RESUMEN

Diisopropylfluorophosphate exerts its toxic effect by irreversibly inhibiting acetylcholinesterase. This results in over-stimulation of central and peripheral cholinergic activity. The aim of the present study was to evaluate the possible preventive effects of acute treatment with reversible acetylcholinesterase inhibitor galantamine against the signs of cholinergic toxic syndrome provoked by diisopropylfluorophosphate, such as hypothermia, muscular fasciculations, oral dyskinesia and decreased locomotor performance in a rat model of intoxication. The effects of these two anticholinesterases on acetylcholinesterase activity and on the expression of mRNA of the immediate early response gene c-fos in the brain were assessed by histochemical acetylcholinesterase staining and by in situ hybridization, respectively. Diisopropylfluorophosphate induced rapidly progressing hypothermia, muscular fasciculations, oral dyskinesia and decreased locomotor performance. The increased cholinergic cortical and hippocampal activity due to irreversible acetylcholinerase inhibition were indicated by the increased c-fos mRNA autoradiographic signal and by the inhibition of acetylcholinesterase staining, respectively. Galantamine by itself provoked transient and relatively weak inhibition of the acetylcholinesterase staining, while it did not induce increased c-fos mRNA expression or significant behavioural signs of cholinergic toxicity. Galantamine significantly reduced the rate of the onset, but not the maximal hypothermia induced by diisopropylfluorophosphate. Importantly, all the above-mentioned behavioural and neurochemical effects of diisopropylfluorophosphate were significantly reduced by galantamine. These results indicate that the acute pre-treatment with galantamine may have prophylactic effects against the intoxication by diisopropylfluorophosphate.

Asunto(s)

Encéfalo/efectos de los fármacos , Encéfalo/patología , Galantamina/farmacología , Isoflurofato/toxicidad , Fármacos Neuroprotectores/farmacología , Acetilcolinesterasa/metabolismo , Animales , Temperatura Corporal/efectos de los fármacos , Encéfalo/enzimología , Encéfalo/fisiopatología , Fasciculación/inducido químicamente , Fasciculación/genética , Fasciculación/patología , Fasciculación/fisiopatología , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Actividad Motora/efectos de los fármacos , Trastornos del Movimiento/genética , Trastornos del Movimiento/patología , Trastornos del Movimiento/fisiopatología , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Wistar

RESUMEN

Alternative splicing is an important mechanism for expanding proteome diversity from a limited number of genes, especially in higher vertebrates. Brain-specific splicing factors play an important role in establishing specific patterns of alternative splicing in the brain and thereby contribute to its complex architecture and function. Nova proteins are splicing factors that are expressed specifically in the central nervous system, where they regulate a large number of pre-mRNAs encoding synaptic proteins that are important for the balance of neuronal excitation and inhibition. Since this balance is interrupted in epileptic seizures, we explored whether LiCl/pilocarpine- or kainate-induced epileptic seizures would induce changes in the levels of Nova mRNAs in the rat brain. We found that the muscarinic agonist, pilocarpine, but not the glutamatergic agonist, kainate, induced a significant downregulation of Nova2 mRNA and upregulation of all three Nova1 mRNA isoforms in the striatum. Treatment with the muscarinic antagonist, scopolamine, at the onset of pilocarpine-induced seizures inhibited the seizures and the changes in Nova mRNA levels. Therefore it seems likely that pilocarpine stimulation of muscarinic acetylcholine receptors was a prerequisite for the observed changes, while the contribution of other striatal neurotransmitter systems activated by seizures could not be excluded. We propose that the LiCl/pilocarpine seizure model could serve as a valuable tool for studying mechanisms of Nova-regulated alternative splicing in rat striatum.

Asunto(s)

Antígenos de Neoplasias/biosíntesis , Cuerpo Estriado/metabolismo , Agonistas Muscarínicos/farmacología , Proteínas del Tejido Nervioso/biosíntesis , Pilocarpina/farmacología , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/biosíntesis , Convulsiones/metabolismo , Animales , Antígenos de Neoplasias/genética , Cuerpo Estriado/efectos de los fármacos , Agonistas de Aminoácidos Excitadores/farmacología , Ácido Kaínico/farmacología , Cloruro de Litio/farmacología , Masculino , Antagonistas Muscarínicos/farmacología , Proteínas del Tejido Nervioso/genética , Antígeno Ventral Neuro-Oncológico , Proteínas de Unión al ARN/genética , Ratas , Ratas Wistar , Escopolamina/farmacología

