Calcium homeostasis following traumatic neuronal injury

Cell death and dysfunction following traumatic brain injury (TBI) consists of a primary phase, which causes immediate consequences to cells by direct mechanical disruption of the brain, and a secondary phase which consists of delayed events initiated at the time of insult. One of the major culprits that contributes to delayed neuronal damage and death after a traumatic insult is the calcium ion. The original calcium hypothesis suggests that a large, sustained influx of calcium into cells initiates cell death signalling cascades. While much of this original tenant remains true, recent findings suggest that the role of calcium in traumatic neuronal injury may be more complex. For example, a sustained level of intracellular free calcium is not necessarily lethal, but the specific route of calcium entry may couple calcium directly to cell death pathways. Other sources of calcium, such as intracellular calcium stores, can also cause cell damage. In addition, calcium-mediated signal transduction pathways have been found to be altered following injury. These alterations are sustained for several hours and may contribute to dysfunction in neurons that do not necessarily die after a traumatic episode. This review provides an overview of experimental evidence that has led to our current understanding of the role of calcium in neuronal death and dysfunction after TBI. While the focus is on alterations in neuronal calcium homeostasis following mechanical injury, these findings may have implications for other pathological states of the brain, such as ischaemia and neurodegenerative disease.     

Recombinant human ciliary neurotrophic factor alters the threshold of hippocampal pyramidal neuron sensitivity to excitotoxin damage: synergistic effects of monosialogangliosides.

Ciliary neurotrophic factor (CNTF) is a multifunctional protein which not only promotes neuronal survival in vitro and in vivo but also controls cell division of neuronal precursors, transmitter differentiation, and glial cell differentiation. Recent studies have indicated that neurotrophic factors can alter hippocampal neuronal threshold to excitotoxin sensitivity. To examine such a role for CNTF, cultures of rat embryonic hippocampal neurons were maintained with recombinant human CNTF for different times, prior to exposure to a toxic dose of glutamate at 5 days in vitro for a further 24 hr. The cytotoxic action of 200 microM glutamate (approximately 40% of pyramidal neurons remaining after 24 hr) was reduced in a concentration-dependent manner in cultures receiving a prior exposure to CNTF within the first 3 days of cell plating: 30 ng/ml CNTF permitted about 75% of the initial number of pyramidal neurons to survive. Presentation of CNTF less than 48 hr before glutamate challenge was ineffective at up to 100 ng/ml. When pyramidal neurons were cultured with a subthreshold concentration (2 ng/ml) of CNTF together with 10 microM of the monosialoganglioside GM1 (or its inner ester form) in the same paradigm, the resulting neuronal survival was similar to that seen with 30 ng/ml CNTF in the face of a glutamate challenge. Such low doses of either CNTF or ganglioside alone were ineffective. The ability of trophic factors to influence the threshold of neuronal sensitivity to excitatory amino acid injury suggests that these proteins could play an important role in the reparative capacity of acutely traumatized central neurons and in neurodegenerative diseases linked to an excitotoxic mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neuronal cell death in neurodegenerative disorders and oxidative stress]

Mechanisms of the process of neuronal degeneration in neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and Alzheimer's disease (AD) remain unsolved. Oxidative stress might be a possible mechanism of neuronal cell death. Glutamate is an excitatory amino acid and its excessive release can cause intracellular calcium influx, activation of calcium-dependent enzymes such as nitric oxide (NO) synthase (NOS), and production of toxic oxygen radicals. Excessive release of glutamate, therefore, can be used as a model of experimental oxidative stress. Continuous exposure to low levels of glutamate potentiates selective motor neuronal death mediated by NO, which inversely protects nonmotor neurons through the guanylyl cyclase-cGMP cascade. Mesencephalic dopaminergic neurons are resistant to cytotoxicity induced by NO. The protecting mechanism from NO neurotoxicity in dopaminergic neurons is based on inhibition of conversion of NO to peroxynitrite anion, and is possibly due to suppression of superoxide anion production. Dopamine D 2 agonists provide protection mediated not only by the inhibition of dopamine turnover but also via D 2-type dopamine receptor stimulation and the subsequent synthesis of proteins that scavenge free radicals. In addition, nicotinic receptor stimulation may be able to protect neurons from oxidative stress induced by A beta.     

