Ca2+-Permeable Acid-sensing Ion Channels and Ischemic Brain Injury

Acidosis is a common feature of brain in acute neurological injury, particularly in ischemia where low pH has been assumed to play an important role in the pathological process. However, the cellular and molecular mechanisms underlying acidosis-induced injury remain unclear. Recent studies have demonstrated that activation of Ca²⁺-permeable acid-sensing ion channels (ASIC1a) is largely responsible for acidosis-mediated, glutamate receptor-independent, neuronal injury. In cultured mouse cortical neurons, lowering extracellular pH to the level commonly seen in ischemic brain activates amiloride-sensitive ASIC currents. In the majority of these neurons, ASICs are permeable to Ca²⁺, and an activation of these channels induces increases in the concentration of intracellular Ca²⁺ ([Ca²⁺]_i). Activation of ASICs with resultant [Ca²⁺]_i loading induces time-dependent neuronal injury occurring in the presence of the blockers for voltage-gated Ca²⁺ channels and the glutamate receptors. This acid-induced injury is, however, inhibited by the blockers of ASICs, and by reducing [Ca²⁺]_o. In focal ischemia, intracerebroventricular administration of ASIC1a blockers, or knockout of the ASIC1a gene protects brain from injury and does so more potently than glutamate antagonism. Furthermore, pharmacological blockade of ASICs has up to a 5 h therapeutic time window, far beyond that of glutamate antagonists. Thus, targeting the Ca²⁺-permeable acid-sensing ion channels may prove to be a novel neuroprotective strategy for stroke patients.     

Acid‐sensing ion channel 1a is involved in ischaemia/reperfusion induced kidney injury by increasing renal epithelia cell apoptosis

Acidic microenvironment is commonly observed in ischaemic tissue. In the kidney, extracellular pH dropped from 7.4 to 6.5 within 10 minutes initiation of ischaemia. Acid‐sensing ion channels (ASICs) can be activated by pH drops from 7.4 to 7.0 or lower and permeates to Ca²⁺entrance. Thus, activation of ASIC1a can mediate the intracellular Ca²⁺ accumulation and play crucial roles in apoptosis of cells. However, the role of ASICs in renal ischaemic injury is unclear. The aim of the present study was to test the hypothesis that ischaemia increases renal epithelia cell apoptosis through ASIC1a‐mediated calcium entry. The results show that ASIC1a distributed in the proximal tubule with higher level in the renal tubule ischaemic injury both in vivo and in vitro. In vivo, Injection of ASIC1a inhibitor PcTx‐1 previous to ischaemia/reperfusion (I/R) operation attenuated renal ischaemic injury. In vitro, HK‐2 cells were pre‐treated with PcTx‐1 before hypoxia, the intracellular concentration of Ca²⁺, mitochondrial transmembrane potential (?ψm) and apoptosis was measured. Blocking ASIC1a attenuated I/R induced Ca²⁺ overflow, loss of ?ψm and apoptosis in HK‐2 cells. The results revealed that ASIC1a localized in the proximal tubular and contributed to I/R induced kidney injury. Consequently, targeting the ASIC1a may prove to be a novel strategy for AKI patients.     

High-Density Expression of Ca2+-Permeable ASIC1a Channels in NG2 Glia of Rat Hippocampus

NG2 cells, a fourth type of glial cell in the mammalian CNS, undergo reactive changes in response to a wide variety of brain insults. Recent studies have demonstrated that neuronally expressed acid-sensing ion channels (ASICs) are implicated in various neurological disorders including brain ischemia and seizures. Acidosis is a common feature of acute neurological conditions. It is postulated that a drop in pH may be the link between the pathological process and activation of NG2 cells. Such postulate immediately prompts the following questions: Do NG2 cells express ASICs? If so, what are their functional properties and subunit composition? Here, using a combination of electrophysiology, Ca²⁺ imaging and immunocytochemistry, we present evidence to demonstrate that NG2 cells of the rat hippocampus express high density of Ca²⁺-permeable ASIC1a channels compared with several types of hippocampal neurons. First, nucleated patch recordings from NG2 cells revealed high density of proton-activated currents. The magnitude of proton-activated current was pH dependent, with a pH for half-maximal activation of 6.3. Second, the current-voltage relationship showed a reversal close to the equilibrium potential for Na⁺. Third, psalmotoxin 1, a blocker specific for the ASIC1a channel, largely inhibited proton-activated currents. Fourth, Ca²⁺ imaging showed that activation of proton-activated channels led to an increase of [Ca²⁺]_i. Finally, immunocytochemistry showed co-localization of ASIC1a and NG2 proteins in the hippocampus. Thus the acid chemosensor, the ASIC1a channel, may serve for inducing membrane depolarization and Ca²⁺ influx, thereby playing a crucial role in the NG2 cell response to injury following ischemia.     

