Time Course Of Neuronal Death Research Articles

Time course of neuronal death following endothelin-1 induced focal ischemia in rats.

Endothelin-1 (ET-1) induced focal ischemia is increasingly being used as a preclinical model of stroke. Here, we described for the first time, the time course of neuronal death and infarct evolution during the first 7 days following ischemia. We used hematoxylin and eosin (H&E) staining to evaluate infarct progression and Fluoro-Jade C (FJC) to quantify neuronal degeneration at 24, 48, 72h and 7 days after ET-1 injection to the forelimb motor cortex in Sprague-Dawley rats. We found that infarct volume and neuronal degeneration are maximal at 24h post-stroke. Neuronal degeneration is also significantly reduced within 7 days of stroke induction. This study is the first to provide a direct evaluation of both infarct volume evolution and neuronal death time course following ET-1 induced focal ischemia in the forelimb motor cortex. This study describes the short-term time course of neuronal death and brain injury in the ET-1 stroke model, which provides a significant reference when determining the appropriate time to commence neuroprotective or recovery promoting strategies.

Brain Protection

A growing community of researchers has questioned the merits of the translational process in stroke and has championed the need for proof-of-concept studies to explain why >100 clinical stroke studies have failed.1,2 On the positive side, no published laboratory results or strong theoretical arguments have been advanced to indicate that human neurons, glia, and blood vessels are necessarily resistant to protection with drugs or devices. In fact, proof of principle has been established for many agents in cultured cells as well as in rodent and subhuman primate stroke models. Therefore, it is important to understand the reasons why an apparent disconnect exists between preclinical and clinical trial results. The answers may lie partly in shortcomings and breaches in the translational process that repeatedly compromise adequate drug testing in both preclinical and clinical arenas. On the positive side, it may be argued that a formula for human brain protection has been established, that is, by delivering in a timely manner oxygen as well as glucose and by removing wastes from ischemic tissue. That formula has met with success for reperfusion strategies in vivo and for maintaining cultured cells and brain slices in vitro. Some success has also been achieved using hypothermia to treat victims of cardiac arrest and global ischemia. The pleiotropic actions of hypothermia have taught us that protection may be better achieved by targeting multiple rather than individual death pathways as well as by reducing energy demand and energy consumption. Some of the same lessons may also apply to the promising and emerging field of preconditioning whose tissue-sparing effects appear to rely on upregulation of multiple endogenous protective mechanisms. Lessons from the transplant field are also instructive as surgeons have become increasingly sophisticated about maintaining organ viability ex vivo for many hours in the absence of a blood …

Neuronal death after peripheral nerve injury and experimental strategies for neuroprotection

Objective: Despite considerable microsurgical innovation in peripheral nerve repair, the outcome has improved little since the 1940s, reflecting surgical inability to adequately address the complex neurobiology of nerve injury and regeneration. Axotomy-induced neuronal death is potentially the most fundamental problem, and given recently published data, a review is timely.Methods: Initial review of relevant doctoral theses from the University of Umeå, and Blond-McIndoe Research Laboratories, the University of Manchester, plus initial PubMed search including terms 'neuron death' and 'neuroprotection', subsequently expanded to relevant quoted articles.Results: Various factors related to patient (principally age) and injury (Sunderland grade, proximity to cell body and mechanism) determine the extent of neuronal death, the mechanism of which is reviewed. A considerable proportion of sensory neurons (particularly small cutaneous afferents) die after distal injury and death is more widespread after proximal injury. Motor neurons are susceptible to post-ganglionic plexus and spinal root level injury. Root avulsion causes the greatest cell death. The time course of neuronal death is fortuitously slow and mainly occurs by a process akin to apoptosis. A therapeutic window therefore exists, as do potential neuroprotective targets. Nerve repair is partly neuroprotective, but must be performed early. Exogenous neurotrophic factor administration (e.g. in tissue engineered conduits) is beneficial, but not practical for various reasons. In contrast, adjuvant neuroprotective pharmacotherapy is practical, and two clinically safe agents are reviewed. Acetyl-L-carnitine arrests sensory neuronal death and speeds up regeneration. N-acetyl-cysteine provides comparable sensory neuronal protection via mitochondrial preservation and protects motor neurons. Both agents are well characterized experimentally and highly effective even after clinically relevant delays between injury and treatment. Barriers to translational research are being addressed.Discussion: The future of peripheral nerve repair lies in modulating neurobiology at the time of injury, repair and during regeneration. Neuroprotection may be an essential component of that therapeutic package.