RESUMEN

Reversible acetylcholinesterase inhibitor donepezil displays prophylactic effects against intoxication with irreversible organophosphorous acetylcholinesterase inhibitors. We used behavioural observation of yawning and epileptic seizures, histochemical acetylcholinesterase staining, and in situ hybridization of the immediate early genes, c-fos and synaptotagmin 4 (Syt4) mRNAs in the brain, to evaluate whether donepezil could protect the brain against the effects of the organophosphate anticholinesterase, diisopropylfluorophosphate, in a rat model of intoxication. Diisopropylfluorophosphatetreated animals exhibited frequent yawning, significant inhibition of acetylcholinesterase staining and upregulation of c-fos mRNA, but not the epileptic seizures or significant change of Syt4 mRNA levels. In order to reduce the threshold for the induction of cholinergic seizures, additional groups of rats were pre-treated with LiCl 24 h before the treatment with diisopropylfluorophosphate. These rats exhibited the seizures, a significant inhibition of acetylcholinesterase staining and significant upregulation of c-fos and Syt4 mRNA levels. All the above-mentioned effects of diisopropylfluorophosphate were inhibited by donepezil pre-treatment. Donepezil pre-treatment by itself induced only a comparatively weaker inhibition of acetylcholinesterase staining and infrequent yawning. We conclude that donepezil protects the brain against diisopropylfluorophosphate-induced effects and that Syt4 mRNA upregulation may serve as a novel marker for organophosphate-induced seizures.

Asunto(s)

Anticonvulsivantes/farmacología , Inhibidores de la Colinesterasa/farmacología , Indanos/farmacología , Isoflurofato/farmacología , Piperidinas/farmacología , Convulsiones/prevención & control , Sinaptotagminas/genética , Acetilcolina/metabolismo , Acetilcolinesterasa/efectos de los fármacos , Acetilcolinesterasa/metabolismo , Animales , Encéfalo/patología , Donepezilo , Genes fos/efectos de los fármacos , Cloruro de Litio/farmacología , Masculino , Modelos Animales , ARN Mensajero/genética , Ratas , Ratas Wistar , Receptores Colinérgicos/efectos de los fármacos , Convulsiones/inducido químicamente , Convulsiones/patología , Regulación hacia Arriba , Bostezo/efectos de los fármacos

RESUMEN

Synaptotagmins (Syts) constitute a multi-gene family of 15 putative membrane trafficking proteins. The expression of some of the Syts in the brain might be dopaminergically controlled and thus affected by dopamine depletion in Parkinson's disease. We used hemiparkinsonian rats to investigate the effects of chronic striatal dopamine depletion and the acute effects of antiparkinsonic drug L-DOPA or D1 agonist (+/-)-6-chloro-7,8-dihydroxy-3-allyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrobromide (SKF82958) on the levels of striatal Syt I, II, IV, VI, VII, X, XI mRNA isoforms. On the 6-hydroxydopamine (6-OHDA)-lesioned side we observed a nearly total loss of tyrosine hydroxylase (TH), synaptotagmin I, Syt IV, Syt VII and Syt XI mRNA levels in the substantia nigra compacta (SNc). In dopamine-depleted striatum we also found a significant down-regulation Syt II and up-regulation of Syt X mRNA levels that could not be reversed by the acute treatment either with L-DOPA or SKF82958. By contrast, these two drugs induced an increase of Syt IV and Syt VII mRNA levels. A time-course study revealed the highest levels of Syt IV and VII mRNAs to occur at two hours and 12 hours after the treatment with SKF82958, respectively. D1 antagonist (+/-)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochloride (SCH23390) but not D2 antagonist haloperidol prevented the L-DOPA-driven increase of Syt IV and VII mRNAs. These results imply that synaptic plasticity in response to chronic striatal dopamine depletion involves a complex pattern of changes in striatal Syt mRNA expression. The L-DOPA treatment does not reverse the changes in Syt II and Syt X gene expression, but recruits additional, D1 receptor-mediated changes in Syt IV and Syt VII gene expression. Whether these D1 receptor-mediated changes play a role in the alterations of synaptic transmission that results in the unwanted side effects of chronic L-DOPA treatment in Parkinson's disease remains to be determined.