Inhibition of acid-sensing ion channel 1a in hepatic stellate cells attenuates PDGF-induced activation of HSCs through MAPK pathway

Acid-sensing ion channels (ASICs), a group of Na⁺-selective and Ca²⁺-permeant ligand-gated cation channels, can be transiently activated by extracellular acid. Among seven subunits of ASICs, acid-sensing ion channel 1a (ASIC1a), which is responsible for Ca²⁺ transportation, is elevated in response to inflammation, tumor, and ischemic injury in central nervous system and non-neuronal tissues. In this study, we demonstrated for the first time the presence of ASIC1a in rat liver and hepatic stellate cells (HSCs). Furthermore, the expression of ASIC1a was increased in primary HSCs and liver tissues of CCl₄-treated rats, suggesting that ASIC1a may play certain role in liver fibrosis. Interestingly, we identified that the level of ASIC1a was significantly elevated in response to platelet-derived growth factor (PDGF) induction in a time- and dose-dependent manner. It was also established that Ca²⁺-transporting ASIC1a was involved in acid-induced injury of different cell types. Moreover, inhibition or silencing of ASIC1a was able to inhibit PDGF-induced pro-fibrogenic effects of activated rat HSCs, including cell activation, de novo synthesis of extracellular matrix components through mitogen-activated protein kinase signaling pathway. Collectively, our studies identified that ASIC1a was expressed in rat liver and HSCs and provided a strong evidence for the involvement of the ASIC1a in the progression of hepatic fibrosis.     

Native and recombinant ASIC1a receptors conduct negligible Ca2+ entry

Acid Sensing Ion Channels (ASICs) are a family of proton-gated cation channels that play a role in the sensation of noxious stimuli. Of these, ASIC1a is the only family member that is reported to be permeable to Ca²⁺, although the absolute magnitude of the Ca²⁺ current is unclear. Here, we used patch-clamp photometry to determine the contribution of Ca²⁺ to total current through native and recombinant ASIC1a receptors. We found that acidification of the extracellular medium evoked amiloride and psalmotoxin 1-sensitive currents in isolated chick dorsal root ganglion neurons and human embryonic kidney cells, but did not alter fura-2 fluorescence when the bath concentration of Ca²⁺ was close to that found in normal physiological conditions. Further, activation of recombinant ASIC1a receptors also failed to produce measurable changes in fluorescence despite of the fact that the total cation current through the over-expressed receptor was ten-fold larger than that of the native channels. Finally, we imaged a field of intact DRG neurons loaded with the Ca²⁺-sensing dye Fluo-4, and found that acidification increased [Ca²⁺]_i in a small population of cells. Thus, although our whole-field imaging data agree with previous studies that activation of ASIC1a receptors can potentially cause elevations in intracellular free Ca²⁺, our single cell data strongly challenges the view that Ca²⁺ entry through the ASIC1a receptor itself contributes to this response.     

Expression and Activity of Acid-Sensing Ion Channels in the Mouse Anterior Pituitary

Acid sensing ion channels (ASICs) are proton-gated cation channels that are expressed in the nervous system and play an important role in fear learning and memory. The function of ASICs in the pituitary, an endocrine gland that contributes to emotions, is unknown. We sought to investigate which ASIC subunits were present in the pituitary and found mRNA expression for all ASIC isoforms, including ASIC1a, ASIC1b, ASIC2a, ASIC2b, ASIC3 and ASIC4. We also observed acid-evoked ASIC-like currents in isolated anterior pituitary cells that were absent in mice lacking ASIC1a. The biophysical properties and the responses to PcTx1, amiloride, Ca²⁺ and Zn²⁺ suggested that ASIC currents were mediated predominantly by heteromultimeric channels that contained ASIC1a and ASIC2a or ASIC2b. ASIC currents were also sensitive to FMRFamide (Phe-Met-Arg-Phe amide), suggesting that FMRFamide-like compounds might endogenously regulate pituitary ASICs. To determine whether ASICs might regulate pituitary cell function, we applied low pH and found that it increased the intracellular Ca²⁺ concentration. These data suggest that ASIC channels are present and functionally active in anterior pituitary cells and may therefore influence their function.     

