of spinal trauma requires modeling the different etiologic aspects of the injury.. To fulfill these 2 conditions a new model of mouse spinal cord compression injury was devised using a thread-driven olive-shaped compressive device. The authors characterized early motor, sensory, and histological outcomes using 3 olive diameters and different compression durations.. A gradual and reproducible functional severity that correlated with lesion extension was demonstrated in 76 mice. To further substantiate the characterization of this model, a noncompetitive N-methyl-d-aspartate antagonist was administered in 30 mice, which demonstrated the involvement of excitotoxicity in this model.. The study demonstrated that spinal olive-compression injury in the mouse is a reproducible, well-characterized, and predictable model for analyzing early events after SCI. The nonmagnetic and remotely controlled design of this model will allow completion of the lesion while the animal is in the MR imaging apparatus, thus permitting further real-time MR imaging studies that will provide insights into the characterization of early events in the spatial and temporal evolution of SCI. Moreover, this model lays the foundation for future in vivo studies of functional and histological outcomes following SCI in genetically engineered animals.

Topics: Animals; Behavior, Animal; Cyclohexenes; Disease Models, Animal; Female; Magnetic Resonance Imaging; Mice; Mice, Inbred C57BL; Motor Activity; Neuroprotective Agents; Piperidines; Receptors, N-Methyl-D-Aspartate; Recovery of Function; Spinal Cord Compression; Time Factors

Over-stimulation of NMDA receptors (NMDARs) is involved in many neurodegenerative disorders. Thus, developing safe NMDAR antagonists is of high therapeutic interest. GK11 is a high affinity uncompetitive NMDAR antagonist with low intrinsic neurotoxicity, shown to be promising for treating CNS trauma. In the present study, we investigated the molecular basis of its interaction with NMDARs and compared this with the reference molecule MK801. We show, on primary cultures of hippocampal neurons, that GK11 exhibits neuroprotection properties similar to those of MK801, but in contrast with MK801, GK11 is not toxic to neurons. Using patch-clamp techniques, we also show that on NR1a/NR2B receptors, GK11 totally blocks the NMDA-mediated currents but has a six-fold lower IC(50) than MK801. On NR1a/NR2A receptors, it displays similar affinity but fails to totally prevent the currents. As NR2A is preferentially localized at synapses and NR2B at extrasynaptic sites, we investigated, using calcium imaging and patch-clamp approaches, the effects of GK11 on either synaptic or extrasynaptic NMDA-mediated responses. Here we demonstrate that in contrast with MK801, GK11 better preserve the synaptic NMDA-mediated currents. Our study supports that the selectivity of GK11 for NR2B containing receptors accounts contributes, at least partially, for its safer pharmacological profile.

Topics: Animals; Cells, Cultured; Cyclohexenes; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Hippocampus; Mice; Neuroprotective Agents; Piperidines; Rats; Receptors, N-Methyl-D-Aspartate

Exposure to the organophosphate nerve agent soman produces seizures that in turn lead to neuropathology. This study describes the temporal and spatial evolution of brain pathology following soman-induced convulsions and the attenuation of these alterations after neuroprotective intervention with magnetic resonance imaging (MRI). Neuroimaging 12 h after soman exposure, the hippocampus and thalamus exhibited significant decreases (23%) in apparent diffusion coefficients (ADC). These acute effects were resolved by 7 days. In addition, T2 measurements declined significantly at 12 h (37%) returning to near normal values by 24 h. Histopathological analyses confirmed moderate cell loss within the hippocampus and piriform cortex. Together these findings suggest that initial cell death was resolved through regional cellular remodeling. Pharmacological countermeasures were administered in the form of diazepam, a benzodiazepine anticonvulsant, or gacyclidine (GK-11), an anti-glutamatergic compound. Diazepam therapy applied immediately after soman exposure prevented acute ADC changes. However the presence of edema, using T2 measurements, was detected at 3 h within the retrosplenial, amygdala and piriform cortices and at 12 h in the thalamus (34% below normal). GK-11 therapy appeared to prevent most of these changes. However at 7 days after soman, a decrease (17%) in ADC was observed in the piriform cortex. Pathology was confined to the piriform cortex suggesting that this region is more difficult to protect. This is the first report that provides temporal and spatial resolution using MRI with histological correlation of pharmacological interventions against soman-mediated seizure-induced neuropathology.

Topics: Animals; Anticonvulsants; Brain; Cell Death; Cyclohexanes; Cyclohexenes; Diazepam; Diffusion Magnetic Resonance Imaging; Magnetic Resonance Imaging; Male; Neuroprotective Agents; Piperidines; Rats; Rats, Sprague-Dawley; Seizures; Soman; Status Epilepticus