Asunto(s)

Cuerpo Estriado/metabolismo , Regulación de la Expresión Génica/fisiología , Expresión Génica/fisiología , Trastornos Parkinsonianos/metabolismo , Sinaptotagminas/metabolismo , Animales , Apomorfina/farmacología , Conducta Animal/efectos de los fármacos , Conducta Animal/fisiología , Benzazepinas/administración & dosificación , Benzazepinas/farmacología , Encéfalo/anatomía & histología , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Agonistas de Dopamina/farmacología , Antagonistas de Dopamina/administración & dosificación , Interacciones Farmacológicas , Femenino , Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Hibridación in Situ/métodos , Levodopa/administración & dosificación , Oxidopamina , Trastornos Parkinsonianos/inducido químicamente , Trastornos Parkinsonianos/tratamiento farmacológico , Isoformas de Proteínas/genética , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Rotación , Sustancia Negra/lesiones , Sustancia Negra/metabolismo , Sinaptotagminas/genética , Factores de Tiempo , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismo

RESUMEN

The effects of kainic acid (KA)-induced limbic seizures have been investigated on cytochrome c oxidase (COx) activity, COx subunit IV mRNA abundance, ATP and phosphocreatine (PCr) levels in amygdala, hippocampus and frontal cortex of rat brain. Rats were killed either 1 h, three days or seven days after the onset of status epilepticus (SE) by CO2 and decapitation for the assay of COx activity and by head-focused microwave for the determination of ATP and PCr. Within 1 h COx activity and COx subunit IV mRNA increased in all brain areas tested between 120% and 130% of control activity, followed by a significant reduction from control, in amygdala and hippocampus on day three and seven, respectively. In amygdala, ATP and PCr levels were reduced to 44% and 49% of control 1 h after seizures. No significant recovery was seen on day three or seven. Pretreatment of rats with the spin trapping agent N-tert-butyl-alpha-phenylnitrone (PBN, 200 mg kg(-1), i.p.) 30 min before KA administration had no effect on SE, but protected COx activity and attenuated changes in energy metabolites. Pretreatment for three days with the endogenous antioxidant vitamin E (Vit-E, 100 mg/kg, i.p.) had an even greater protective effect than PBN. Both pretreatment regimens attenuated KA-induced neurodegenerative changes, as assessed by histology and prevention of the decrease of COx subunit IV mRNA and COx activity in hippocampus and amygdala, otherwise seen following KA-treatment alone. These findings suggest a close relationship between SE-induced neuronal injury and deficits in energy metabolism due to mitochondrial dysfunction.

Asunto(s)

Encéfalo/enzimología , Complejo IV de Transporte de Electrones/metabolismo , Metabolismo Energético/fisiología , Degeneración Nerviosa/enzimología , Estrés Oxidativo/fisiología , Estado Epiléptico/enzimología , Adenosina Trifosfato/metabolismo , Amígdala del Cerebelo/efectos de los fármacos , Amígdala del Cerebelo/enzimología , Amígdala del Cerebelo/fisiopatología , Animales , Antioxidantes/farmacología , Encéfalo/efectos de los fármacos , Encéfalo/fisiopatología , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/enzimología , Corteza Cerebral/fisiopatología , Óxidos N-Cíclicos , Complejo IV de Transporte de Electrones/efectos de los fármacos , Complejo IV de Transporte de Electrones/genética , Metabolismo Energético/efectos de los fármacos , Agonistas de Aminoácidos Excitadores/farmacología , Hipocampo/efectos de los fármacos , Hipocampo/enzimología , Hipocampo/fisiopatología , Ácido Kaínico/farmacología , Degeneración Nerviosa/fisiopatología , Degeneración Nerviosa/prevención & control , Neurotoxinas/farmacología , Óxidos de Nitrógeno/farmacología , Estrés Oxidativo/efectos de los fármacos , Fosfocreatina/metabolismo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Especies Reactivas de Oxígeno/metabolismo , Estado Epiléptico/inducido químicamente , Estado Epiléptico/fisiopatología , Vitamina E/farmacología

RESUMEN

Reversible inhibitors of acetylcholinesterase improve spatial learning and memory in animal models of cognitive impairment. Here we investigate if the beneficial effects of free radical scavenger N-tert-butyl-alpha-phenylnitrone (PBN) on cognitive performance could be explained by its recently discovered anticholinesterase activity. Morris water maze experiment was performed to examine the effect of PBN on the impairment of spatial learning and memory induced by the antagonist of cholinergic muscarinic transmission scopolamine. In situ hybridization histochemistry experiment was performed to study its effects on the induction of immediate early gene expression (c-fos, c-jun) by dopamine D1 receptor agonist SKF-82958 and on the augmentation of the SKF-82958-induced expression of these genes by scopolamine. In both experiments, the effects of PBN were compared to the effects of reversible anticholinesterase physostigmine. We found that physostigmine but not PBN significantly reversed the cognitive impairment in scopolamine-challenged rats, prevented the induction of c-fos and c-jun mRNAs by SKF-82958 and attenuated the augmentation of the SKF-82958-induced expression of these genes by scopolamine. The present experiments did not reveal a significant in vivo anticholinesterase activity of PBN.