In vitro characterization of scaffold-free three-dimensional mesenchymal stem cell aggregates

Cerebral ischemia is a key pathophysiological feature of various brain insults. Inadequate oxygen supply can manifest regionally in stroke or as a result of traumatic brain injury or globally following cardiac arrest, all leading to irreversible brain damage. Mitochondrial function is essential for neuronal survival, since neurons critically depend on ATP synthesis generated by mitochondrial oxidative phosphorylation. Mitochondrial activity depends on Ca²⁺ and is fueled either by Ca²⁺ from the extracellular space when triggered by neuronal activity or by Ca²⁺ released from the endoplasmic reticulum (ER) and taken up through specialized contact sites between the ER and mitochondria known as mitochondrial-associated ER membranes. The coordination of these Ca²⁺ pools is required to synchronize mitochondrial respiration rates and ATP synthesis to physiological demands. In this review, we discuss the role of the proteins involved in mitochondrial Ca²⁺ homeostasis in models of ischemia. The proteins include those important for the Ca²⁺-dependent motility of mitochondria and for Ca²⁺ transfer from the ER to mitochondria, the tethering proteins that bring the two organelles together, inositol 1,4,5-triphosphate receptors that enable Ca²⁺ release from the ER, voltage-dependent anion channels that allow Ca²⁺ entry through the highly permeable outer mitochondrial membrane and the mitochondrial Ca²⁺ uniporter together with its regulatory proteins that permit Ca²⁺ entry into the mitochondrial matrix. Finally, we address those proteins important for the extrusion of Ca²⁺ from the mitochondria such as the mitochondrial Na⁺/Ca²⁺ exchanger or, if the mitochondrial Ca²⁺ concentration exceeds a certain threshold, the mitochondrial permeability transition pore.     

Neuroprotection in ischemia: blocking calcium-permeable acid-sensing ion channels

Ca2+ toxicity remains the central focus of ischemic brain injury. The mechanism by which toxic Ca2+ loading of cells occurs in the ischemic brain has become less clear as multiple human trials of glutamate antagonists have failed to show effective neuroprotection in stroke. Acidosis is a common feature of ischemia and is assumed to play a critical role in brain injury; however, the mechanism(s) remain ill defined. Here, we show that acidosis activates Ca2+ -permeable acid-sensing ion channels (ASICs), inducing glutamate receptor-independent, Ca2+ -dependent, neuronal injury inhibited by ASIC blockers. Cells lacking endogenous ASICs are resistant to acid injury, while transfection of Ca2+ -permeable ASIC1a establishes sensitivity. In focal ischemia, intracerebroventricular injection of ASIC1a blockers or knockout of the ASIC1a gene protects the brain from ischemic injury and does so more potently than glutamate antagonism. Thus, acidosis injures the brain via membrane receptor-based mechanisms with resultant toxicity of [Ca2+]i, disclosing new potential therapeutic targets for stroke.     

Acid-sensing ion channel 3 decreases phosphorylation of extracellular signal-regulated kinases and induces synoviocyte cell death by increasing intracellular calcium

Introduction

Acid-sensing ion channel 3 (ASIC3) is expressed in synoviocytes, activated by decreases in pH, and reduces inflammation in animal models of inflammatory arthritis. The purpose of the current study was to characterize potential mechanisms underlying the control of inflammation by ASIC3 in fibroblast-like synoviocytes (FLS).

Methods

Experiments were performed in cultured FLS from wild-type (WT) and ASIC3-/- mice, ASIC1-/- mice, and people with rheumatoid arthritis. We assessed the effects of acidic pH with and without interleukin-1β on FLS and the role of ASICs in modulating intracellular calcium [Ca²⁺]_i, mitogen activated kinase (MAP kinase) expression, and cell death. [Ca²⁺]_i was assessed by fluorescent calcium imaging, MAP kinases were measured by Western Blots; ASIC, cytokine and protease mRNA expression were measured by quantitative PCR and cell death was measured with a LIVE/DEAD assay.