Prenatal infection constitutes an important risk factor for brain injury, in both premature and full-term infants. Unfortunately, as the mechanisms involved are far from understood, no therapeutic strategy emerges to prevent the damage. We tested the hypothesis that administration of lipopolysaccharide (LPS) to gravid female rats enhanced glutamate-induced oxidative stress in brain of pups. A microdialysis probe was implanted into the striatum of 14-day-old animals and the release of hydroxyl radicals (.OH) in the perfusion medium was evaluated. Glutamate promoted a delayed.OH release in the offspring of dams given LPS, contrasting with the.OH decreases observed in control animals. A similar response occurred after infusion of (R,S)-3,5-dihydroxyphenylglycine (DHPG), a Group I metabotropic glutamate receptor (mGluR) agonist. This response was not consecutive to a remote activation of N-methyl-D-aspartate (NMDA) receptors, as it was unaffected by an NMDA receptor antagonist. Furthermore, the response to NMDA itself decreased in the offspring of dams given LPS. Massive amounts of DHPG, however, likely internalizing the mGlu receptor, still blunted the response to NMDA, as in controls. No quantitative variation occurred in mGluR1, mGluR5, or the NR1 subunit of the NMDA receptor between controls and neonates born from LPS-treated dams. Direct LPS injection into age-matched pups, by contrast, affected the response to neither glutamate nor DHPG. These results confirm that normally during perinatal development, the brain is protected from any oxidative stress resulting from excess glutamate, and the results support the hypothesis that maternal infection before delivery may lead to critical brain damage via the release of toxic free radicals.

Topics: 2,3-Diketogulonic Acid; Aging; Animals; Animals, Newborn; Brain; Cell Membrane; Chromatography, High Pressure Liquid; Cyclohexanes; Cyclohexenes; Female; Fetal Diseases; Functional Laterality; Glutamic Acid; Hydroxyl Radical; Lipopolysaccharides; Male; Methoxyhydroxyphenylglycol; Neuroprotective Agents; Oxidative Stress; Piperidines; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Wistar; Receptors, Metabotropic Glutamate; Subcellular Fractions; Time Factors

TCP and its derivative gacyclidine (+/- GK11) are high-affinity non-competitive antagonists of N-methyl-D-aspartate (NMDA) receptors (NMDARs) and as such exhibit significant neuroprotective properties. These compounds also bind with a low affinity to binding sites whose pharmacological profiles are different from that of NMDARs. With the intention to develop new strategies of neuroprotection, we found it mandatory to investigate whether 1-[1-(2-thienyl)cyclohexyl]piperidine (TCP) and gacyclidine low-affinity sites are similar. The effects of several drugs selective for either NMDARs or the [(3)H]TCP low-affinity site (or PCP(3) site) on (+), (-)[(3)H]GK11 and [(3)H]TCP specific binding were investigated. Competition experiments on cerebellum homogenates revealed substantial differences between the pharmacological profiles of the PCP(3) site and that of gacyclidine's enantiomers low-affinity sites. Under experimental conditions preventing the interaction of the radioligands with NMDARs, the autoradiographic study showed, however, that the distributions of both [(3)H]TCP and (-)[(3)H]GK11 specific binding were similar. The specific labelling was low and uniform in telencephalic structures, whereas in the cerebellum it was higher in the molecular than in the granular layer. Finally, the analysis of competition experiments performed on tissues slices demonstrated that PCP(3) selective ligands were unable to prevent [(3)H]TCP or (-)[(3)H]GK11 binding to "non-NMDA" binding sites. As a whole, our data suggest that: (1) the different pharmacological profiles of [(3)H]TCP and [(3)H]gacyclidine enantiomers on low-affinity sites are due to their selectivity for specific NMDARs subpopulations; (2) the pharmacological isolation of TCP and gacyclidine "non-NMDA" binding sites is the most appropriate way to further study the low-affinity component of their specific binding. Obtaining reliable and specific pharmacological tools for those binding sites is of particular interest, since it is likely that they play a substantial role in the low neurotoxicity, and therefore tolerability, of gacyclidine, a new neuroprotective drug currently evaluated in clinical trials for the treatment of brain and spinal cord injuries.

Topics: Animals; Binding Sites; Binding, Competitive; Brain; Brain Injuries; Cerebellum; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Neurons; Neuroprotective Agents; Phencyclidine; Piperidines; Radioligand Assay; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Stereoisomerism; Subcellular Fractions; Synapses; Synaptic Transmission; Tritium

The pharmacokinetics of gacyclidine enantiomers, a noncompetitive N-methyl-D-aspartate (NMDA) antagonist, were studied in plasma and spinal cord extracellular fluid (ECF) after experimental spinal cord injury in rats. Spinal cord trauma was produced by introducing an inflatable balloon in the dorsal subdural space. Upon implantation of microdialysis probes in spinal cord (T9) and intravenous (iv) bolus administration of (+/-)-gacyclidine (2.5 mg/kg), concentrations in plasma and ECF were monitored over 5 h and analyzed by a stereospecific gas chromatography-mass spectrometry (GC-MS) assay. In plasma, concentrations of (+)-gacyclidine were approximately 25% higher than those of (-)-gacyclidine over the duration of the experiment and decayed in parallel (t(1/2 alpha) approximately 7 min; t(1/2 beta) approximately 90 min) with no significant difference between the two enantiomers. Clearance (CL) and volume of distribution (Vd) of (-)-gacyclidine were approximately 20% higher than those of its optical antipode (CL: 285 versus 236 mL. kg(-1). min(-1); Vd(beta): 39.3 versus 31.2 l/kg). Protein binding (approximately 91%) was not stereoselective. In spinal cord ECF, both enantiomers were quantifiable within 10 min after drug administration, and their concentration remained stable over the duration of the experiment in spite of changing blood concentrations. Repeated iv bolus injections of gacyclidine did not modify these profiles. Areas under the curves (AUCs) of concentration in ECF versus time were similar for both enantiomers and not correlated with AUCs in plasma. Penetration of (-)-gacyclidine was, however, significantly higher (approximately 30%) than that of (+)-gacyclidine. In summary, the disposition of gacyclidine enantiomers is stereoselective. Both enantiomers exhibit a high affinity for spinal cord tissue, and the drug exchange between plasma and spinal cord ECF involves an active transport system. These findings contribute to the explanation of the discrepancy between drug concentrations in plasma and spinal cord ECF.