Zn2+ entry produces oxidative neuronal necrosis in cortical cell cultures

Evidence has accumulated that Zn²⁺ plays a central role in neurodegenerative processes following brain injuries including ischaemia or epilepsy. In the present study, we examined patterns and possible mechanisms of Zn²⁺ neurotoxicity. Inclusion of 30–300 μm Zn²⁺ for 30 min caused neuronal necrosis apparent by cell body and mitochondrial swelling in cortical cell cultures. This Zn²⁺ neurotoxicity was not attenuated by antiapoptosis agents, inhibitors of protein synthesis or caspase. Blockade of glutamate receptors or nitric oxide synthase showed no beneficial effect against Zn²⁺ neurotoxicity. Interestingly, antioxidants, trolox or SKF38393, attenuated Zn²⁺-induced neuronal necrosis. Pretreatment with insulin or brain-derived neurotrophic factor increased the Zn²⁺-induced free radical injury. Kainate or AMPA facilitated Zn²⁺ entry and potentiated Zn²⁺ neurotoxicity in a way sensitive to trolox. Reactive oxygen species and lipid peroxidation were generated in the early phase of Zn²⁺ neurotoxicity. These findings indicate that entry and accumulation of Zn²⁺ result in generation of toxic free radicals and then cause necrotic neuronal degeneration under certain pathological conditions in the brain.     

Caspase-mediated suppression of glutamate (AMPA) receptor channel activity in hippocampal neurons in response to DNA damage promotes apoptosis and prevents necrosis: implications for neurological side effects of cancer therapy and neurodegenerative disorders

DNA damage in neurons is implicated in the pathogenesis of several neurodegenerative disorders and may also contribute to the often severe neurological complications in cancer patients treated with chemotherapeutic agents. DNA damage can trigger apoptosis, a form of controlled cell death that involves activation of cysteine proteases called caspases. The excitatory neurotransmitter glutamate plays central roles in the activation of neurons and in processes such as learning and memory, but overactivation of ionotropic glutamate receptors can induce either apoptosis or necrosis. Glutamate receptors of the AMPA (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate) type mediate such physiological and pathological processes in most neurons. We now report that DNA damage can alter glutamate receptor channel activity by a mechanism involving activation of caspases. Whole-cell patch clamp analyses revealed a marked decrease in AMPA-induced currents after exposure of neurons to camptothecin, a topoisomerase inhibitor that induces DNA damage; N-methyl-d-aspartate (NMDA)-induced currents were unaffected by camptothecin. The decrease in AMPA-induced current was accompanied by a decreased calcium response to AMPA. Pharmacological inhibition of caspases abolished the effects of camptothecin on AMPA-induced current and calcium responses, and promoted excitotoxic necrosis. Combined treatment with glutamate receptor antagonists and a caspase inhibitor prevented camptothecin-induced neuronal death. Caspase-mediated suppression of AMPA currents may allow neurons with damaged DNA to withdraw their participation in excitatory circuits and undergo apoptosis, thereby avoiding widespread necrosis. These findings have important implications for treatment of patients with cancer and neurodegenerative disorders.     

Excitotoxic neuroprotection and vulnerability with CaMKII inhibition

Aberrant calcium signaling is a common feature of ischemia and multiple neurodegenerative diseases. While activation of calcium-calmodulin (CaM)-dependent protein kinase II (CaMKII) is a key event in calcium signaling, its role in excitotoxicity is controversial. Our findings demonstrate neuroprotection in neuronal cultures treated with the small molecule (KN-93) and peptide (tat-AIP and tat-CN21) inhibitors of CaMKII immediately prior to excitotoxic glutamate/glycine insult. Unlike KN-93 which blocks CaMKII activation, but not constitutively active forms of CaMKII, tat-CN21 and tat-AIP significantly reduced excitotoxicity in cultured neurons when applied post-insult. We observed that the neuroprotective effects of tat-CN21 are greatest when applied before the toxic glutamate challenge and diminish with time, with the neuroprotection associated with CaMKII inhibition diminishing back to control 3h post glutamate insult. Mechanistically, tat-CN21 inhibition of CaMKII resulted in an increase in CaMKII activity and the percentage of soluble αCaMKII observed in neuronal lysates 24h following glutamate stimulation. To address the impact of prolonged CaMKII inhibition prior to excitotoxic insult, neuronal cultures were treated with CaMKII inhibitors overnight and then subjected to a sub-maximal excitotoxic insult. In this model, CaMKII inhibition prior to insult exacerbated neuronal death, suggesting that a loss of CaMKII enhances neuronal vulnerability to glutamate. Although changes in αCaMKII or NR2B protein levels are not responsible for this enhanced glutamate vulnerability, this process is blocked by the protein translation inhibitor cycloheximide. In total, the neuroprotection afforded by CaMKII inhibition can be seen as neuroprotective immediately surrounding the excitotoxic insult, whereas sustained CaMKII inhibition produced by excitotoxicity leads to neuronal death by enhancing neuronal vulnerability to glutamate.     

