Pathological changes induced in cerebrocortical neurons by phencyclidine and related drugs.

Phencyclidine (PCP), a dissociative anesthetic and widely abused psychotomimetic drug, and MK-801, a potent PCP receptor ligand, have neuroprotective properties stemming from their ability to antagonize the excitotoxic actions of endogenous excitatory amino acids such as glutamate and aspartate. There is growing interest in the potential application of these compounds in the treatment of neurological disorders. However, there is an apparent neurotoxic effect of PCP and related agents (MK-801, tiletamine, and ketamine), which has heretofore been overlooked: these drugs induce acute pathomorphological changes in specific populations of brain neurons when administered subcutaneously to adult rats in relatively low doses. These findings raise new questions regarding the safety of these agents in the clinical management of neurodegenerative diseases and reinforce concerns about the potential risks associated with illicit use of PCP.

L-cysteine, a bicarbonate-sensitive endogenous excitotoxin.

After systemic administration to immature rodents, L-cysteine destroys neurons in the cerebral cortex, hippocampus, thalamus, and striatum, but the underlying mechanism has never been clarified. This neurotoxicity of L-cysteine, in vitro or in vivo, has now been shown to be mediated primarily through the N-methyl-D-aspartate subtype of glutamate receptor (with quisqualate receptor participation at higher concentrations). In addition, the excitotoxic potency of L-cysteine was substantially increased in the presence of physiological concentrations of bicarbonate ion. L-Cysteine is naturally present in the human brain and in the environment, and is much more powerful than beta-N-methylamino-L-alanine, a bicarbonate-dependent excitotoxin, which has been implicated in an adult neurodegenerative disorder endemic to Guam. Thus, the potential involvement of this common sulfur-containing amino acid in neurodegenerative processes affecting the central nervous system warrants consideration.

NMDA antagonist neurotoxicity: mechanism and prevention.

Antagonists of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor, including phencyclidine (PCP) and ketamine, protect against brain damage in neurological disorders such as stroke. However, these agents have psychotomimetic properties in humans and morphologically damage neurons in the cerebral cortex of rats. It is now shown that the morphological damage can be prevented by certain anticholinergic drugs or by diazepam and barbiturates, which act at the gamma-aminobutyric acid (GABA) receptor-channel complex and are known to suppress the psychotomimetic symptoms caused by ketamine. Thus, it may be possible to prevent the unwanted side effects of NMDA antagonists, thereby enhancing their utility as neuroprotective drugs.

Ethanol-induced neuronal apoptosis in vivo requires BAX in the developing mouse brain.

A single episode of ethanol intoxication triggers widespread apoptotic neurodegeneration in the infant rat or mouse brain. The cell death process occurs over a 6-16 h period following ethanol administration, is accompanied by a robust display of caspase-3 enzyme activation, and meets ultrastructural criteria for apoptosis. Two apoptotic pathways (intrinsic and extrinsic) have been described, either of which may culminate in the activation of caspase-3. The intrinsic pathway is regulated by Bax and Bcl-XL and involves Bax-induced mitochondrial dysfunction and release of cytochrome c as antecedent events leading to caspase-3 activation. Activation of caspase-8 is a key event preceding caspase-3 activation in the extrinsic pathway. In the present study, following ethanol administration to infant mice, we found no change in activated caspase-8, which suggests that the extrinsic pathway is not involved in ethanol-induced apoptosis. We also found that ethanol triggers robust caspase-3 activation and apoptotic neurodegeneration in C57BL/6 wildtype mice, but induces neither phenomenon in homozygous Bax-deficient mice. Therefore, it appears that ethanol-induced neuroapoptosis is an intrinsic pathway-mediated phenomenon involving Bax-induced disruption of mitochondrial membranes and cytochrome c release as early events leading to caspase-3 activation.

Motor neuron degeneration induced by excitotoxin agonists has features in common with those seen in the SOD-1 transgenic mouse model of amyotrophic lateral sclerosis.

A superoxide dismutase 1 (SOD-1)genetic defect has been identified in familial amyotrophic lateral sclerosis (ALS) and motor neuron degeneration has been described in SOD-1 transgenic mice. Because an excitotoxic mechanism has been implicated in ALS, we undertook studies to provide a description of excitotoxic degeneration of spinal motor neurons for comparison with the degenerative process observed in SOD-1 transgenic mice. Excitotoxin agonists selective for each of the three major types of inotropic glutamate receptors were applied directly onto the lumbar spinal cord of 21-day old rats following posterior laminectomy. N-methyl-D-aspartate (NMDA) preferentially affected dorsal horn neurons, whereas the non-NMDA agonist, kainic acid, preferentially affected motor neurons. Cytopathological changes in motor neurons closely resembled those described in SOD-1 mice. These changes consist of massively swollen dendritic processes in the presence of well-preserved presynaptic axon terminals; cell bodies of motor neurons filled with vacuoles that originate both from endoplasmic reticulum and mitochondria; pleomorphic changes in mitochondria; axons of motor neuron becoming swollen proximally with accumulation of vacuoles, organelles, filaments, and degeneration products in the swollen segment. The observed changes in motor axons resemble changes described in the spinal cord of ALS patients. These findings are consistent with the proposal that motor neuron degeneration in ALS may be mediated by an excitotoxic process involving hyperactivation with non-NMDA glutamate receptors.