Topics: Animals; Biological Transport, Active; Calibration; Cyclohexanes; Cyclohexenes; Disease Models, Animal; Dose-Response Relationship, Drug; Male; Neuroprotective Agents; Piperidines; Protein Binding; Rats; Rats, Wistar; Spinal Cord Injuries; Stereoisomerism; Tissue Distribution

Gacyclidine, a channel blocker of N-methyl-D-aspartate receptors (NMDAR), exhibits potent neuroprotective properties and a low self-neurotoxicity. Preventing its interaction with NMDARs we demonstrate, through the use of its enantiomers, that gacyclidine also interacts with other ('non-NMDA') binding sites. The autoradiographic study showed that these sites displayed a uniform specific binding in the forebrain and a more discrete distribution in the molecular layer of the cerebellum. The 'non-NMDA' binding sites could exert a modulatory control on glutamatergic neurotransmission.

Topics: Animals; Autoradiography; Binding Sites; Brain Chemistry; Cerebellum; Cyclohexanes; Cyclohexenes; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Glutamic Acid; Glycine; Male; Piperidines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Stereoisomerism; Tritium

Gacyclidine is a non-competitive NMDA receptor antagonist with potent neuroprotective properties. However, we have previously demonstrated that gacyclidine enantiomers [(-) and (+)GK11] interact with other ('non-NMDA') binding sites which may play a role in the lower self-neurotoxicity of this compound relative to the other NMDA receptor antagonists. Evidence for these binding sites has been obtained from autoradiographic and membrane binding experiments. They were found to be expressed at high levels in the molecular layer of the cerebellum, although they can also been seen in the granular layer and in telencephalic regions. The present study was designed to further characterize these gacyclidine 'non-NMDA' binding sites. The pharmacological profiles obtained on cerebellar and telencephalic membrane homogenates showed that they could not be linked directly to the main receptors or uptake complexes of the central nervous system (CNS). However, the comparison of (-) and (+)[(3)H]GK11 binding distribution in different mutant animals bearing specific cellular deficits in the cerebellum has demonstrated that the gacyclidine 'non-NMDA' binding sites are associated with the dendritic trees of Purkinje cells. Interestingly, our study also shows that the radioligand binding to both cerebellar and telencephalic structures could be modulated by endogenous factors which can be removed by a stringent prewashing procedure.

Topics: Animals; Binding Sites; Binding, Competitive; Cell Membrane; Cerebellum; Cyclohexanes; Cyclohexenes; Dendrites; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Neurologic Mutants; Neurotransmitter Agents; Organ Specificity; Piperidines; Purkinje Cells; Radioligand Assay; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Stereoisomerism; Telencephalon

The purpose of this study was to determine the pharmacokinetics of gacyclidine, a non-competitive NMDA antagonist, in plasma and spinal cord extracellular fluid (ECF) after IV administration of single enantiomers in rats. After implantation of microdialysis probes in spinal cord, concentrations in plasma and ECF dialysates were determined by a chiral GC/MS assay over 5 h after administration of either (+)-gacyclidine or (-)-gacyclidine (1.25 mg/kg). Plasma protein binding was estimated in vitro by equilibrium dialysis. Plasma concentrations decayed in parallel in a biphasic manner (t(1/2)alpha approximately 9 min; t(1/2)beta approximately 90 min) with no significant difference between the two enantiomers. Clearance of (+)-gacyclidine and (-)-gacyclidine (291 versus 275 ml/min per kg, respectively), volume of distribution (Vdbeta: 38 versus 40 l/kg), and protein binding (90 versus 89%) were not stereoselective. Both gacyclidine enantiomers were quantifiable in spinal cord ECF 10 min after drug administration and their concentrations remained stable over the duration of the experiment in spite of changing blood concentrations. Penetration of the two enantiomers in spinal cord ECF was similar although highly variable between animals. Exposure of spinal cord ECF was comparable for both enantiomers, and not correlated with plasma AUCs. This study showed the absence of any pharmacokinetic difference between the two enantiomers when administered individually, and no enantiomeric inversion. Both gacyclidine enantiomers penetrate rapidly and extensively into spinal cord ECF, and their distribution may involve an active transport system.