Asunto(s)

Inhibidores de la Colinesterasa/farmacología , Cognición/efectos de los fármacos , Depuradores de Radicales Libres/farmacología , Antagonistas Muscarínicos/farmacología , Óxidos de Nitrógeno/farmacología , Escopolamina/farmacología , Animales , Benzazepinas/farmacología , Óxidos N-Cíclicos , Agonistas de Dopamina/farmacología , Expresión Génica/efectos de los fármacos , Genes Inmediatos-Precoces/efectos de los fármacos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Memoria/efectos de los fármacos , Fisostigmina/farmacología , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-jun/genética , Ratas , Ratas Wistar , Percepción Espacial/efectos de los fármacos

RESUMEN

Impaired energy metabolism may play a critical role in the neuronal injury caused by kainic acid (KA) induced status epilepticus (SE). Following an acute dose of KA (15 mg/kg, s.c.) rats developed SE within 1 h. Rats were sacrificed 1 or 72 h after the onset of SE using a head focused microwave technique and the brain regions (pyriform cortex, amygdala, and hippocampus) were assayed for energy metabolites: ATP, ADP, AMP, phosphocreatine (PCr) and creatine (Cr) using reversed-phase HPLC (RP-HPLC). Control values were significantly higher in cortex (23-32%) than in other brain regions. Within 1 h, SE caused a marked decline in ATP (44-56%), PCr (49-64%), total adenine nucleotides (TAN, 45-50%) and total creatine compounds (TCC, 32-51%). Within three days, the hippocampus showed the greatest recovery, as the reduced values returned to normal. Pretreatment of rats with an antioxidant (PBN, 200 mg/kg, i.p., 30 min prior to KA; or vitamin E (Vit-E), 100 mg/kg, i.p./day for 3 days), which did not prevent seizure activity, attenuated depletion of high-energy phosphates caused by KA. These findings suggest that the depletion of energy metabolites caused by KA-induced seizures may be linked to oxidative stress mediated toxicity.

Asunto(s)

Antioxidantes/farmacología , Metabolismo Energético/efectos de los fármacos , Óxidos de Nitrógeno/farmacología , Estado Epiléptico/metabolismo , Vitamina E/farmacología , Animales , Encéfalo/metabolismo , Óxidos N-Cíclicos , Agonistas de Aminoácidos Excitadores/farmacología , Ácido Kaínico/farmacología , Masculino , Fosfatos/metabolismo , Ratas , Ratas Sprague-Dawley , Estado Epiléptico/inducido químicamente , Distribución Tisular

RESUMEN

N-tert-butyl-alpha-phenylnitrone (PBN), a widely used nitrone-based free radical trap was recently shown to prevent acetylcholinesterase (AChE) inhibitors induced muscle fasciculations and brain seizures while being ineffective against glutamergic or cholinergic receptor agonist induced seizures. In the present study we compared the effects on AChE activity of four free radical spin traps PBN, alpha-(4-pyridil-1)-N-tert-butyl nitrone (POBN), N-tert-butyl-alpha-(2-sulfophenyl)-nitrone (S-PBN) and 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO). The kinetics of AChE inhibition were studied in vitro using a spectrophotometric kinetic assay with AChE from rat brain, diaphragm, electric eel and mouse brain. Spin trapping compounds S-PBN and DEPMPO, in concentrations up to 3 mM did not inhibit hydrolysis of ACh, while PBN and POBN inhibited hydrolysis of ACh in a reversible and concentration-dependent manner. Double reciprocal plots of the reaction velocity against varying ACh concentrations at each inhibitor concentration were linear and generally indicated mixed type inhibition. PBN was the most potent inhibitor of mouse AChE with Ki and Ki' of 0.58 and 2.99 mM, respectively, and the weakest inhibitor of electric eel AChE. In contrast, POBN showed the highest affinity for electric eel enzyme, with Ki and Ki' values of 1.065 and 3.15 mM, respectively. These findings suggest that the effect of PBN and POBN on AChE activity does not depend on trapping of damaging reactive oxygen and that in addition to their antioxidant action other pharmacological effects of these compounds should be considered when neuroprotective actions of PBN or POBN are investigated.