Age-specific neurotoxicity in the rat associated with NMDA receptor blockade: potential relevance to schizophrenia?

Agents that block the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor induce a schizophrenialike psychosis in adult humans and injure or kill neurons in several corticolimbic regions of the adult rat brain. Susceptibility to the psychotomimetic effects of the NMDA antagonist, ketamine is minimal or absent in children and becomes maximal in early adulthood. We examined the sensitivity of rats at various ages to the neurotoxic effects of the powerful NMDA antagonist, MK-801. Vulnerability was found to be age dependent, having onset at approximately puberty (45 days of age) and becoming maximal in early adulthood. This age-dependency profile (onset of susceptibility in late adolescence) in the rat is similar to that for ketamine-induced psychosis or schizophrenia in humans. These findings suggest that NMDA receptor hypofunction, the mechanism underlying the neurotoxic and psychotomimetic actions of NMDA antagonists, may also play a role in schizophrenia.

An ex vivo rat retinal preparation for excitotoxicity studies.

Although the isolated chicken embryo retina has been a very useful in vitro preparation for studying mechanisms of excitotoxicity, it is an avian rather than mammalian tissue and its embryonic age makes it unsuitable for a full range of developmental and aging studies. Therefore, we have explored the feasibility of using the rat retina at various ages for in vitro excitotoxicity studies. In this model, retinal segments were isolated in artificial cerebrospinal fluid (CSF) at 5 degrees C then incubated under various conditions at 30 degrees C and assessed histologically for signs of neurodegenerative changes. Retinal segments from 7-, 30-, 120- and 660-day-old rats incubated in CSF for 3 h and from 30-day-old rats incubated for 24 h retained a normal histological appearance. Thus, this preparation is suitable for in vitro studies pertaining to either acute or delayed excitotoxic phenomena in the mammalian CNS at any age from infancy to old age. Excitotoxin agonist experiments in the 30-day-old rat retina revealed the surprising result that the non-NMDA agonists, kainate and AMPA, at a low concentration (100 microM) damaged a much larger number of retinal neurons than NMDA did at a very high concentration (10 mM).

D-aminophosphonovalerate is 100-fold more powerful than D-alpha-aminoadipate in blocking N-methylaspartate neurotoxicity.

Here we report that the D-isomers of 2-amino-5-phosphonovalerate (D-APV) and alpha-amino-adipate (D-alpha AA) protect arcuate hypothalamic neurons from the potent excitotoxic activity of N-methylaspartate (NMA). Consistent with evidence that APV is much more powerful than alpha AA in antagonizing the neuroexcitatory activity of NMA, we found D-APV nearly 100 times more powerful than D-alpha AA in preventing NMA from destroying arcuate neurons.

MK-801 powerfully protects against N-methyl aspartate neurotoxicity.

Using the ex vivo chick embryo retina to study the efficacy of antagonists in blocking the excitotoxic effects of excitatory amino acid agonists, we previously identified phencyclidine as the most powerful known anti-excitotoxin. Here we show that MK-801 is 5 times more powerful than phencyclidine as an anti-excitotoxin, that its antagonism is specific for N-methyl-asparate toxicity, is non-competitive and does not entail inhibition of excitatory amino acid receptor binding.

The role of specific ions in glutamate neurotoxicity.

When the chick embryo retina is incubated in balanced salt solution containing glutamate (Glu) in 1 mM concentration, a neurodegenerative reaction occurs within 30 min. Here we report that the neurotoxic action of Glu on retinal neurons is dependent on the presence of Na+ and Cl-, but not Ca2+, in the incubation medium. Also, we report that depolarizing concentrations of K+ can induce a severe cytotoxic reaction in chick retina which, like the depolarization-linked neurotoxicity of Glu, is a Cl- dependent phenomenon.

Intrastriatal folic acid mimics the distant but not local brain damaging properties of kainic acid.