Results

Acidic pH increased [Ca²⁺]_i and decreased p-ERK expression in WT FLS; these effects were significantly smaller in ASIC3-/- FLS and were prevented by blockade of [Ca²⁺]_i. Blockade of protein phosphatase 2A (PP2A) prevented the pH-induced decreases in p-ERK. In WT FLS, IL-1β increases ASIC3 mRNA, and when combined with acidic pH enhances [Ca²⁺]_i, p-ERK, IL-6 and metalloprotienase mRNA, and cell death. Inhibitors of [Ca²⁺]_i and ERK prevented cell death induced by pH 6.0 in combination with IL-1β in WT FLS.

Conclusions

Decreased pH activates ASIC3 resulting in increased [Ca²⁺]_i, and decreased p-ERK. Under inflammatory conditions, acidic pH results in enhanced [Ca²⁺]_i and phosphorylation of extracellular signal-regulated kinase that leads to cell death. Thus, activation of ASIC3 on FLS by acidic pH from an inflamed joint could limit synovial proliferation resulting in reduced accumulation of inflammatory mediators and subsequent joint damage.     

大鼠全脑缺血后海马CA1区锥体神经元L型钙通道电流的改变(英)

已有研究表明在脑缺血期间及再灌流后早期,海马CA1锥体神经元细胞内钙浓度明显升高,这一钙超载被认为是缺血性脑损伤的重要机制之一.电压依赖性钙通道是介导正常CA1神经元钙内流的主要途径.实验观察了脑缺血再灌流后早期海马CA1锥体神经元电压依赖性L型钙通道的变化.以改良的四血管闭塞法制作大鼠15 min前脑缺血模型,在急性分离的海马CA1神经元上,采用膜片钳细胞贴附式记录L型电压依赖性钙通道电流.脑缺血后CA1神经元L型钙通道的总体平均电流明显增大,这是由于通道的开放概率增加所致.进一步分析单通道动力学显示,脑缺血后通道的开放时间变长,通道的开放频率增大.研究结果提示L型钙通道功能活动增强可能参与了缺血后海马CA1锥体神经元的细胞内钙浓度升高.     

Roles Played by the Na+/Ca2+ Exchanger and Hypothermia in the Prevention of Ischemia-Induced Carrier-Mediated Efflux of Catecholamines into the Extracellular Space: Implications for Stroke Therapy

The release of [³H]dopamine ([³H]DA) and [³H]noradrenaline ([³H]NA) in acutely perfused rat striatal and cortical slice preparations was measured at 37 °C and 17 °C under ischemic conditions. The ischemia was simulated by the removal of oxygen and glucose from the Krebs solution. At 37 °C, resting release rates in response to ischemia were increased; in contrast, at 17 °C, resting release rates were significantly reduced, or resting release was completely prevented. The removal of extracellular Ca²⁺ further increased the release rates of [³H]DA and [³H]NA induced by ischemic conditions. This finding indicated that the Na⁺/Ca²⁺ exchanger (NCX), working in reverse in the absence of extracellular Ca²⁺, fails to trigger the influx of Ca²⁺ in exchange for Na⁺ and fails to counteract ischemia by further increasing the intracellular Na⁺ concentration ([Na⁺]_i). KB-R7943, an inhibitor of NCX, significantly reduced the cytoplasmic resting release rate of catecholamines under ischemic conditions and under conditions where Ca²⁺ was removed. Hypothermia inhibited the excessive release of [³H]DA in response to ischemia, even in the absence of Ca²⁺. These findings further indicate that the NCX plays an important role in maintaining a high [Na⁺]_i, a condition that may lead to the reversal of monoamine transporter functions; this effect consequently leads to the excessive cytoplasmic tonic release of monoamines and the reversal of the NCX. Using HPLC combined with scintillation spectrometry, hypothermia, which enhances the stimulation-evoked release of DA, was found to inhibit the efflux of toxic DA metabolites, such as 3,4-dihydroxyphenylacetaldehyde (DOPAL). In slices prepared from human cortical brain tissue removed during elective neurosurgery, the uptake and release values for [³H]NA did not differ from those measured at 37 °C in slices that were previously maintained under hypoxic conditions at 8 °C for 20 h. This result indicates that hypothermia preserves the functions of the transport and release mechanisms, even under hypoxic conditions. Oxidative stress (H₂O₂), a mediator of ischemic brain injury enhanced the striatal resting release of [³H]DA and its toxic metabolites (DOPAL, quinone). The study supports our earlier findings that during ischemia transmitters are released from the cytoplasm. In addition, the major findings of this study that hypothermia of brain slice preparations prevents the extracellular calcium concentration ([Ca²⁺]_o)-independent non-vesicular transmitter release induced by ischemic insults, inhibiting Na⁺/Cl^?-dependent membrane transport of monoamines and their toxic metabolites into the extracellular space, where they can exert toxic effects.

