[German S1 Guideline Long-/Post-COVID]. / S1-Leitlinie Long-/Post-COVID.

The German Society of Pneumology initiated 2021 the AWMF S1 guideline Long COVID/Post-COVID. In a broad interdisciplinary approach, this S1 guideline was designed based on the current state of knowledge.The clinical recommendations describe current Long COVID/Post-COVID symptoms, diagnostic approaches, and therapies.In addition to the general and consensus introduction, a subject-specific approach was taken to summarize the current state of knowledge.The guideline has an explicit practical claim and will be developed and adapted by the author team based on the current increase in knowledge.

The Traditional Chinese Medicine MLC901 inhibits inflammation processes after focal cerebral ischemia.

Inflammation is considered as a major contributor to brain injury following cerebral ischemia. The therapeutic potential of both MLC601/MLC901, which are herbal extract preparations derived from Chinese Medicine, has been reported both in advanced stroke clinical trials and also in animal and cellular models. The aim of this study was to investigate the effects of MLC901 on the different steps of post-ischemic inflammation in focal ischemia in mice. In vivo injury was induced by 60 minutes of middle cerebral artery occlusion (MCAO) followed by reperfusion. MLC901 was administered in post-treatment 90 min after the onset of ischemia and once a day during reperfusion. MLC901 treatment resulted in a reduction in infarct volume, a decrease of Blood Brain Barrier leakage and brain swelling, an improvement in neurological scores and a reduction of mortality rate at 24 hours after MCAO. These beneficial effects of MLC901 were accompanied by an inhibition of astrocytes and microglia/macrophage activation, a drastically decreased neutrophil invasion into the ischemic brain as well as by a negative regulation of pro-inflammatory mediator expression (cytokines, chemokines, matrix metalloproteinases). MLC901 significantly inhibited the expression of Prx6 as well as the transcriptional activity of NFκB and the activation of Toll-like receptor 4 (TLR4) signaling, an important pathway in the immune response in the ischemic brain. MLC901 effects on the neuroinflammation cascade induced by cerebral ischemia probably contribute, in a very significant way, in its potential therapeutic value.

Unravelling the dissociation pathways of acetic acid upon electron transfer in potassium collisions: experimental and theoretical studies.

Electron transfer in alkali-molecule collisions with gas phase acetic acid and its deuterated analogues resulting in OH- formation requires considerable internal rearrangement in the temporary negative ion. At a collision energy well above the threshold of negative ion formation, electron transfer from potassium to CH3COOH/CH3COOD and CD3COOH results not only in H transfer from CH3 to COOH/COOD, but also in H release from COOH and subsequent rearrangement to eliminate OH-. These processes are also investigated by theoretical post-Hartree-Fock and DFT calculations. The combination of both studies reveals that the most favourable intermediate mechanism occurs via diol formation. Such intramolecular H transfer is reported here for the first time in the context of electron transfer induced dissociation experiments in alkali-molecule collisions. A comprehensive fragmentation study is presented and dissociation mechanisms are suggested.

Predictors of hippocampal atrophy in critically ill patients.

BACKGROUND AND PURPOSE: Hippocampal atrophy is presumably one morphological sign of critical illness encephalopathy; however, predictors have not yet been determined. METHODS: The data for this report derived from patients treated at the intensive care units (ICUs) of the University Hospital in Bonn in the years 2004-2006. These patients underwent structural magnetic resonance imaging 6-24 months after discharge. Volumes (intracranial, whole brain, white matter, grey matter, cerebral spinal fluid, bilateral hippocampus) were compared with healthy controls. Pro-inflammatory parameters and ICU scoring systems were explored in conjunction with brain volumes. Cut-scores were defined to differentiate patients with high from those with low inflammatory response. RESULTS: Hippocampal and white matter volume were reduced in critically ill patients compared with healthy controls. Procalcitonin showed a very strong correlation (r = -0.903, P = 0.01) and interleukin-6 a moderate correlation (r = -0.538, P = 0.031) with hippocampal volume, but not with other brain volumes. C-reactive protein was linked to grey matter volume. There was no correlation with systemic inflammatory response syndrome criteria (body temperature, heart rate, respiratory rate, white blood cell count) or for hippocampal or whole brain volume. Furthermore, parameters representing severity of disease (APACHE II score, SOFA score, duration of stay and duration of mechanical ventilation) were not associated with hippocampal or other brain volumes. CONCLUSIONS: This analysis suggests that high levels of procalcitonin and interleukin-6 in the blood serum of critically ill patients are associated with a high likelihood of hippocampal atrophy irrespective of the severity of disease measured by ICU scoring systems and other inflammatory parameters.