Asunto(s)

Acetilcolinesterasa/metabolismo , Bencenosulfonatos/farmacología , Inhibidores de la Colinesterasa/farmacología , Óxidos N-Cíclicos/farmacología , Óxidos de Nitrógeno/farmacología , Marcadores de Spin , Animales , Encéfalo/enzimología , Diafragma/enzimología , Electrophorus , Depuradores de Radicales Libres/farmacología , Cinética , Ratones , Músculo Esquelético/enzimología , Piridinas , Ratas , Ratas Sprague-Dawley

RESUMEN

Indirect evidence suggests that reactive oxygen species (ROS) may mediate muscle fiber necrosis following muscle hyperactivity induced by the anticholinesterase diisopropylphosphorofluoridate (DFP). Pronounced muscle fasciculations and muscle fiber necrosis were seen when acetylcholinesterase (AChE) activity was reduced to less than 30% of control. The spin trapping agent phenyl-N-tert-butylnitrone (PBN) was used in vivo to directly assess the formation of ROS during DFP (1.75 mg/kg, s.c.) induced muscle hyperactivity. Pretreatment with PBN (300 mg/kg, i.p.), the concentration necessary for in vivo spin trapping, prevented muscle hyperactivity as well as necrosis and attenuated the DFP induced AChE inhibition otherwise seen in DFP only treated rats. PBN had no effect when given after fasciculations were established. Muscle extracts from PBN and DFP treated rats subjected to electron spin resonance (ESR) spectroscopy tested negative for ROS. While the role of PBN as an antioxidant is well established, its prophylactic effect against excitotoxity induced by an AChE inhibitor are due to its protection of AChE, an unexpected non-antioxidant action.

Asunto(s)

Acetilcolinesterasa/fisiología , Inhibidores de la Colinesterasa/toxicidad , Agonistas de Aminoácidos Excitadores/toxicidad , Fasciculación/prevención & control , Depuradores de Radicales Libres/farmacología , Isoflurofato/antagonistas & inhibidores , Contracción Muscular/efectos de los fármacos , Músculo Esquelético/efectos de los fármacos , Óxidos de Nitrógeno/farmacología , Marcadores de Spin , Animales , Atropina/farmacología , Óxidos N-Cíclicos , Fasciculación/inducido químicamente , Isoflurofato/toxicidad , Masculino , Antagonistas Muscarínicos/farmacología , Músculo Esquelético/patología , Necrosis , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno

RESUMEN

Synaptotagmins (Syts) I and IV are synaptic proteins involved in the regulation of neurosecretion. Dopaminergic drugs have been shown to modulate their expression. Here we investigate whether dopaminergic regulation of syt I and syt IV expression could play a role in the hypersensitive striatum of rats with unilateral lesions of dopaminergic nigrostriatal neurons with 6-hydroxydopamine. We show that chronic dopaminergic denervation resulted in a small down-regulation of striatal syt I mRNA, whereas acute treatment with SKF-82958, a dopamine D1 receptor agonist, induced a massive syt IV mRNA upregulation in the striatum on the lesioned side. We conclude that chronic lack of dopamine and treatment with dopamine D1 receptor agonists alter the synaptic plasticity in dopamine depleted basal ganglia.

Asunto(s)

Proteínas de Unión al Calcio , Dopamina/deficiencia , Glicoproteínas de Membrana/genética , Neostriado/metabolismo , Proteínas del Tejido Nervioso/genética , Trastornos Parkinsonianos/metabolismo , Animales , Benzazepinas/farmacología , Desnervación/efectos adversos , Agonistas de Dopamina/farmacología , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/fisiología , Masculino , Neostriado/efectos de los fármacos , Neostriado/fisiopatología , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Oxidopamina/efectos adversos , Trastornos Parkinsonianos/genética , Trastornos Parkinsonianos/fisiopatología , ARN Mensajero/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptores de Dopamina D1/efectos de los fármacos , Receptores de Dopamina D1/metabolismo , Sinaptotagmina I , Sinaptotagminas

RESUMEN

LEK-8829 [9,10-didehydro-N-methyl-(2-propynyl)-6-methyl-8- aminomethylergoline bimaleinate] is an antagonist of dopamine D2 receptors and serotonin (5-HT)2 and 5-HT1A receptors in intact animals and a D1 receptor agonist in dopamine-depleted animals. In the present study, we used rats with unilateral striatal lesions with ibotenic acid (IA) to investigate the dopamine receptor activities of LEK-8829 in a model with innervated dopamine receptors. The IA-lesioned rats circled ipsilaterally when challenged with apomorphine, the mixed agonist on D1/D2 receptors. LEK-8829 induced a dose-dependent contralateral turning that was blocked by D1 receptor antagonist SCH-23390. The treatment with D1 receptor agonist SKF-82958 induced ipsilateral turning, whereas the treatment with D2 receptor antagonist haloperidol induced contralateral posture. The combined treatment with SKF-82958 and haloperidol resulted in a weak contralateral turning, indicating the possible receptor mechanism of contralateral turning induced by LEK-8829. Bromocriptine induced a weak ipsilateral turning that was blocked by haloperidol. The ipsilateral turning induced by bromocriptine was significantly potentiated by the coadministration of a low dose but not by a high dose of LEK-8829. The potentiation of turning was blocked either by SCH-23390 or by haloperidol. The potentiation of ipsilateral turning suggests the costimulation of D2 and D1 receptors by bromocriptine and LEK-8829, respectively, whereas the lack of potentiation by the highest dose of LEK-8829 may be explained by the opposing activity of LEK-8829 and bromocriptine at D2 receptors. We propose that the D2 and 5HT2 receptor-blocking and D1 receptor-stimulating profile of LEK-8829 is promising for the treatment of negative symptoms of schizophrenia.