     

Neuronal calcium signaling via store-operated channels in health and disease

Store-operated calcium entry (SOCE) is the flow of calcium ions (Ca²⁺) into cells in response to the depletion of intracellular Ca²⁺ stores that reside predominantly in the endoplasmic reticulum (ER). The role of SOCE has been relatively well understood for non-excitable cells. It is mediated mostly by the ER Ca²⁺ sensor STIM1 and plasma membrane Ca²⁺ channel Orai1 and serves to sustain Ca²⁺ signaling and refill ER Ca²⁺ stores. In contrast, because of the complexity of Ca²⁺ influx mechanisms that are present in excitable cells, our knowledge about the function of neuronal SOCE (nSOCE) is still nascent. This review summarizes the available data on the molecular components of nSOCE and their relevance to neuronal signaling. We also present evidence of disturbances of nSOCE in neurodegenerative diseases (namely Alzheimer’s disease, Huntington’s disease, and Parkinson’s disease) and traumatic brain injury. The emerging important role of nSOCE in neuronal physiology and pathology makes it a possible clinical target.     

The SERCA2: A Gatekeeper of Neuronal Calcium Homeostasis in the Brain

Calcium (Ca²⁺) ions are prominent cell signaling regulators that carry information for a variety of cellular processes and are critical for neuronal survival and function. Furthermore, Ca²⁺ acts as a prominent second messenger that modulates divergent intracellular cascades in the nerve cells. Therefore, nerve cells have developed intricate Ca²⁺ signaling pathways to couple the Ca²⁺ signal to their biochemical machinery. Notably, intracellular Ca²⁺ homeostasis greatly relies on the rapid redistribution of Ca²⁺ ions into the diverse subcellular organelles which serve as Ca²⁺ stores, including the endoplasmic reticulum (ER). It is well established that Ca²⁺ released into the neuronal cytoplasm is pumped back into the ER by the sarco-/ER Ca²⁺ ATPase 2 (SERCA2), a P-type ion-motive ATPase that resides on the ER membrane. Even though the SERCA2 is constitutively expressed in nerve cells, its precise role in brain physiology and pathophysiology is not well-characterized. Intriguingly, SERCA2-dependent Ca²⁺ dysregulation has been implicated in several disorders that affect cognitive function, including Darier’s disease, schizophrenia, Alzheimer’s disease, and cerebral ischemia. The current review summarizes knowledge on the expression pattern of the different SERCA2 isoforms in the nervous system, and further discusses evidence of SERCA2 dysregulation in various neuropsychiatric disorders. To the best of our knowledge, this is the first literature review that specifically highlights the critical role of the SERCA2 in the brain. Advancing knowledge on the role of SERCA2 in maintaining neuronal Ca²⁺ homeostasis may ultimately lead to the development of safer and more effective pharmacotherapies to combat debilitating neuropsychiatric disorders.     

Effect of ischemia-reperfusion on the post-rest inotropy of isolated perfused rat heart