Glutamate excitotoxicity attenuates insulin-like growth factor-I prosurvival signaling

Recent evidence suggests that impaired insulin/insulin-like growth factor I (IGF-I) input may be associated to neurodegeneration. Several major neurodegenerative diseases involve excitotoxic cell injury whereby excess glutamate signaling leads to neuronal death. Recently it was shown that glutamate inactivates Akt, a serine-kinase crucially involved in the prosurvival actions of IGF-I. We now report that excitotoxic doses of glutamate antagonize Akt activation by IGF-I and inhibit the neuroprotective effects of this growth factor on cultured neurons. Glutamate induces loss of sensitivity to IGF-I by phosphorylating the IGF-I receptor docking protein insulin-receptor-substrate (IRS)-1 in Ser(307) through a pathway involving activation of PKA and PKC in a hierarchical fashion. Administration of Ro320432, a selective PKC inhibitor, abrogates the inhibitory effects of glutamate on IGF-I-induced Akt activation in vitro and in vivo and is sufficient to block the neurotoxic action of glutamate on cultured neurons. Notably, administration of Ro320432 after ischemic insult, a major form of excitotoxic injury in vivo, results in a marked decrease ( approximately 50%) in infarct size. Therefore, uncoupling of IGF-I signaling by glutamate may constitute an additional route contributing to excitotoxic neuronal injury. Further work should determine the potential use of PKC inhibitors as a novel therapeutic strategy in ischemia and other excitotoxic insults.     

NT-3 and BDNF protect CNS neurons against metabolic/excitotoxic insults

Neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) were recently shown to have biological activity in central neurons. In the present study, NT-3 and BDNF attenuated glucose deprivation-induced neuronal damage dose-dependently in rat hippocampal, septal and cortical cultures. Direct measurements of intraneuronal free calcium levels ([Ca²⁺]_i) and manipulations of calcium inlux demonstrated that NT-3 and BDNF each prevented the elevation of [Ca²⁺]_i that mediated glucose deprivation-induced injury. Studies in cultures depleted of glia indicateda direct action of NT-3 and BDNF on neurons. Neurons pretreated with NT-3 or BDNF for 24 hr were more resistant to glutamate neurotoxicity, and showed attenuated [Ca²⁺]_i responses to glutamate. TrkB (BDNF receptor) and trkC (NT-3 receptor) proteins were present in hippocampal, cortical and septal cultures where they were localied to neuronal cell bodies and neurites. The data demonstrate that NT-3 and BDNF can protect neurons against metabolic and excitotoxic insults, and suggest that these neurotrophins may serve [Ca²⁺]_i-stabilizing and neuroprotective functions in the brain.     

Neuroprotection and antioxidants

Ischemia as a serious neurodegenerative disorder causes together with reperfusion injury many changes in nervous tissue. Most of the neuronal damage is caused by complex of biochemical reactions and substantial processes, such as protein agregation, reactions of free radicals, insufficient blood supply, glutamate excitotoxicity, and oxidative stress. The result of these processes can be apoptotic or necrotic cell death and it can lead to an irreversible damage. Therefore, neuroprotection and prevention of the neurodegeneration are highly important topics to study. There are several approaches to prevent the ischemic damage. Use of many modern therapeutical methods and the incorporation of several substances into the diet of patients is possible to stimulate the endogenous protective mechanisms and improve the life quality.     