Asunto(s)

Antipsicóticos/farmacología , Bromocriptina/farmacología , Cuerpo Estriado/efectos de los fármacos , Ácido Lisérgico/análogos & derivados , Receptores Dopaminérgicos/efectos de los fármacos , Animales , Conducta Animal/efectos de los fármacos , Benzazepinas/farmacología , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Agonistas de Aminoácidos Excitadores , Haloperidol/farmacología , Ácido Iboténico , Ácido Lisérgico/antagonistas & inhibidores , Ácido Lisérgico/farmacología , Masculino , Ratas , Ratas Wistar , Rotación

RESUMEN

The neuroprotection afforded by spin trapping agents such as N-tert-butyl-alpha-phenylnitrone (PBN) has lent support to the hypothesis that increased production of reactive oxygen species (ROS) is a major contributing factor to excitotoxicity, aging and cognitive decline. Little is known, however, about the pharmacological properties of PBN. We have compared the acute effects of PBN on the development of seizures induced by the irreversible acetylcholinesterase (AChE) inhibitor diisopropylphosphorofluoridate (DFP), the reversible AChE inhibitor physostigmine (PHY), the muscarinic cholinergic receptor agonist pilocarpine (PIL) and the glutamatergic receptor agonist kainic acid (KA). Rats were sacrificed 90 min after the injection of seizure-inducing agents. In situ hybridization was used to detect the induction of immediate early gene (IEG) c-fos and c-jun mRNA's and the levels of AChE mRNA. The activity of AChE was visualized by AChE staining and quantified using an in vitro AChE assay. The seizures correlated with the induction of IEG mRNA's with all agents used. The pre-treatment with 150 mg/kg of PBN prevented DFP- and PHY-induced seizures and the related expression of IEG mRNA's, but had no effect on PIL- or KA-induced seizures and associated IEG mRNA's changes. PBN prevented seizures and significantly protected AChE activity against DFP inhibition when given before, but not when given after DFP. This study shows that PBN specifically protects against anticholinesterase-induced seizures by reversible protection of AChE activity and not by the blockade of muscarinic or glutamate receptors, reactivation of AChE or scavenging of ROS. The anticholinesterase properties should be considered when using PBN in studies of cholinergic dysfunction.

Asunto(s)

Anticonvulsivantes/farmacología , Inhibidores de la Colinesterasa , Fármacos Neuroprotectores/farmacología , Óxidos de Nitrógeno/farmacología , Convulsiones/prevención & control , Detección de Spin , Acetilcolinesterasa/metabolismo , Animales , Conducta Animal/efectos de los fármacos , Encéfalo/enzimología , Encéfalo/patología , Óxidos N-Cíclicos , Agonistas de Aminoácidos Excitadores , Genes fos/efectos de los fármacos , Genes jun/efectos de los fármacos , Hibridación in Situ , Isoflurofato , Ácido Kaínico , Masculino , Agonistas Muscarínicos , Fisostigmina , Pilocarpina , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ratas , Ratas Sprague-Dawley , Convulsiones/inducido químicamente , Convulsiones/patología , Marcadores de Spin