Topics: Animals; Blood Proteins; Calibration; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Extracellular Space; Half-Life; Injections, Intravenous; Male; Microdialysis; Piperidines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Stereoisomerism

Classical emergency treatment of organophosphate poisoning includes the combined administration of a cholinesterase reactivator (an oxime), a muscarinic cholinergic receptor antagonist (atropine) and a benzodiazepine anticonvulsant (diazepam). In subjects taking pyridostigmine and trained to autoadminister at least one autoinjector at the first signs of poisoning, classical emergency treatment ensures survival but only an antiglutamatergic compound like gacyclidine appears to be able to ensure optimal management of nerve agent poisoning in terms of rapid normalization of EEG activity, clinical recovery and total neuroprotection. All of this reinforces the therapeutical value of gacyclidine, which is in the process of approval for human use in France for treatment of head injury, as a central nervous system protective agent for the treatment of OP poisoning.

Topics: Atropine; Cholinesterase Reactivators; Cyclohexanes; Cyclohexenes; Electroencephalography; Excitatory Amino Acid Antagonists; Humans; Organophosphate Poisoning; Piperidines; Poisoning; Pyridostigmine Bromide; Self Administration

The aim of this study was to evaluate the efficacy, optimal dose, and optimal time-window of gacyclidine, a novel N-methyl-D-aspartate (NMDA) receptor antagonist, in terms of its functional, histopathological, and electrophysiological effects after experimental spinal cord injury. The spinal cord of rats was damaged by a photochemical method and the animals were treated by saline or gacyclidine at doses of 1, 2.5, or 5 mg/kg 10 min after injury or gacyclidine 1 mg/kg 10, 30, 60, and 120 min after injury. The time-course of the motor score (walking and inclined-plane stability) was evaluated until day 18, and somatosensory evoked potentials were determined on day 18. The animals were then sacrificed, 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 in most of the groups treated after injury than in the untreated injured animals. Motor performances were related to preservation of a larger undamaged area of spinal cord at the level of the injury and, interestingly, with prevention of extension of the anatomical lesion above the level of the injury. Somatosensory evoked potential amplitudes were often higher in treated groups. These results confirm that gacyclidine induces dose-dependent and time-dependent attenuation of spinal cord damage after an experimental vascular lesion. Although all three doses induced neuroprotective effects, recovery was greater and very homogeneous in the group treated with 1 mg/kg. Moreover, recovery was slightly better and more homogeneous within the groups treated 10 and 30 min after injury compared to the other groups. It appears that, according to the existing evidence, NMDA antagonists are an essential component in the elaboration of a neuroprotective strategy after spinal cord trauma.

Topics: Animals; Cyclohexanes; Cyclohexenes; Disease Progression; Dose-Response Relationship, Drug; Evoked Potentials, Somatosensory; Excitatory Amino Acid Antagonists; Light Coagulation; Male; Motor Activity; Neuroprotective Agents; Piperidines; Rats; Rats, Sprague-Dawley; Reaction Time; Receptors, N-Methyl-D-Aspartate; Spinal Cord; Spinal Cord Injuries; Spinal Cord Ischemia

Gacyclidine (1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine), the racemate of (+)-and (-)-GK11, exhibits potent neuroprotective properties due to its antagonism at the NMDA receptor. In its tritiated form, gacyclidine showed a binding distribution similar to that of NMDA receptors in the rat brain. With membrane preparations, the (-)-enantiomer of gacyclidine exhibited an affinity similar to that of MK-801 (dizocilpine, (+)-5-methyl-10,11-dihydro-5H-dibenzo[a, d]cyclohepten-5,10-imine) in the low nanomolar range, while the (+)-enantiomer was about 10 times less potent. Gacyclidine affinity was lower in the cerebellum than in the forebrain or the spinal cord. In this latter region and in the cerebellum, two binding sites were evidenced, one of which was a low-affinity site insensitive to MK-801. In all regions, PRE-084 (2-(4-morpholino)ethyl-1-phenylcyclohexane-1-carboxylate), a sigma receptor ligand, had no effect on [3H]gacyclidine binding.

Topics: Animals; Autoradiography; Cerebellum; Cyclohexanes; Cyclohexenes; In Vitro Techniques; Male; Membranes; Neuroprotective Agents; Organ Specificity; Piperidines; Radioligand Assay; Rats; Rats, Wistar; Spinal Cord; Telencephalon; Tritium

Gacyclidine, a novel, noncompetitive NMDA receptor antagonist, was injected (i.v.) into rats at three different doses to determine if the drug could promote behavioral recovery and reduce the behavioral and anatomical impairments that occur after bilateral contusions of the medial frontal cortex (MFC). In the Morris water maze, contused rats treated with gacyclidine at a dosage of 0.1 mg/kg performed better than their vehicle-treated conspecifics. Rats given gacyclidine at either 0.3 or 0.03 mg/kg performed better than brain-injured controls, but not as well as those treated with 0.1 mg/kg. Counts of surviving neurons in the nucleus basalis magnocellularis (NBM) and the medial dorsal nucleus (MDN) of the thalamus were used to determine whether gacyclidine treatment attenuated secondary cell death. In both the NBM and the MDN, the counts revealed fewer surviving neurons in untreated contused rats than in gacyclidine-treated rats. Increases in the size and number of microglia and astrocytes were observed in the striatum of gacyclidine-treated contused brains. Although most consequences of MFC contusions were attenuated, we still observed increases in ventricle dilation and thinning of the cortex. In fact, the ventricles of rats treated with 0.1 mg/kg of gacyclidine were larger than those of their vehicle treated counterparts, although we observed no behavioral impairment.