Folic acid (pteroyl-L-glutamine acid, PGA), when injected into the rat striatum, has the kainic acid (KA) property of inducing sustained seizures and a disseminated pattern of distant brain damage, but lacks the KA property of destroying neurons locally at the injection site. This suggests the interesting possibility that one component of KA neurotoxicity (seizure-related distant damage) may involve interaction with a folate system. Folates are promising tools for exploring the neurotoxic properties of KA and, more importantly, for studying mechanisms of epilepsy and epileptic brain damage.

The anti-excitotoxic effects of certain anesthetics, analgesics and sedative-hypnotics.

Various agents were tested for their ability to antagonize the acute excitotoxic action of N-methyl-DL-aspartate (NMA) and kainic acid (KA) on neurons in the in vitro chick embryo retina. The following compounds (in order of descending potencies) were effective in completely blocking the neurotoxic activity of NMA: phencyclidine, ketamine, (+/-)-SKF 10,047, pentazocine, D-aminophosphonovalerate, D-amino-phosphonoheptanoate, D-alpha-aminoadipate, OH-quinoxaline carboxylate, kynurenate, (+/-)-cis-2,3-piperidine dicarboxylate, secobarbital, amobarbital and pentobarbital. The latter 6 agents also protected against KA toxicity but complete protection was observed only from relatively high concentrations. At 20 mM, Mg2+ blocked NMA toxicity but at concentrations up to 30 mM did not block KA toxicity. Compounds that failed to block either NMA or KA toxicity include D- and L-aminophosphonobutyrate, L-glutamic acid diethyl ester, xanthurenate, GABA and taurine. The chick embryo retina is a useful preparation for identifying agents that have either excitotoxic or anti-excitotoxic activity.

Calcium influx accompanies but does not cause excitotoxin-induced neuronal necrosis in retina.

Several authors have recently proposed that excessive calcium (Ca++) influx into postsynaptic cells may be the mechanism by which excitotoxins such as glutamate (Glu), N-methylaspartate (NMA) and kainic acid (KA) cause neuronal necrosis. Here we have undertaken both in vivo and in vitro studies to explore this hypothesis. Our findings indicate that Ca++ does accumulate selectively in neural elements undergoing degeneration in the in vivo mouse hypothalamus following subcutaneous administration of NMA. However, pretreatment with the putative Ca++ channel blocker nimodipine resulted in augmentation rather than suppression of the toxic action of NMA and Glu on the mouse hypothalamus and eliminating Ca++ from the incubation medium did not interfere with the toxic action of Glu, NMA or KA on the chick embryo retina in vitro. We conclude, therefore, that Ca++ influx is an unlikely explanation for excitotoxin-induced degeneration of retinal or hypothalamic neurons.

L-homocysteic acid: an endogenous excitotoxic ligand of the NMDA receptor.

L-Homocysteic acid (L-HCA) has been proposed as a natural transmitter at the N-methyl-D-aspartate (NMDA) subtype of excitatory amino acid receptor based on recent evidence that L-HCA occurs L-HCA occurs naturally in the mammalian CNS, is released from K+ stimulated brain slices in a calcium-dependent manner and may be contained in nerve terminals located in certain brain regions that have a high density of NMDA receptors. Here we report that L-HCA potently induces a pattern of cytopathology in the ex vivo chick retina which mimics the pattern of NMDA but not kainic acid (KA) neurotoxicity. We also show that known NMDA antagonists, including Mg++, D-aminophosphonopentanoate and certain anesthetics, analgesics, and sedative hypnotics block the neurotoxic actions of L-HCA in direct proportion to their efficacy in blocking NMDA neurotoxicity. While there is a perfect correspondence between agents that block NMDA and L-HCA neurotoxicity, only a few such agents are active against KA neurotoxicity. We find that 3H-Glu binding is inhibited more potently by L-HCA (Ki = 67 microM). Moreover the patterns with which L-HCA and NMDA displace 3H-Glu binding in autoradiograms appear essentially identical. These findings are consistent with the proposal that L-HCA is an endogenous ligand at NMDA receptors.

Early defibrillation by EMTs: the Brussels experience.

Considering that in Brussels the first-aid ambulance team reaches the patient in cardiac arrest 10 min before the physician-manned ambulance, we instituted a feasibility study of early defibrillation by emergency medical technicians (EMTs). Three hundred EMTs received a 20-h automatic external defibrillation (AED) training course followed by a refresher course every 6 months. Of 316 cardiac arrests included in this study, asystole was encountered in 53% and ventricular fibrillation/ventricular tachycardia (VF/VT) in 33% of the cases on arrival of the EMTs. In the VF/VT group, defibrillation was performed by EMTs with a Laerdal Heartstart 7-9 min before the medical team arrived. The overall cardiac arrest survival rate improved from 7% in 1989 to 19% in 1992. However, the long-term survival rate (14/105) of ventricular fibrillation remained low because of excessive delays in emergency medical service (EMS) access and in early ACLS. In conclusion, this work shows that in Brussels: (1) early defibrillation of cardiac arrest victims in VF is feasible by EMTs when a training and a follow-up program are implemented; (2) the weakest link of the chain of survival is the early EMS access, and the early ACLS; and (3) AED program increases the interest and the efficacy of EMTs and medical teams in the management of cardiac arrests.