RESUMEN

The ergoline derivative, LEK-8829 (9,10-didehydro-N-methyl-(2-propynyl)-6-methyl-8-aminomethylerg oline), has been proposed as a potential atypical antipsychotic drug with antagonistic actions at dopamine D2 and serotonin 5-HT2 and 5-HT1A receptors (Krisch et al., 1994, 1996). LEK-8829 also induces contralateral turning in rats with 6-hydroxydopamine-induced unilateral lesion of dopamine nigrostriatal neurons. Turning is blocked by SCH-23390 (R(+)-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzaze pine), a dopamine D1 receptor antagonist. It has been suggested that LEK-8829 could have beneficial effects in parkinsonian patients suffering from psychotic episodes induced as a side-effect of antiparkinsonian treatment with dopamine D2 receptor agonists. Therefore, we now investigated the interaction of LEK-8829 with the dopamine D2 receptor agonist bromocriptine (2-bromo-alpha-ergokryptine) in 6-hydroxydopamine-lesioned rats. Treatment with either LEK-8829 (3 mg kg(-1)) or bromocriptine (3 mg kg(-1)) induced a vigorous contralateral turning response. The cumulated number of turns induced by the treatment with both drugs combined was not significantly different from the cumulated number of turns induced by single-drug treatment. The pretreatment with SCH-23390 (1 mg kg(-1)) did not have a significant effect on the bromocriptine-induced turning but significantly decreased the turning observed after the combined LEK-8829/bromocriptine treatment. We conclude that in the 6-hydroxydopamine model, the turning behaviour mediated by the LEK-8829/bromocriptine combination may be the result of opposing activity of both drugs at dopamine D2 receptors with concomitant stimulation of dopamine D1 receptors by LEK-8829. Therefore, LEK-8829 may have a potential for the therapy of parkinsonism complicated by dopamine D2 receptor agonist drug-induced psychosis.

Asunto(s)

Antiparkinsonianos/farmacología , Antagonistas de Dopamina/farmacología , Ácido Lisérgico/análogos & derivados , Conducta Estereotipada/efectos de los fármacos , Animales , Benzazepinas/farmacología , Bromocriptina/farmacología , Haloperidol/farmacología , Ácido Lisérgico/farmacología , Masculino , Actividad Motora/efectos de los fármacos , Ratas , Ratas Wistar

RESUMEN

1. Previous experiments have suggested a potential atypical antipsychotic activity of the ergoline derivative LEK-8829. In vitro experiments showed a high affinity to 5-HT1A, 5-HT2 and D2 receptors (the ratio of pKi values 5-HT2/D2 = 1.11) and a moderate affinity to D1 receptors. In vivo experiments showed antagonism of dopamine and 5-hydroxytryptamine (5-HT) receptor-linked behaviours. 2. In the present study, the rats with unilateral dopaminergic deafferentation of the striatum, induced by the lesion of the median forebrain bundle with 6-hydroxydopamine (6-OHDA), were used to determine the effects of LEK-8829 on turning behaviour and on striatal c-fos mRNA levels. 3. The administration of LEK-8829 induced a long lasting contralateral turning behaviour that was dose-dependent. It was found that the specific D1 receptor antagonist SCH-23390 but not the D2 receptor antagonist haloperidol or 5-HT1A antagonist pindolol, dose-dependently inhibited the turning behaviour induced by LEK-8829. 4. In an attempt to clarify the D1:D2 receptor interactions involved in the action of LEK-8829 in the 6OHDA model, we used in situ hybridization histochemistry to compare the effect of SCH-23390 pretreatment on striatal c-fos mRNA expression induced either by LEK-8829 or by the typical antipsychotic haloperidol. 5. LEK-8829 induced a bilateral striatal c-fos mRNA expression that was significantly higher in the denervated striatum as compared to the intact striatum and was completely blocked on both sides by pretreatment with SCH-23390. In contrast, haloperidol-induced striatal c-fos mRNA expression was limited to the innervated striatum and was not blocked by SCH-23390. 6. Our data demonstrate an intrinsic activity of LEK-8829 on D1 receptors that is potentiated in the dopamine-depleted striatum. We conclude, therefore, that the putative atypical antipsychotic LEK-8829 may prove useful as an experimental tool for the study of D1:D2 receptor interactions and could have beneficial effects in the treatment of drug-induced psychosis in patients with Parkinson's disease.

Asunto(s)

Antipsicóticos/farmacología , Ácido Lisérgico/análogos & derivados , Receptores de Dopamina D1/efectos de los fármacos , Animales , Apomorfina/farmacología , Conducta Animal/efectos de los fármacos , Benzazepinas/farmacología , Relación Dosis-Respuesta a Droga , Genes fos/efectos de los fármacos , Haloperidol/farmacología , Ácido Lisérgico/farmacología , Masculino , Oxidopamina , Pindolol/farmacología , Ratas , Ratas Wistar , Receptores de Dopamina D1/fisiología

RESUMEN

In this preliminary report nonradioactive digoxigenine-based and radioactive in situ hybridization procedures for the localization of acetylcholinesterase mRNA were tested and compared in rat brain. General patterns of Ache mRNA localization observed by both techniques did not differ significantly and were practically the same as reported in previous in situ studies on the mammalian brain. Shorter procedure time and avoidance of precautions necessary at work with radioactive materials are major advantages of nonradioactive technique. Under- and over- staining can be prevented by direct examination of coloring reaction. Faint staining in the control experiment with heterologous DNA suggests that proper stringency is essential for the specificity of staining.