Time-course of neuronal death in the mouse pilocarpine model of chronic epilepsy using Fluoro-Jade C staining

Epilepsy is a serious neurological disorder in human beings and the long-term pathological events remain largely obscure. We are interested in elucidating long-term brain injury that may occur in the temporal lobe epilepsy, and time-course of neuronal death was examined in a mouse pilocarpine model of chronic epilepsy by Fluoro-Jade C (FJC) dye that can specifically stain the degenerative neurons in the central nervous system. The FJC stain combined with immunohistochemistry to neuronal nuclear specific protein revealed that pilocarpine-induced status epilepticus (SE) resulted in massive degenerative death of neuronal cells in brains with their dense distribution in the cerebral cortex and hippocampus. The FJC-positive degenerating neurons, most of them also expressed apoptosis signaling molecules such as caspase-9 and activated caspase-3, occurred at 4h, increased into peak levels at 12h–3d, and then gradually went down at 7d–14d after onset of SE. More interestingly, a large percentage (about 88%) of FJC-positive degenerative neurons were GABAergic as indicated with their immunoreactivity to glutamic acid decarboxylase-67, implying that inhibitory function of GABAergic neural system might by seriously damaged in brains subject to SE attack in this mouse pilocarpine model. Taken together with previous studies, time-course of degenerative neurons in the mouse pilocarpine model by Fluoro-Jade C staining further benefits understanding of long-term brain pathological changes and recurrent seizure mechanism, and may also result in finding the most suitable time-window in therapeutic manipulation of the chronic epilepsy in human beings.

Time course and mechanism of hippocampal neuronal death in an in vitro model of status epilepticus: Role of NMDA receptor activation and NMDA dependent calcium entry

The hippocampus is especially vulnerable to seizure-induced damage and excitotoxic neuronal injury. This study examined the time course of neuronal death in relationship to seizure duration and the pharmacological mechanisms underlying seizure-induced cell death using low magnesium (Mg 2+) induced continuous high frequency epileptiform discharges (in vitro status epilepticus) in hippocampal neuronal cultures. Neuronal death was assessed using cell morphology and fluorescein diacetate–propidium iodide staining. Effects of low Mg 2+ and various receptor antagonists on spike frequency were assessed using patch clamp electrophysiology. We observed a linear and time-dependent increase in neuronal death with increasing durations of status epilepticus. This cell death was dependent upon extracellular calcium (Ca 2+) that entered primarily through the N-methyl- d-aspartate (NMDA) glutamate receptor channel subtype. Neuronal death was significantly decreased by co-incubation with the NMDA receptor antagonists and was also inhibited by reduction of extracellular (Ca 2+) during status epilepticus. In contrast, neuronal death from in vitro status epilepticus was not significantly prevented by inhibition of other glutamate receptor subtypes or voltage-gated Ca 2+ channels. Interestingly this NMDA-Ca 2+ dependent neuronal death was much more gradual in onset compared to cell death from excitotoxic glutamate exposure. The results provide evidence that in vitro status epilepticus results in increased activation of the NMDA-Ca 2+ transduction pathway leading to neuronal death in a time-dependent fashion. The results also indicate that there is a significant window of opportunity during the initial time of continuous seizure activity to be able to intervene, protect neurons and decrease the high morbidity and mortality associated with status epilepticus.

Increased synthesis of specific proteins during glutamate-induced neuronal death in cerebellar culture

We have previously shown that glutamate-induced neurotoxicity is mediated by a sodium-chloride component and a calcium component in our cerebellar granule cell culture. In order to further characterize these two different components, the time course of neuronal death induced by glutamate (100 μM) in basal solution and in low sodium-chloride solution was studied by morphological and biochemical criteria. As shown by phase-contrast microscopy, cerebellar granule cells exhibited clear neuronal degeneration within 4 h after exposure to this excitotoxin. These morphological changes correlated [ 35S]methionine incorporation into proteins which rapidly declined during the first hour of treatment. Qualitative change in [ 35S]methionine incorporation into proteins was further investigated by two-dimensional gel electrophoresis performed after glutamate exposure in basal solution and in low sodium-chloride solution. Most of the proteins showed a decreased labelling after glutamate exposure as expected, but some polypeptides showed an increased labelling or appeared to be newly synthetized. Furthermore, a different pattern of protein synthesis was observed when glutamate exposure was performed in basal solution or in low sodium-chloride solution. The identification of these polypeptides and their implication in the neuronal death are discussed.

Biological differences between ischemia, hypoglycemia, and epilepsy.

Ischemia, hypoglycemia, and epilepsy have long been thought to produce similar or identical brain damage. Furthermore, these insults have been assumed to be additive in their damaging effects. These notions have been based on neuropathological observations in the hippocampus and cerebral cortex, and on the tenet that energy failure (ischemia, hypoglycemia) and increased demand for energy (epilepsy) similarly give rise to selective neuronal necrosis. Recently, other bases for considering these three insults identical have grown out of observations that loss of calcium homeostasis is common to all and that an excitotoxic mechanism of selective neuronal necrosis exists in all three conditions. Fundamental differences between ischemia, hypoglycemia, and epilepsy include the underlying neurochemical changes induced, the neuronal revival times, the time course of neuronal death, the distribution of selective neuronal necrosis, and the likely excitotoxins released. Lactic acid accumulation, implicated in damage to the neuropil as well as to neuronal cell bodies, also occurs to different degrees and in different distributions in the three conditions. The degree and distribution of pannecrosis is thus also different in ischemia, hypoglycemia, and epilepsy.