Topics: Animals; Basal Nucleus of Meynert; Brain Injuries; Cell Count; Cerebral Cortex; Cerebral Ventricles; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Frontal Lobe; gamma-Aminobutyric Acid; Male; Maze Learning; Microglia; Motor Activity; Neurons; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Recovery of Function; Swimming

Gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) is a TCP derivative, which exhibits potent neuroprotective properties against glutamate-induced neurotoxicity in vitro and in vivo. In order to better understand gacyclidine pharmacological properties, the binding parameters of its enantiomers ((-) and (+)[3H]GK11) were determined in the rat central nervous system (CNS). An autoradiographic study has shown that their binding distributions are correlated with those of N-methyl-D-aspartate (NMDA) receptors throughout the CNS. Globally, the labeling was the highest with (-)[3H]GK11. In the cerebellum, both radioligands similarly labeled the molecular layer. For both radioligands, on telencephalic, cerebellum and spinal cord homogenates, the association and dissociation kinetics were accounted for by multiphasic process. In all regions, (-)[3H]GK11 exhibited the highest affinity in the nanomolar range. The pharmacological study revealed that both enantiomers labeled both high and low affinity sites in all regions. The pharmacological profile of high affinity sites was correlated with those of NMDA receptors. Those of low affinity sites were different in telencephalic and cerebellar homogenates. Overall, this study showed that low affinity sites might constitute a heterogeneous population, which could include sigma receptors in the cerebellum. The autoradiographic study has shown that these sites may be located in the molecular layer. The contribution of low affinity sites to the neuroprotective properties of gacyclidine remains to be investigated.

Topics: Animals; Binding Sites; Binding, Competitive; Central Nervous System; Cerebellum; Cyclohexanes; Cyclohexenes; In Vitro Techniques; Kinetics; Male; Neuroprotective Agents; Piperidines; Prosencephalon; Radioligand Assay; Rats; Rats, Wistar; Spinal Cord; Stereoisomerism; Subcellular Fractions; Tritium

Determination of the pharmacokinetics of gacyclidine enantiomers, a non-competitive NMDA antagonist, in plasma and spinal cord extracellular fluid (ECF) of rats.. Implantation of microdialysis probes in spinal cord (T9). Serial collection of plasma samples and ECF dialysates over 5 hours after IV bolus administration of (+/-)-gacyclidine (2.5 mg/kg). Plasma protein binding determined in vivo by equilibrium dialysis. Chiral GC/ MS assay.. Plasma concentrations of (+)-gacyclidine were approximately 25% higher than those of (-)-gacyclidine over the duration of the experiment in all animals. Plasma concentrations decayed in parallel in a biphasic manner (t1/2alpha approximately 9 min; t1/2beta approximately 90 min) with no significant difference between enantiomers. Clearance and volume of distribution of (-)-gacyclidine were approximately 20% higher than those of its optical antipode (CL: 248 vs 197 ml.kg(-1)x min(-1); Vdbeta: 31.6 vs 23.5 1/kg). Protein binding (approximately 90%) was not stereoselective. Both gacyclidine enantiomers were quantifiable in spinal cord ECF 10 min after drug administration and remained stable over the duration of the experiment in spite of changing blood concentrations. Penetration of (-)-gacyclidine was significantly higher (approximately 40%) than that of (+)-gacyclidine in all animals. Yet, exposure of spinal cord ECF was similar for both enantiomers, and not correlated with plasma AUCs.. The disposition of gacyclidine enantiomers is stereoselective. Both enantiomers exhibit a high affinity for spinal cord tissue and their distribution may involve a stereoselective and active transport system. This hypothesis could also explain the discrepancy between drug concentrations in plasma and spinal cord ECE

Topics: Animals; Blood Proteins; Catheterization; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Extracellular Space; Injections, Intravenous; Male; Microdialysis; Piperidines; Protein Binding; Rats; Rats, Wistar; Spinal Cord; Stereoisomerism

Gacyclidine, a new neuroprotectant aimed to non-competitively antagonize N-methyl-D-aspartate (NMDA) receptors, and dizocilpine blocked the hydroxyl radical response to toxic amounts of glutamate, perfused through a microdialysis implanted in the striatum of conscious rats. Furthermore, the hydroxyl radical response resulting from the infusion of lower doses of glutamate, which could not be inhibited by the same amounts of dizocilpine nor by acute alcohol exposure, still remained sensitive to gacyclidine inhibition. Thus, oxidative stress resulting from a glutamate discharge involves the activation of both NMDA receptors, and of non-NMDA mechanism(s) which, with moderate glutamate levels, were still antagonized by gacyclidine. Enhanced blockage of toxic hydroxyl radicals might explain the different and possibly higher neuroprotective property of gacyclidine as compared with other non-competitive NMDA antagonists.

Topics: Animals; Binding, Competitive; Corpus Striatum; Cyclohexanes; Cyclohexenes; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Glutamates; Hydroxyl Radical; Male; Microdialysis; N-Methylaspartate; Neuroprotective Agents; Oxidative Stress; Piperidines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

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.