Inhibition of cell migration and invasion mediated by the TAT-RasGAP317-326 peptide requires the DLC1 tumor suppressor.

TAT-RasGAP(317-326), a peptide corresponding to the 317-326 sequence of p120 RasGAP coupled with a cell-permeable TAT-derived peptide, sensitizes the death response of various tumor cells to several anticancer treatments. We now report that this peptide is also able to increase cell adherence, prevent cell migration and inhibit matrix invasion. This is accompanied by a marked modification of the actin cytoskeleton and focal adhesion redistribution. Interestingly, integrins and the small Rho GTP-binding protein, which are well-characterized proteins modulating actin fibers, adhesion and migration, do not appear to be required for the pro-adhesive properties of TAT-RasGAP(317-326). In contrast, deleted in liver cancer-1, a tumor suppressor protein, the expression of which is often deregulated in cancer cells, was found to be required for TAT-RasGAP(317-326) to promote cell adherence and inhibit migration. These results show that TAT-RasGAP(317-326), besides its ability to favor tumor cell death, hampers cell migration and invasion.

NeuroAiD: properties for neuroprotection and neurorepair.

BACKGROUND: Treatments for stroke and other brain injuries are limited. NeuroAiD has been shown to be beneficial in clinical studies. We reviewed the pharmacological effects of NeuroAiD on the normal and ischemic brain and neurons. METHODS: In vivo and in vitro experiments using mouse model of stroke (focal ischemia), rat model of cardiac arrest (global ischemia) and cortical neurons in culture were reviewed and summarized. RESULTS: NeuroAiD improved survival, attenuated infarct size, improved functional recovery in the model of focal ischemia, and protected neurons against glutamate-induced injury. Furthermore, it enhanced cognitive recovery by reducing hippocampal CA1 cell degeneration, DNA fragmentation, Bax expression and ma-londialdehyde release in the model of global ischemia. Activation of the Akt survival pathway and opening of KATP channels may contribute to the neuroprotective properties of NeuroAiD. NeuroAiD increased BDNF expression and induced proliferation of cells which differentiate and mature into neurons. It enhanced rosette formation of human embryonic stem cells. NeuroAiD-treated embryonic cortical neurons developed into neurons with longer neurites, denser outgrowths and networks, and more synaptic release sites. CONCLUSIONS: NeuroAiD demonstrated both neuroprotective and neuroregenerative properties in rodent models of focal and global ischemia and in cortical cell cultures. These properties would be important for developing a treatment strategy in reducing the long-term disability of stroke, cardiac arrest and other brain injuries.

Activation of ATP-sensitive potassium channels as an element of the neuroprotective effects of the Traditional Chinese Medicine MLC901 against oxygen glucose deprivation.