Adenoviral expression of CREB protects neurons from apoptotic and excitotoxic stress

In this study we have used a molecular approach to manipulate CREB gene expression to study its role in the regulation of neuronal cell death. To achieve this, adenoviral (Ad) vectors encoding EGFP, CREB, and a powerful CREB dominant-negative, known as A-CREB were constructed. The over-expression of CREB but not A-CREB was found to protect primary hippocampal neurons from staurosporine-induced apoptosis, glutamate induced excitotoxicity and exposure to an in vitro ischaemic stress. Hence, manipulating CREB-regulated pathways may provide a means of delaying or preventing the neuronal cell death associated with ischaemic related injury, and in neurodegenerative diseases such as Huntington's and Alzheimer's disease.     

Preclinical testing of neuroprotective neurotrophic factors in a model of chronic motor neuron degeneration.

Many neurotrophic factors have been shown to enhance survival of embryonic motor neurons or affect their response to injury. Few studies have investigated the potential effects of neurotrophic factors on more mature motor neurons that might be relevant for neurodegenerative diseases. Using organotypic spinal cord cultures from postnatal rats, we have demonstrated that insulin-like growth factor-I (IGF-I) and glial-derived neurotrophic factor (GDNF) significantly increase choline acetyltransferase (ChAT) activity, but brain-derived neurotrophic factor (BDNF), neurotrophin-4 (NT-4/5), and neurotrophin-3 (NT-3) do not. Surprisingly, ciliary neurotrophic factor (CNTF) actually reduces ChAT activity compared to age-matched control cultures. Neurotrophic factors have also been shown to alter the sensitivity of some neurons to glutamate neurotoxicity, a postulated mechanism of injury in the neurodegenerative disease, amyotrophic lateral sclerosis (ALS). Incubation of organotypic spinal cord cultures in the presence of the glutamate transport inhibitor threo-hydroxyaspartate (THA) reproducibly causes death of motor neurons which is glutamate-mediated. In this model of motor neuron degeneration, IGF-I, GDNF, and NT-4/5 are potently neuroprotective, but BDNF, CNTF, and NT-3 are not. The organotypic glutamate toxicity model appears to be the best preclinical predictor to date of success in human clinical trials in ALS.     

Uric acid protects neurons against excitotoxic and metabolic insults in cell culture,and against focal ischemic brain injury in vivo

Uric acid is a well-known natural antioxidant present in fluids and tissues throughout the body. Oxyradical production and cellular calcium overload are believed to contribute to the damage and death of neurons that occurs following cerebral ischemia in victims of stroke. We now report that uric acid protects cultured rat hippocampal neurons against cell death induced by insults relevant to the pathogenesis of cerebral ischemia, including exposure to the excitatory amino acid glutamate and the metabolic poison cyanide. Confocal laser scanning microscope analyses showed that uric acid suppresses the accumulation of reactive oxygen species (hydrogen peroxide and peroxynitrite), and lipid peroxidation, associated with each insult. Mitochondrial function was compromised by the excitotoxic and metabolic insults, and was preserved in neurons treated with uric acid. Delayed elevations of intracellular free calcium levels induced by glutamate and cyanide were significantly attenuated in neurons treated with uric acid. These data demonstrate a neuroprotective action of uric acid that involves suppression of oxyradical accumulation, stabilization of calcium homeostasis, and preservation of mitochondrial function. Administration of uric acid to adult rats either 24 hr prior to middle cerebral artery occlusion (62.5 mg uric acid/kg, intraperitoneally) or 1 hr following reperfusion (16 mg uric acid/kg, intravenously) resulted in a highly significant reduction in ischemic damage to cerebral cortex and striatum, and improved behavioral outcome. These findings support a central role for oxyradicals in excitotoxic and ischemic neuronal injury, and suggest a potential therapeutic use for uric acid in ischemic stroke and related neurodegenerative conditions. J. Neurosci. Res. 53:613–625, 1998. © 1998 Wiley-Liss, Inc.     