Topics: Animals; Contusions; Cyclohexanes; Cyclohexenes; Electrophysiology; Evoked Potentials, Somatosensory; Excitatory Amino Acid Antagonists; Male; Motor Activity; N-Methylaspartate; Neuroprotective Agents; Piperidines; Rats; Rats, Sprague-Dawley; Spinal Cord; Spinal Cord Injuries; Time Factors; Weight Gain

Prevention of the immediate excitotoxic phase occurring in response to spinal cord injury (SCI) is a major issue to reduce the neuronal damage responsible for any ensuing motor deficits. The present study evaluated the neuroprotective efficacy of three noncompetitive NMDA receptor antagonists: Gacyclidine (GK-11), a new compound, Dizocilpine (MK-801), and Cerestat (CNS-1102) in a rat spinal cord contusion model. To mimic human SCI, a standardized model of rat spinal cord closed contusion in which animals spontaneously and progressively recover from the induced paraplegia was employed. Such model, characterized by a slow recovery of hindlimb locomotor function enables easy quantification of the neuroprotection at both the behavioral and cellular level. The animals were treated intravenously with the respective drugs 10 min after the spinal contusion. The dose range study suggested that 1 mg/kg of Gacyclidine was the most effective dose to promote functional recovery in reducing by half the time needed to reach full locomotor recovery. Racemate and enantiomers of Gacyclidine showed similar neuroprotective effects, but treatment with the enantiomers were not as efficacious in promoting full functional recovery. Similarly, a prolonged treatment with the racemate was not as efficious as a single dose, suggesting that a prolonged blockade of the amino-excitatory neurotransmission may be deleterious. Finally, Dizocilpine and Cerestat treatments induced only a partial and delayed neuroprotective effect compared to Gacyclidine. Neuroprotection characterized by a reduction of the cystic cavity and of the astrogliosis was observed with all treatments. As Gacyclidine is already in clinical trials, the present findings suggest the premise that it is a promising agent for limiting the initial neuronal damage induced by CNS trauma leading to better functional recovery.

Topics: Animals; Astrocytes; Cyclohexanes; Cyclohexenes; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Administration Schedule; Excitatory Amino Acid Antagonists; Female; Gliosis; Guanidines; Neuroprotective Agents; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Recovery of Function; Spinal Cord; Spinal Cord Injuries; Stereoisomerism

Organophosphorus (OP) nerve agents are still used as warfare and terrorism compounds. Classical delayed treatment of victims of organophosphate poisoning includes combined i.v. administration of a cholinesterase reactivator (an oxime), a muscarinic cholinergic receptor antagonist (atropine) and a benzodiazepine anticonvulsant (diazepam). The objective of this study was to evaluate, in a realistic setting, the therapeutic benefit of administration of GK-11 (gacyclidine), an antiglutamatergic compound, as a complement to the above therapy against organophosphate poisoning. Gacyclidine was injected (i.v.) in combination with atropine/diazepam/pralidoxime at man-equivalent doses after a 45- or 30-min latency period to intoxicated primates (2 LD50). The effects of gacyclidine on the animals' survival, electroencephalographic (EEG) activity, signs of toxicity, recovery after challenge and central nervous system histology were examined. The present data demonstrated that atropine/diazepam/pralidoxime alone or combined with gacyclidine did not prevent signs of soman toxicity when treatment was delayed 45 min after poisoning. Atropine/diazepam/pralidoxime also did not control seizures or prevent neuropathology in primates exhibiting severe signs of poisoning when treatment was commenced 30 min after intoxication. However, in this latter case, EEG recordings revealed that additional treatment with gacyclidine was able to stop soman-induced seizures and restore normal EEG activity. This drug also totally prevented the neuropathology observed 5 weeks after soman exposure in animals treated with atropine/diazepam/pralidoxime alone. Overall, in the case of severe OP-poisoning, gacyclidine represents a promising adjuvant therapy to the currently available polymedication to ensure optimal management of organophosphate poisoning in man. This drug is presently being evaluated in a human clinical trial for a different neuroprotective indication. However, it should always be kept in mind that, in the case of severe OP-poisoning, medical intervention must be conducted as early as possible.

Topics: Animals; Anticonvulsants; Antidotes; Atropine; Brain; Cerebellum; Cerebral Cortex; Chemical Warfare Agents; Cholinesterase Reactivators; Cyclohexanes; Cyclohexenes; Diazepam; Drug Therapy, Combination; Electrocardiography; Excitatory Amino Acid Antagonists; Haplorhini; Macaca fascicularis; Male; Piperidines; Poisoning; Pralidoxime Compounds; Seizures; Soman; Time Factors