Semi-automatic external defibrillation.

Malignant arrhythmia, which is responsible for most of the out-of-hospital cardiac arrests, is ventricular fibrillation (VF). The best treatment of VF is a controlled electric shock on the chest administered in a short delay. The emergency medical technicians (EMTs) qualified to carry out this treatment in Belgium and in districts often succeed in arriving on the spot 8 minutes earlier than the people of the Service Mobile d'Urgence et de Réanimation (SMUR). The delegation of defibrillation to ambulance crew members however implies a specific teaching, training and a medical control. The Brussels experience shows that semi-automatic external defibrillation by EMT-Ds (SAED) is feasible when criteria for applying SAED in the pre-hospital phase are applicable.

Computed tomography of ununited anconeal process in the dog.

OBJECTIVE: The purpose of this study was to describe computed tomography (CT) features of the ununited anconeal process and relate them with the following elbow dysplasia signs: medial coronoid disease, medial humeral condyle changes, osteoarthritis (OA), and radioulnar incongruence. METHODS: Computed tomographic images of dogs older than six months with an ununited anconeal process were evaluated (n = 13). Ununited anconeal process features were described as being complete or incomplete, and the degree of displacement, volume, and presence of cysts and sclerosis were also evaluated. Medial coronoid disease was defined as an irregular medial coronoid process shape, presence of sclerosis and fragmentation. Medial humeral condyle changes were defined as subchondral bone flattening, lucencies, and sclerosis. Osteoarthritis was graded depending on the osteophytes size. Radioulnar incongruence was measured on a sagittal view at the base of the medial coronoid process. RESULTS: Eleven elbows had a complete and two had an incomplete ununited anconeal process. All ununited anconeal processes had cystic and sclerotic lesions. Seven ununited anconeal processes were displaced and six were non-displaced. Mean ununited anconeal process volume was 1.35 cm3 (0.61 cm³ - 2.08 cm³). Twelve elbows had signs of medial coronoid disease (4 of them with a fragmented medial coronoid process), and one elbow did not show any evidence of medial coronoid disease. Ten elbows had medial humeral condyle changes. One elbow had grade 1 OA, seven elbows had grade 2, and five elbows grade 3. All elbows had radioulnar incongruence: three elbows had a negative and 10 elbows had a positive radioulnar incongruence. Mean radioulnar incongruence was 1.49 mm (0.63 mm - 2.61 mm). Computed tomographic findings were similar in the majority of the elbows studied: complete ununited anconeal processes with signs of medial coronoid disease, positive radioulnar incongruence, high grade of OA, sclerotic medial humeral condyle changes, and large ununited anconeal process volumes. CLINICAL SIGNIFICANCE: Incomplete small ununited anconeal process volumes could be associated with a lower incidence of medial coronoid disease or medial humeral condyle changes. We recommend performing preoperative CT of elbows with an ununited anconeal process to evaluate concurrent lesions.

Contrast-enhanced ultrasonography in the differentiation of retinal detachment and vitreous membrane in dogs and cats.

OBJECTIVES: To evaluate contrast-enhanced ultrasonography (CEU) and colour Doppler imaging (CDI) for detection of persistent vascularisation in retinal detachment. METHODS: In 22 eyes, retinal detachment (n=13) and vitreous membranes (n=9) were confirmed by ophthalmological examination, during cataract surgery, by histopathology or after vitreoretinal surgery. Tentative diagnosis of retinal detachment or vitreous membrane was made using grey-scale B-mode ultrasonography. Assessment of retinal detachment was based on the presence or absence of vascularisation in the membranous structure using CDI and CEU. RESULTS: Sensitivity, specificity, positive-predictive value and negative-predictive value of grey-scale ultrasonography in differentiating retinal detachment from vitreous membrane were 92·3%, 66·6%, 80% and 85·7%. In 91% of eyes, colour Doppler assessment was unsuccessful due to the movement of the eye. Persistent vascularisation was demonstrated in all cases of retinal detachments with CEU. CEU was 100% accurate for detection and differentiation between retinal detachment and vitreous membrane. CLINICAL SIGNIFICANCE: CEU is a useful clinical tool for the diagnosis of retinal detachment and vitreous membrane in dogs and cats.