Calcium-dependent epileptogenesis in an in vitro model of stroke-induced "epilepsy"

PURPOSE: Stroke is the most common cause of acquired epilepsy. The purpose of this investigation was to characterize the role of calcium in the in vitro, glutamate injury-induced epileptogenesis model of stoke-induced epilepsy. METHODS: Fura-2 calcium imaging and whole-cell current clamp electrophysiology techniques were used to measure short-term changes in neuronal free intracellular calcium concentration and long-term alterations in neuronal excitability in response to epileptogenic glutamate injury (20 microM, 10 min) under various extracellular calcium conditions and in the presence of different glutamate-receptor antagonists. RESULTS: Glutamate injury-induced epileptogenesis was associated with prolonged, reversible elevations of free intracellular calcium concentration during and immediately after injury and chronic hyperexcitability manifested as spontaneous recurrent epileptiform discharges for the remaining life of the cultures. Epileptogenic glutamate exposure performed in solutions containing low extracellular calcium, barium substituted for calcium, or N-methyl-d-aspartate (NMDA)-receptor antagonists reduced the duration of intracellular calcium elevation and inhibited epileptogenesis. Antagonism of non-NMDA-receptor subtypes had no effect on glutamate injury-induced calcium changes or the induction epileptogenesis. The duration of the calcium elevation and the total calcium load statistically correlated with the development of epileptogenesis. Comparable elevations in neuronal calcium induced by non-glutamate receptor-mediated pathways did not cause epileptogenesis. CONCLUSIONS: This investigation indicates that the glutamate injury-induced epileptogenesis model of stroke-induced epilepsy is calcium dependent and requires NMDA-receptor activation. Further, these experiments suggest that prolonged, reversible elevations in neuronal free intracellular calcium initiate the long-term plasticity changes that underlie the development of injury-induced epilepsy.     

Calbindin‐D28K is increased in the ventral horn of spinal cord by neuroprotective factors for motor neurons

Slow glutamate‐mediated neuronal degeneration is implicated in the pathophysiology of motor neuron diseases such as amyotrophic lateral sclerosis (ALS). The calcium‐binding proteins calbindin‐D_28K and parvalbumin have been reported to protect neurons against excitotoxic insults. Expression of calbindin‐D_28K is low in adult human motor neurons, and vulnerable motor neurons additionally may lack parvalbumin. Thus, it has been speculated that the lack of calcium‐binding proteins may, in part, be responsible for early degeneration of the population of motor neurons most vulnerable in ALS. Using a rat organotypic spinal cord slice system, we examined whether the most potent neuroprotective factors for motor neurons can increase the expression of calbindin‐D_28K or parvalbumin proteins in the postnatal spinal cord. After 4 weeks of incubation of spinal cord slices with 1) glial cell line‐derived neurotrophic factor (GDNF), 2) neurturin, 3) insulin‐like growth factor I (IGF‐I), or 4) pigment epithelium‐derived factor (PEDF), the number of calbindin‐D_28K‐immunopositive large neurons (>20 μm) in the ventral horn was higher under the first three conditions, but not after PEDF, compared with untreated controls. Under the same conditions, parvalbumin was not upregulated by any neuroprotective factor. The same calbindin increase was true of IGF‐I and GDNF in a parallel glutamate toxicity model of motor neuron degeneration. Taken together with our previous reports from the same model, which showed that all these neurotrophic factors can potently protect motor neurons from slow glutamate injury, the data here suggest that upregulation of calbindin‐D_28K by some of these factors may be one mechanism by which motor neurons can be protected from glutamate‐induced, calcium‐mediated excitotoxicity. © 2015 Wiley Periodicals, Inc.     

Reduced expression of neuronal nicotinic acetylcholine receptors during the early stages of damage by oxidative stress in PC12 cells.

The mechanism for a large loss of neuronal nicotinic acetylcholine receptors (nAChRs) in brains with neurodegenerative diseases remains unclear. Based on our previous results of [(3)H]epibatidine binding influenced by lipid peroxidation, we suggest that nAChR deficit in neurodegenerative diseases might be related to the neurons attacked by free radicals. To further understand how free radicals influence the expression of nAChRs, we detected [(125)I]alpha-bungarotoxin binding, nAChR subunit protein and mRNA during the early stage of damage by oxidative stress in PC12 cells in the present study. The results showed that free radical insult (FeSO(4)) within the concentration range (1 -100 microM) used in the study induced dose-dependent increases in lipid peroxidation and toxicity to PC12 cells, but did not result in apoptosis or necrosis. Significant reductions in [(125)I]alpha-bungarotoxin binding site, protein level for the alpha3 and alpha7 subunits, and mRNA level for the alpha7 subunit were observed in PC12 cells treated by FeSO(4) at the concentrations without inducing cell death compared to control. Pretreatment of cultural cells with antioxidant such as Vitamin E and reduced glutathione prevented the inhibiting effect of free radicals on [(125)I]alpha-bungarotoxin and [(3)H]epibatidine bindings. The present results further demonstrate that oxidative stress might reduce the number of [(125)I]alpha-bungarotoxin binding site and selectively suppress the expression of the nAChR subunits at protein and mRNA levels during the early stages of damage in PC12 cells.     