NeuroAid (MLC601 and MLC901), a Traditional Medicine used in China for patients after stroke has been reported in preclinical models of ischemia to induce neuroprotection and neuroplasticity. This work shows the effects of MLC901 on an in vitro model of oxygen glucose deprivation (OGD). MLC901 prevents neuronal death induced by 120 min OGD and decreases the exaggerated Ca²âº entry in mature cortical neurons exposed to 120 min OGD. The neuroprotective effect of MLC901 is associated with a large hyperpolarization of ∼20 mV which is antagonized by glibenclamide, the specific inhibitor of K(ATP) channels. In addition MLC901 strengthens the activation of K(ATP) channels. MLC901 has been directly shown to act as an activator of K(ATP) channels as potent as the classical K(ATP) channel opener. The capacity of MLC901 to produce a large hyperpolarization, particularly in neurons that have suffered from energy deprivation probably plays an important role in the neuroprotective effects of this traditional medicine that comes in addition to its previously demonstrated neuroregenerative properties.

Spadin as a new antidepressant: absence of TREK-1-related side effects.

Despite several decades of research, current antidepressant (AD) treatments remain of a limited efficacy justifying the need to find new drugs. These drugs have to be more efficacious, more rapid and display lesser side effects. Using rodent models, we recently identified spadin as a new antidepressant molecule that acts more quickly than classical ADs, working within 4 days to get same effects obtained with other ADs after 21 days. Spadin blocks TREK-1 K(2P) potassium channels that are considered as new targets for ADs. Deletion of the TREK-1 channel is known to increase sensitivity to pain, seizures and ischemia. Thus blocking these channels could result in deleterious side effects. In this study we showed that spadin did not interfere with other TREK-1 controlled functions such as pain, epilepsy and ischemia. We also demonstrated that spadin was unable to inhibit currents generated by TREK-2, TRAAK, TASK and TRESK four other K2P channels. More importantly, spadin did not induce cardiac dysfunctions, did not block I(Kr) and I(Ks) and did not modify the systolic pressure or cardiac pulses. After a three week treatment spadin remained an efficacious AD and did not modify the infarct size in brain following focal ischemia. Finally, we showed that kainate induced seizures and glycemia were not modified by spadin treatments. These data, together with those previously published reinforce the idea that spadin represents a good candidate for a new generation of ADs. This article is part of a Special Issue entitled 'Anxiety and Depression'.

MLC901, a traditional Chinese medicine protects the brain against global ischemia.

Global ischemia leads to damage in the hippocampal CA1 region and is associated with behavioral deficits. NeuroAid (MLC601 and MLC901), a Traditional Chinese Medicine is used in China for patients after stroke. We have investigated here the effects of MLC901 on brain injury and deficits after global ischemia in the rat. Global ischemia induced by four-vessel occlusion resulted in degeneration of CA1 neurons. MLC901 (0.074 mg/ml) prevented both necrosis and apoptosis of neurons up to 3 h after ischemia. These positive MLC901 effects were associated with a decrease in Bax expression and in levels of the lipid peroxidation product malondialdehyde. Using the PI3-kinase inhibitor LY294002 we also demonstrated the critical role of the Akt pathway in MLC901-mediated neuroprotection. MLC901 enhanced neurogenesis. Furthermore, MLC901 improved functional recovery of rats after global ischemia as assessed by the Morris water maze. In this test MLC901 reduced the increase in escape latency and in swim distance induced by ischemia. MLC901 also improved post-ischemic grip strength. If observations made with rats can be extended to humans, then MLC901 will represent a novel therapeutic strategy after cardiac arrest with a clinically interesting time window of protection.

Neuroprotective and neuroproliferative activities of NeuroAid (MLC601, MLC901), a Chinese medicine, in vitro and in vivo.

Although stroke remains a leading cause of death and adult disability, numerous recent failures in clinical stroke trials have led to some pessimism in the field. Interestingly, NeuroAid (MLC601), a traditional medicine, particularly used in China, South East Asia and Middle East has been reported to have beneficial effects in patients, particularly in post-stroke complications. Here, we demonstrate in a rodent model of focal ischemia that NeuroAid II (MLC901) pre- and post-treatments up to 3 h after stroke improve survival, protect the brain from the ischemic injury and drastically decrease functional deficits. MLC601 and MLC901 also prevent neuronal death in an in vitro model of excitotoxicity using primary cultures of cortical neurons exposed to glutamate. In addition, MLC601/MLC901 treatments were shown to induce neurogenesis in rodent and human cells, promote cell proliferation as well as neurite outgrowth and stimulate the development of a dense axonal and dendritic network. MLC601 and MLC901 clearly represent a very interesting strategy for stroke treatment at different stages of the disease.