Asunto(s)

Acetilcolinesterasa/biosíntesis , Sistema Nervioso Central/enzimología , ARN Mensajero/biosíntesis , Animales , Química Encefálica/fisiología , Hibridación in Situ , Sondas de Oligonucleótidos , Ratas , Radioisótopos de Azufre

RESUMEN

In situ hybridization histochemistry was used to localize tyrosine hydroxylase (TH) mRNA and cholecystokinin (CCK) mRNA-expressing cells in the ventral mesencephalon of the common marmoset (Callithrix jacchus) and to examine the effects of the dopaminergic (DA) neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on these two populations of neurons in the pars compacta of the substantia nigra (SNc) and ventral tegmental area (VTA). X-ray film and liquid emulsion autoradiography of brain sections hybridized with an 35S-labelled synthetic 45-mer antisense human TH oligonucleotide probe showed strong hybridization signals and dense populations of TH mRNA expressing cells in the SNc and VTA at all levels, in the control marmoset brain. In the MPTP-treated brain, there was a substantial reduction of TH mRNA in the ventral midbrain. The loss of TH mRNA-expressing cells amounted to 98% in the lateral SNc, 88% in the medial SNc and 33% in the VTA. In situ hybridization of adjacent sections with an 35S-labelled synthetic 45-mer antisense human CCK oligonucleotide probe showed a weak hybridization signal for CCK mRNA in the ventral midbrain of the control brain. Emulsion autoradiography demonstrated CCK mRNA expressing cells in the SNc and VTA at all levels with the number of cells in the VTA similar to that for TH mRNA. However, the number of cells in the SNc expressing CCK mRNA was a fraction (1/4) of that expressing TH mRNA; moreover, the level of expression per cell was substantially less than that for TH mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/farmacología , Colecistoquinina/genética , Mesencéfalo/fisiología , Neuronas/fisiología , Tirosina 3-Monooxigenasa/genética , Animales , Autorradiografía , Callithrix , Femenino , Mesencéfalo/efectos de los fármacos , Neuronas/efectos de los fármacos , Sondas de Oligonucleótidos , ARN Mensajero/análisis , ARN Mensajero/metabolismo , Valores de Referencia , Radioisótopos de Azufre

RESUMEN

Three days after systemic administration of kainic acid (15 mg/kg, s.c.), selected cholinergic markers (choline acetyltransferase, acetylcholinesterase, muscarinic acetylcholine receptor, and high-affinity choline uptake) and GABAergic parameters [benzodiazepine and gamma-aminobutyric acid (GABA) receptors] were studied in the frontal and piriform cortex, dorsal hippocampus, amygdaloid complex, and nucleus basalis. Kainic acid treatment resulted in a significant reduction of choline acetyltransferase activity in the piriform cortex (by 20%), amygdala (by 19%), and nucleus basalis (by 31%) in comparison with vehicle-injected control rats. A lower activity of acetylcholinesterase was also determined in the piriform cortex following parenteral kainic acid administration. [3H]Quinuclidinyl benzilate binding to muscarinic acetylcholine receptors was significantly decreased in the piriform cortex (by 33%), amygdala (by 39%), and nucleus basalis (by 33%) in the group treated with kainic acid, whereas such binding in the hippocampus and frontal cortex was not affected by kainic acid. Sodium-dependent high-affinity choline uptake into cholinergic nerve terminals was decreased in the piriform cortex (by 25%) and amygdala (by 24%) after kainic acid treatment. In contrast, [3H]flunitrazepam binding to benzodiazepine receptors and [3H]muscimol binding to GABA receptors were not affected 3 days after parenteral kainic acid application in any of the brain regions studied. The data indicate that kainic acid-induced limbic seizures result in a loss of cholinergic cells in the nucleus basalis that is paralleled by degeneration of cholinergic fibers and cholinoceptive structures in the piriform cortex and amygdala, a finding emphasizing the important role of cholinergic mechanisms in generating and/or maintaining seizure activity.

Asunto(s)

Ganglios Basales/metabolismo , Ácido Kaínico/farmacología , Sistema Límbico/metabolismo , Sistema Nervioso Parasimpático/metabolismo , Sustancia Innominada/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Amígdala del Cerebelo/metabolismo , Animales , Conducta Animal/efectos de los fármacos , Biomarcadores/metabolismo , Corteza Cerebral/metabolismo , Masculino , Vías Olfatorias/metabolismo , Ratas , Receptores de GABA-A/metabolismo , Convulsiones/inducido químicamente