A modified method for the determination of gacyclidine enantiomers in human plasma by GC-MS with selected-ion monitoring using the deuterated derivative of gacyclidine (d3-gacyclidine) as internal standard was developed. Following a single-step liquid-liquid extraction with hexane, drug enantiomers were separated on a chiral fused-silica capillary column (CP-Chirasil-Dex; Chrompack). The fragment ion, m/z 266, was selected for monitoring d3-gacyclidine (retention times of 35.2 and 35.6 min for the (+)- and (-)-enantiomer, respectively) whereas the fragment ion, m/z 263, was selected for quantitation of gacyclidine (retention times of 35.4 and 35.9 min for the (+)- and (-)-enantiomer, respectively). The limit of quantitation for each enantiomer was 0.3 ng/ml, using 1 ml of sample, with a relative standard deviation (RSD) < 14% and a signal-to-noise ratio of 5. The extraction recovery of both gacyclidine enantiomers from human plasma was about 75%. The calibration curves were linear (r2 > 0.996) over the working range of 0.312 to 20 ng/ml. Within- and between-day RSD were < 9% at 5, 10 and 20 ng/ml, and < 16% at 0.312, 0.625, 1.25 and 2.5 ng/ml. Intraday and interday bias were less than 11% for both enantiomers. The chromatographic behavior of d3-gacyclidine remained satisfactory even after more than 500 injections. Applicability of this specific and stereoselective assay is demonstrated for a clinical pharmacokinetic study with racemic gacyclidine.

Topics: Calibration; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Gas Chromatography-Mass Spectrometry; Humans; Piperidines; Receptors, N-Methyl-D-Aspartate; Reproducibility of Results; Sensitivity and Specificity

A sensitive gas chromatographic-mass spectrometric (GC-MS) procedure is described for the selective determination of gacyclidine (a non-competitive N-methyl-D-aspartate antagonist) in rat plasma and spinal cord dialyzates. It involves a single-step liquid-liquid extraction of plasma samples and dialyzates with hexane (pH 8.0) and the use of phencyclidine as an internal standard. The compounds were separated on a GC capillary column and specifically detected by MS in the selected-ion monitoring mode. Gacyclidine and its internal standard were monitored by using the fragment ions at m/z 206 and 200, respectively. The method was accurate and reproducible (intra- and inter-day reproducibility < 12%) with a limit of quantification of 1.6 ng ml-1 using 100 microliters plasma of dialyzate samples. The calibration curves for rat plasma and Ringer's solution were linear (r2 > 0.996) over a range from 1.6 to 200 ng ml-1. The extraction efficiency was close to 100%. This simple and rapid assay (total run time < 10 min) was validated for a pilot pharmacokinetic study in healthy rats after intravenous injection of a bolus dose of gacyclidine (2.5 mg kg-1).

Topics: Animals; Cyclohexanes; Cyclohexenes; Gas Chromatography-Mass Spectrometry; Male; Microdialysis; N-Methylaspartate; Piperidines; Rats; Rats, Wistar; Sensitivity and Specificity

Organophosphorus nerve agents are still in use today in warfare and as terrorism compounds. Classical emergency treatment of organophosphate poisoning includes the combined administration of a cholinesterase reactivator (an oxime), a muscarinic cholinergic receptor antagonist (atropine) and a benzodiazepine anticonvulsant (diazepam). However, recent experiments with primates have demonstrated that such treatment, even when administered immediately after organophosphate exposure, does not rapidly restore normal electroencephalographic (EEG) activity and fails to totally prevent neuronal brain damage. The objective of this study was to evaluate, in a realistic setting, the therapeutic benefit of administration of GK-11 (gacyclidine), an antiglutamatergic compound, as a complement to the available emergency therapy against organophosphate poisoning. GK-11 was injected at a dose of 0.1 mg/kg (i.v) after a 45-min latency period to heavily intoxicated (8 LD50) primates. Just after intoxication, man-equivalent doses of one autoinjector containing atropine/pralidoxime/diazepam were administered. The effects of GK-11 were examined on survival, EEG activity, signs of toxicity, recovery after challenge and central nervous system histology. The present data demonstrate that treatment with GK-11 prevents the mortality observed after early administration of classical emergency medication alone. EEG recordings and clinical observations also revealed that GK-11 prevented soman-induced seizures and motor convulsions. EEG analysis within the classical frequency bands (beta, theta, alpha, delta) demonstrated that central activity was totally restored to normal after GK-11 treatment, but remained profoundly altered in animals receiving atropine/pralidoxime/diazepam alone. GK-11 also markedly accelerated clinical recovery of soman-challenged primates. Lastly, this drug totally prevented the neuropathology observed 3 weeks after soman exposure in animals treated with classical emergency treatment alone. GK-11 represents a promising adjuvant therapy to the currently available emergency polymedication to ensure optimal management of organophosphate poisoning in man. This drug is presently being evaluated in a human clinical trial for a different neuroprotective indication.

Topics: Animals; Anticonvulsants; Atropine; Brain; Chemical Warfare Agents; Cholinesterase Inhibitors; Cholinesterase Reactivators; Cyclohexanes; Cyclohexenes; Diazepam; Drug Combinations; Excitatory Amino Acid Antagonists; Injections, Intravenous; Macaca fascicularis; Male; Muscarinic Antagonists; Neuroprotective Agents; Piperidines; Pralidoxime Compounds; Seizures; Soman; Staining and Labeling; Time Factors

Primary dissociated cultures of rhombencephalon were prepared from 5-9-week-old human fetuses. Half of some cultures were treated by two non-competitive N-methyl-D-aspartate antagonists, namely 1-(2-thienyl)cyclohexylpiperidine (TCP) and cis-Pip/Mel-[1-(2-thienyl)-2-methyl-cyclohexyl]piperidine (GK11) in negative enantiomeric form, which enhance the survival of human fetal central nervous system cells in culture. At different days in vitro, the treated and the control cultures were processed for immunocytochemical detection of serotonin-containing neurons which were studied by morphological and morphometric analysis. Statistical analysis showed that the surface of the stained neurons increased as a function of two parameters of time, the gestational age of the cells and the duration of the cultures. The complexity of the shape of the serotonin neurons characterized by the shape factor, the number of bifurcations and the morphological feature (bipolar or multipolar) was found to increase with the gestational age. It appears that the in vitro development of the embryonic cells which represents stages of maturation and differentiation can be specifically evaluated. Such an analysis of fetal central nervous system cells improves the knowledge of factors important in grafting experiments. We verified that the two drugs do not appreciably alter the in vitro development of the treated cells; thus they may be considered as promising drugs for human neuroprotection.