Human high-density lipoprotein particles prevent activation of the JNK pathway induced by human oxidised low-density lipoprotein particles in pancreatic beta cells.

AIMS/HYPOTHESIS: We explored the potential adverse effects of pro-atherogenic oxidised LDL-cholesterol particles on beta cell function. MATERIALS AND METHODS: Isolated human and rat islets and different insulin-secreting cell lines were incubated with human oxidised LDL with or without HDL particles. The insulin level was monitored by ELISA, real-time PCR and a rat insulin promoter construct linked to luciferase gene reporter. Cell apoptosis was determined by scoring cells displaying pycnotic nuclei. RESULTS: Prolonged incubation with human oxidised LDL particles led to a reduction in preproinsulin expression levels, whereas the insulin level was preserved in the presence of native LDL-cholesterol. The loss of insulin production occurred at the transcriptional levels and was associated with an increase in activator protein-1 transcriptional activity. The rise in activator protein-1 activity resulted from activation of c-Jun N-terminal kinases (JNK, now known as mitogen-activated protein kinase 8 [MAPK8]) due to a subsequent decrease in islet-brain 1 (IB1; now known as MAPK8 interacting protein 1) levels. Consistent with the pro-apoptotic role of the JNK pathway, oxidised LDL also induced a twofold increase in the rate of beta cell apoptosis. Treatment of the cells with JNK inhibitor peptides or HDL countered the effects mediated by oxidised LDL. CONCLUSIONS/INTERPRETATION: These data provide strong evidence that oxidised LDL particles exert deleterious effects in the progression of beta cell failure in diabetes and that these effects can be countered by HDL particles.

TREK-1, a K+ channel involved in neuroprotection and general anesthesia.

TREK-1 is a two-pore-domain background potassium channel expressed throughout the central nervous system. It is opened by polyunsaturated fatty acids and lysophospholipids. It is inhibited by neurotransmitters that produce an increase in intracellular cAMP and by those that activate the Gq protein pathway. TREK-1 is also activated by volatile anesthetics and has been suggested to be an important target in the action of these drugs. Using mice with a disrupted TREK-1 gene, we now show that TREK-1 has an important role in neuroprotection against epilepsy and brain and spinal chord ischemia. Trek1-/- mice display an increased sensitivity to ischemia and epilepsy. Neuroprotection by polyunsaturated fatty acids, which is impressive in Trek1+/+ mice, disappears in Trek1-/- mice indicating a central role of TREK-1 in this process. Trek1-/- mice are also resistant to anesthesia by volatile anesthetics. TREK-1 emerges as a potential innovative target for developing new therapeutic agents for neurology and anesthesiology.

Polyunsaturated fatty acids induce ischemic and epileptic tolerance.

The findings reported in this work show that pretreatment with polyunsaturated fatty acids, particularly linolenic acid, present in vegetable oils, can provide a potent tolerance against neurodegeneration in two models of neuronal death-generating treatments such as kainic acid injection and global ischemia. Rats were injected i.v. with 500 nmol/kg of linolenic acid as long as 3 days prior to 6 min global ischemia or received an injection of linolenic acid as long as 3 days prior to a dose of 7.5 mg/kg kainic acid. Neuronal degeneration, assessed by analysis of neuronal density on Cresyl Violet-stained hippocampal sections, was significantly reduced in linolenic acid-treated rats (94-85% of cell survival in the ischemic model and 99-79% of cell survival in the epileptic model in respective CA1 and CA3 subfields). The neuroprotection observed following the injection of linolenic acid 3 days prior to induction of a severe ischemic or epileptic challenge was associated with the induction of the neuroprotective HSP70 heat shock protein within the time window of protection. The injection of 500 nmol/kg of linolenic acid induced a maximal HSP70 expression of 387% at 72 h. In contrast, the overexpression of one well-known protein inducer of neuronal cell death, Bax, which is induced by both ischemic and kainic acid-induced epileptic insults, was prevented by linolenic acid in the 3-day window of protection. These results strengthen the idea of an interesting potential therapeutical value of polyunsaturated fatty acids in neuronal protection.