The neuronal death induced by endotoxic shock but not that induced by excitatory amino acids requires TNF‐α

We studied, using organotypic hippocampal slices in culture, the role of pro‐inflammatory cytokines, oxygen radicals and nitric oxide in neuronal death induced either by endotoxic insult [interferon (IFN) γ, 24 h followed by lipopolysaccharide, 24 h] or by glutamate receptor‐mediated excitotoxic insult. We demonstrated that neuronal death induced by endotoxic insult was absolutely dependent on the synthesis of tumour necrosis factor alpha (TNF‐α). Indeed, TNF‐α antibodies and SB203580, an inhibitor of p38 stress kinase known to block TNF‐α and other cytokine synthesis, completely protected neurons from the endotoxic insult. Inhibiting oxygen radical and nitric oxide productions also reduced the endotoxic shock. We also showed that after priming the cultures with IFN‐γ, TNF‐α was unable to induce neuronal death unless oxygen‐free radicals were exogenously provided. In contrast, although glutamate receptor‐induced excitotoxicity was associated with a low TNF‐α synthesis and a modest activation of p38 stress kinase, neither TNF‐α antibodies nor SB203580 were able to decrease excitotoxic neuronal insult. We did not reduce glutamate receptor‐induced neuronal death with superoxide dismutase plus catalase. In conclusion, although inflammation follows glutamate receptor‐mediated neurotoxicity, the mechanisms by which an endotoxic insult triggers neuronal death are different from those involved in excitotoxicity.     

Differential regulation of trophic and proinflammatory microglial effectors is dependent on severity of neuronal injury

Microglial activation has been reported to promote neurotoxicity and also neuroprotective effects. A possible contributor to this dichotomy of responses may be the degree to which proximal neurons are injured. The aim of this study was to determine whether varying the severity of neuronal injury influenced whether microglia were neuroprotective or neurotoxic. We exposed cortical neuronal cultures to varying degrees of hypoxia thereby generating mild (<20% death, 30 min hypoxia), moderate (40-60% death, 2 h hypoxia), or severe (>70% death, 6 h hypoxia) injuries. Twenty-four hours after hypoxia, the media from the neuronal cultures was collected and incubated with primary microglial cultures for 24 h. Results showed that the classic microglial proinflammatory mediators including inducible nitric oxide synthase, tumor necrosis factor alpha, and interleukin-1-beta were upregulated only in response to mild neuronal injuries, while the trophic microglial effectors brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor were upregulated in response to all degrees of neuronal injury. Microglia stimulated with media from damaged neurons were co-cultured with hypoxic neurons. Microglia stimulated by moderate, but not mild or severe damage were neuroprotective in these co-cultures. We also showed that the severity-dependent phenomenon was not related to autocrine microglial signaling and was dependent on the neurotransmitters released by neurons after injury, namely glutamate and adenosine 5'-triphosphate. Together our results show that severity of neuronal injury is an important factor in determining microglial release of "toxic" versus "protective" effectors and the resulting neurotoxicity versus neuroprotection.     

Ca2+-ATPase isoforms are expressed in neuroprotection in rat, but not human, neurons

Glutamate excitotoxicity has been implicated in neuronal death and damage in many neurodegenerative disorders. The potential neuroprotective role of the plasma membrane calcium ATPase (PMCA) and the NMDA receptor were investigated in rat and human brain neurons after a glutamate insult. Investigation of potential mechanisms of neuronal survival revealed that surviving rat cerebellar granule cells expressed the mRNA of new PMCA isoforms 2b and 2c. There was no observable change in expression of PMCA isoforms or NMDA receptor NR2 subtypes in human cortical neurons. This study shows that subsets of rat and human neurons are resistant to glutamate-induced excitotoxicity and the mechanisms employed to enable survival differ between rat and human neurons.