Topics: Cell Differentiation; Cell Size; Cells, Cultured; Cyclohexanes; Cyclohexenes; Excitatory Amino Acid Antagonists; Fetus; Humans; Illicit Drugs; Neurons; Neuroprotective Agents; Phencyclidine; Piperidines; Receptors, N-Methyl-D-Aspartate; Rhombencephalon; Serotonin

Recent experiments with primates have demonstrated that treatment with atropine/pralidoxime/diazepam, even if administered immediately after organophosphate exposure, does not totally prevent neuronal brain damage. Using primates, we have studied, for the first time, the ability of GK-11 (gacyclidine), an antiglutamatergic drug in the process of agreement for human use, given as an additional therapy, to counteract the neuropathology due to organophosphate exposure that persists after classical treatment with oxime/atropine/benzodiazepine. We have also examined the recovery of the organophosphate-intoxicated primates. Male Cynomolgus monkeys were pretreated 1 hour before poisoning with pyridostigmine, then intoxicated with 8 LD50 of soman and immediately treated with the combination pralidoxime/atropine/diazepam. Some of the animals also received GK-11 at 0.01; 0.03 or 0.1 mg/kg (i.v.) 10 minutes after soman challenge. Recovery of the primates (reflexes, movements, feeding) and the neuropathological changes that occurred three weeks after intoxication (histological examinations and neuronal cell density measurement) were compared in GK-11-treated and control animals. At all doses tested, GK-11 prevented the neuronal rarefaction of the frontoparietal cortex that was observed in soman-intoxicated animals that received only oxime/atropine/diazepam. Moreover, the 0.01 mg/kg dose of GK-11 improved the early recovery of intoxicated primates from 1 day after intoxication. In the view of the most effective management of organophosphate intoxication that is currently available, GK-11 thus appears to be a promising additional neuroprotective therapy. This drug is presently being evaluated in a human clinical trial for a different neuroprotective indication.

Topics: Animals; Cerebral Cortex; Cyclohexanes; Cyclohexenes; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Macaca; Male; Neuroprotective Agents; Organophosphorus Compounds; Piperidines

The non-competitive N-methyl-D-aspartate antagonist cis-Pip/Me 1-[1-(2-thienyl)-2-methyl-cyclohexyl] piperidine (GK11) either in racemic(+/-) or (-) enantiomeric form has been investigated on survival of human fetal spinal cord cells in culture. The treated cultured cells were processed for immunocytochemical detection of GABA at different time intervals ranging from 3 to 27 weeks. The number of GABA-stained cells was found to be greater in the treated cultures than in the control ones. The qualitative and quantitative morphological features of both control and treated cells were analysed by computer-assisted methods which allowed us to individualize three different populations: (1) all control and treated small neuritic field neurons, (2) young control and old treated large neuritic field neurons and (3) old control neurons with large neuritic fields.

Topics: Cells, Cultured; Cyclohexanes; Cyclohexenes; Cyclohexylamines; gamma-Aminobutyric Acid; Humans; Image Processing, Computer-Assisted; Immunohistochemistry; N-Methylaspartate; Neurites; Neuroglia; Neurons; Piperidines; Spinal Cord; Stereoisomerism

Primary cultures of embryonic rat cerebral cortex were treated after 17 days in vitro for 10 min with a single dose of the non-competitive antagonists of N-methyl-D-aspartate (NMDA) receptor MK 801, TCP, and GK 11. They were then maintained in vitro for 31, 59, or 73 days, and then processed for the immunocytochemical detection of neuron-specific enolase (NSE). Immunoreactive cells were counted in treated and control cultures, and it was found that, except at 31 days, treated cultures contained far more NSE immunoreactive cells than controls. Moreover, this effect was dose-dependent, since with both TCP and GK 11 neuron survival was significantly higher with, respectively, 20 microM and 5 microM than with the lowest concentration of 2.5 microM. We tentatively conclude that spontaneous neuron death occurring in primary cultures in vitro is at least partly related to the NMDA-associated Ca++ channel, since the common property of the molecules we used is to block this channel. The relevance of this mechanism of cell death in vitro to neuronal death in vivo is discussed.

Topics: Animals; Cell Survival; Cells, Cultured; Cerebral Cortex; Cyclohexanes; Cyclohexenes; Dizocilpine Maleate; Embryo, Mammalian; Female; N-Methylaspartate; Neurons; Phencyclidine; Piperidines; Pregnancy; Rats; Rats, Inbred Strains