Reovirus infection activates JNK and the JNK-dependent transcription factor c-Jun.

Viral infection often perturbs host cell signaling pathways including those involving mitogen-activated protein kinases (MAPKs). We now show that reovirus infection results in the selective activation of c-Jun N-terminal kinase (JNK). Reovirus-induced JNK activation is associated with an increase in the phosphorylation of the JNK-dependent transcription factor c-Jun. Reovirus serotype 3 prototype strains Abney (T3A) and Dearing (T3D) induce significantly more JNK activation and c-Jun phosphorylation than does the serotype 1 prototypic strain Lang (T1L). T3D and T3A also induce more apoptosis in infected cells than T1L, and there was a significant correlation between the ability of these viruses to phosphorylate c-Jun and induce apoptosis. However, reovirus-induced apoptosis, but not reovirus-induced c-Jun phosphorylation, is inhibited by blocking TRAIL/receptor binding, suggesting that apoptosis and c-Jun phosphorylation involve parallel rather than identical pathways. Strain-specific differences in JNK activation are determined by the reovirus S1 and M2 gene segments, which encode viral outer capsid proteins (sigma1 and mu1c) involved in receptor binding and host cell membrane penetration. These same gene segments also determine differences in the capacity of reovirus strains to induce apoptosis, and again a significant correlation between the capacity of T1L x T3D reassortant reoviruses to both activate JNK and phosphorylate c-Jun and to induce apoptosis was shown. The extracellular signal-related kinase (ERK) is also activated in a strain-specific manner following reovirus infection. Unlike JNK activation, ERK activation could not be mapped to specific reovirus gene segments, suggesting that ERK activation and JNK activation are triggered by different events during virus-host cell interaction.

Antiapoptotic signaling generated by caspase-induced cleavage of RasGAP.

Activation of caspases 3 and 9 is thought to commit a cell irreversibly to apoptosis. There are, however, several documented situations (e.g., during erythroblast differentiation) in which caspases are activated and caspase substrates are cleaved with no associated apoptotic response. Why the cleavage of caspase substrates leads to cell death in certain cases but not in others is unclear. One possibility is that some caspase substrates generate antiapoptotic signals when cleaved. Here we show that RasGAP is one such protein. Caspases cleave RasGAP into a C-terminal fragment (fragment C) and an N-terminal fragment (fragment N). Fragment C expressed alone induces apoptosis, but this effect could be totally blocked by fragment N. Fragment N could also block apoptosis induced by low levels of caspase 9. As caspase activity increases, fragment N is further cleaved into fragments N1 and N2. Apoptosis induced by high levels of caspase 9 or by cisplatin was strongly potentiated by fragment N1 or N2 but not by fragment N. The present study supports a model in which RasGAP functions as a sensor of caspase activity to determine whether or not a cell should survive. When caspases are mildly activated, the partial cleavage of RasGAP protects cells from apoptosis. When caspase activity reaches levels that allow completion of RasGAP cleavage, the resulting RasGAP fragments turn into potent proapoptotic molecules.

Activation of the nuclear factor-kappaB is a key event in brain tolerance.

The transcription factor nuclear factor-kappaB (NFkappaB) is an ubiquitously expressed inducible regulator of a broad range of genes and plays a pivotal role in cell death and survival pathways. Three models of brain tolerance (ischemic, epileptic, and polyunsaturated fatty acid-induced preconditioning), known to confer resistance to neurons against ischemia or status epilepticus, were used to determine whether NFkappaB mediated the late preconditioning. A sublethal 3 min ischemia, a dose of 5 mg/kg kainic acid (KA5) or 500 nmol of linolenic acid (LIN500) led to a rapid increase of NFkappaB DNA-binding activity and nuclear translocation of p65 and p50 subunits of NFkappaB in neurons. Pretreatment with the NFkappaB inhibitor diethyldithiocarbamate or kappaB decoy DNA blocked the increased DNA-binding activity and the nuclear translocation of NFkappaB and abolished the neuroprotective effects of different delayed preconditionings against severe ischemia or epilepsy. The inhibition of NFkappaB observed in rats preconditioned with 3 min ischemia, KA5 or LIN500 treatments compared with ischemic or epileptic controls was correlated with the prevention of the inducible degradation of the inhibitory protein IkappaBalpha. Preconditioning probably inhibits the activation of NFkappaB by interfering with a pathway that leads to the direct transcriptional activation of IkappaBalpha by NFkappaB itself. The present work provides evidence that activation of NFkappaB is a crucial step in the signal transduction pathway that underlies the development of brain tolerance and may open new strategies in the prevention of cerebral diseases, such as ischemia or epilepsy.

In vitro activity of MEKK2 and MEKK3 in detergents is a function of a valine to serine difference in the catalytic domain.

MEKK2 and MEKK3 are mitogen-activated protein kinase kinase kinases (MAP3 kinases) of 70 and 71 kDa respectively that are markedly homologous (94%) in their kinase domains. Both MEKK2 and MEKK3 are able to activate the Jun kinase pathway in vivo. However, following routine immunoprecipitation in Triton X-100, MEKK2 but not MEKK3 is able to effectively phosphorylate both SEK-1 and MEK-1 and to undergo autophosphorylation. Unexpectedly, both MEKK2 and MEKK3 are functional in an in vitro kinase assay when cells are solubilized with the closely related detergent, NP-40. Given the high homology between these kinases, we set out to relate this differential sensitivity to Triton X-100 to differences in primary structure. A set of chimeric molecules were generated and the loss of activity in Triton X-100 mapped to kinase domain II/III and specifically to serine 390 of MEKK3 and valine 384 of MEKK2, residues immediately N-terminal to the active site lysine. Mutation of serine 390 of MEKK3 to a valine (as is found in MEKK2) conferred catalytic activity to MEKK3 in Triton X-100 whereas the reciprocal alteration of valine 384 of MEKK2 to a serine conferred lack of activity in Triton X-100 to MEKK2. Search of the protein database identified only three kinases, MEKK3, Pbs2p and Dd-PKI, with a serine or threonine at this site. The presence of a serine or threonine adjacent to the active site lysine in protein kinases is rare and, in MEKK3, results in detergent instability.

The effects of FK506 on neurologic and histopathologic outcome after transient spinal cord ischemia induced by aortic cross-clamping in rats.

UNLABELLED: Spinal cord injury is a devastating complication of thoracoabdominal aortic surgery. We investigated the effect of the immunosuppressant FK506, a macrolide antibiotic demonstrated to have neuroprotective effects in cerebral ischemia models, in a rat model of transient spinal cord ischemia. Spinal cord ischemia was induced in anesthetized rats by using direct aortic arch plus left subclavian artery cross-clamping through a limited thoracotomy. Experimental groups were as follows: sham-operation; control, receiving only vehicle; FK506 A, receiving FK506 (1 mg/kg IV) before clamping; and FK506 B, receiving FK506 (1 mg/kg IV) at the onset of reperfusion. Neurologic status was assessed at 24 h and then daily up to 96 h with a 0 to 6 scale (0, normal function; 6, severe paraplegia). Rats were randomly killed at 24, 48, or 96 h, and spinal cords were harvested for histopathology. Physiologic variables did not differ significantly among experimental groups. All control rats suffered severe and definitive paraplegia. FK506-treated rats had significantly better neurologic outcome compared with control. Histopathologic analysis disclosed severe injury in the lumbar gray matter of all control rats, whereas most FK506-treated rats had less injury. These data suggest that FK506 can improve neurologic recovery and attenuate spinal cord injury induced by transient thoracic aortic cross-clamping. IMPLICATIONS: A single dose-injection of the immunosuppressant FK506 significantly improved neurologic outcome and attenuated spinal cord injury induced by transient thoracic aortic cross-clamping in the rat.

Reovirus-induced apoptosis is mediated by TRAIL.

Members of the tumor necrosis factor (TNF) receptor superfamily and their activating ligands transmit apoptotic signals in a variety of systems. We now show that the binding of TNF-related, apoptosis-inducing ligand (TRAIL) to its cellular receptors DR5 (TRAILR2) and DR4 (TRAILR1) mediates reovirus-induced apoptosis. Anti-TRAIL antibody and soluble TRAIL receptors block reovirus-induced apoptosis by preventing TRAIL-receptor binding. In addition, reovirus induces both TRAIL release and an increase in the expression of DR5 and DR4 in infected cells. Reovirus-induced apoptosis is also blocked following inhibition of the death receptor-associated, apoptosis-inducing molecules FADD (for FAS-associated death domain) and caspase 8. We propose that reovirus infection promotes apoptosis via the expression of DR5 and the release of TRAIL from infected cells. Virus-induced regulation of the TRAIL apoptotic pathway defines a novel mechanism for virus-induced apoptosis.

Ischemic spinal cord injury induced by aortic cross-clamping: prevention by riluzole.

OBJECTIVE: Recent studies confirmed the deleterious role of glutamate in the pathophysiology of spinal cord ischemia induced by aortic cross-clamping. We investigated the effect of riluzole, an anti-glutamate drug, in a rat model of spinal cord ischemia. MATERIALS AND METHODS: Spinal cord ischemia was induced in normothermia for 14 min in Sprague-Dawley rats using direct aortic arch plus left subclavian artery cross-clamping through a limited thoracotomy. Experimental groups were as follows: sham-operation (n=15), control (n=15) receiving only vehicle, riluzole (n=15) receiving riluzole (4 mg/kg) before clamping and at the onset of reperfusion. Separate animals were used for monitoring physiologic parameters in the sham-operation (n=3), control (n=5), and riluzole (n=5) groups. Neurologic status was assessed at 6, 24 h, and then daily up to 96 h. Rats were randomly killed at 24, 48, or 96 h (n=5 for each time). Spinal cords were harvested for histopathology, immunohistochemistry for microtubule-associated protein 2 (MAP-2), TUNEL staining, and analysis of DNA fragmentation by agarose gel electrophoresis. RESULTS: All sham-operated rats had a normal neurologic outcome, whereas all control rats suffered severe and definitive paraplegia. Riluzole-treated rats had significantly better neurologic function compared to the control. Histopathology disclosed severe neuronal necrosis in the lumbar gray matter of control rats, whereas riluzole-treated rats suffered usually mild to moderate injury. Riluzole particularly prevented motor neurons injury. MAP-2 immunoreactivity was completely lost in control rats, whereas it was preserved either completely or partly in riluzole-treated rats. TUNEL staining revealed numerous apoptotic neurons scattered within the whole gray matter of control rats. Riluzole prevented or dramatically attenuated apoptotic neuronal death in treated rats. DNA extracted from lumbar spinal cords of sham-operated and riluzole-treated rats exhibited no laddering, whereas spinal cords from control rats showed DNA laddering with fragmentation into approximately 180 multiples of base pairs. CONCLUSIONS: Riluzole may protect the spinal cord in a setting of severe ischemia by preventing neuronal necrosis and apoptosis. This drug may therefore be considered for clinical use during 'high risk' surgical procedures on the thoracoabdominal aorta.