This paper aims to study of the effects of ischemia‐reperfusion on the post‐rest inotropy and to characterize post‐rest B1:B2 ratio as an index of intracellular Ca²⁺ overload. When the rest interval between the cardiac beats is increased, the magnitude of the post‐rest beats is increased. First beat (B1) is maximally potentiated with exponental decline of the second (B2) and subsequent beats, thereby establishing a normal B1:B2 ratio of post‐ rest inotropy of the cardiac muscle. The rest potentiation of B1 and subsequent decay in the magnitude B2 is thought to develop from the time‐dependent changes in the Ca²⁺‐uptake and release from the sarcoplasmic reticulum (SR). Ca²⁺‐kinetics of SR can be modulated by a variety of interventions which produce Ca²⁺ loading of the SR. Methods: Isolated perfused (K‐H buffer, 34°C) rat hearts were paced at 1 Hz (steady state frequency). Interbeat intervals between 1s and 10s were introduced and the recovery in the left ventricular contractile force (Pmax) of post‐rest B1 and B2 for each interval was recorded. Their relative relationship was computed and compared under control and experimental conditions. Results: High extracellular Ca²⁺ (2.50 to 7.0 mM) or low extracellular Na⁺ (50% of control), and ischemia (60 min, 34°C) ‐ reperfusion (30 min, 34°C) caused the reversal of the control relationship of the B1 to B2, with B2 being more potentiated than B1, accompanied by the appearance of after‐contractions during the rest intervals of 4s or more. The mean (± SE) control B1:B2 ratio (at 4s interval) of 1.12 ± 0.05 was significantly (P<0.001) reduced to 0.93 ± 0.07; 0.89 ± 0.01; and 0.96 ± 0.02 after high Ca²⁺ (6 mM) perfusion, low Na⁺(50%) perfusion and ischemia‐reperfusion respectively. Simultaneous perfusion with ryanodine (1 μM) abolished the after‐contractions and significantly increased the reduced ratios. The time course of changes in B1:B2 ratio after graded ischemia‐reperfusion showed a significant fall in the ratio between 30 and 60 min of ischemia. A parallel change in Pmax and a significant rise in the left ventricular end‐diastolic pressure, indicating an irreversible phase of the injury was recorded. No significant changes in B1:B2 ratio were detected during the reversible phase (<30 min) of the ischemia‐reperfusion injury. Conclusions: Ischemia‐reperfusion induces significant alterations in the relative ratio of the post‐rest contractions of the left ventricle in isolated perfused rat heart. The altered ratios were characterized to predict the irreversibility of the reperfusion injury and to index the extent of Ca²⁺‐loading of the sarcoplasmic reticulum.     

STIM and Orai in platelet function

Physiological platelet activation and thrombus formation are essential to stop bleeding in case of vascular injury, whereas inadequate triggering of the same process in diseased vessels can lead to fatal thromboembolism and tissue ischemia of vital organs. A central step in platelet activation is agonist-induced elevation of the intracellular Ca²⁺ concentration. This happens on the one hand through the release of Ca²⁺ from intracellular stores and on the other hand through Ca²⁺ influx from the extracellular space. In platelets, the major Ca²⁺ influx pathway is the so-called store operated Ca²⁺ entry (SOCE), induced by store depletion. Studies in the last five years discovered the molecular background of platelet SOCE. Stromal interaction molecule 1 (STIM1) and Orai1, two so far unknown molecules, got in the focus of research. STIM1 was found to be the Ca²⁺ sensor in the endoplasmic reticulum (ER) membrane, whereas Orai1 was identified as the major store operated Ca²⁺ (SOC) channel in the plasma membrane. These two molecules and their role in platelet function and thrombus formation are the topic of the present review with a special focus on apoptosis and apoptosis-like processes in platelet physiology.     

Ischemia-Induced Inhibition of Active Calcium Transport into Gerbil Brain Microsomes: Effect of Anesthetics and Models of Ischemia

The excessive increase in intracellular Ca²⁺ concentration is associated with events linking cerebral blood flow reduction to neuronal cell damage. We have investigated the possible effect of ischemia and ischemia-reperfusion injury on endoplasmic reticulum (ER) Ca²⁺ transport. Two different models of ischemia as well as two different anesthetics were used. 5 min and 15 min of global forebrain ischemia caused significant depression of the rate of microsomal Ca²⁺ accumulation in pentobarbital anesthetised gerbils. The Ca²⁺ uptake activity recovered partially after 1 hour of reperfusion. Unlike pentobarbital anesthetised gerbils, no significant changes were detected in the active microsomal Ca²⁺-transport after 10 min of global forebrain ischemia in gerbil forebrain and hippocampus under halothane anesthesia. In addition, using the model of decapitation ischemia, we observed significant changes of the Ca²⁺ uptake in both halothane and pentobarbital anesthetised gerbils. These findings indicate that ischemic insult alters the brain microsomal Ca²⁺ transport which is not due to inhibition of the Ca²⁺-ATPase activity. However, the effect of ischemia on this transport system is dependent on the model of ischemia and on the type of anesthetics.     

The APPswe/PS1A246E mutations in an astrocytic cell line leads to increased vulnerability to oxygen and glucose deprivation,Ca2+ dysregulation,and mitochondrial abnormalities

Growing evidence suggests a close relationship between Alzheimer′s Disease (AD ) and cerebral hypoxia. Astrocytes play a key role in brain homeostasis and disease states, while some of the earliest changes in AD occur in astrocytes. We have therefore investigated whether mutations associated with AD increase astrocyte vulnerability to ischemia. Two astroglioma cell lines derived from APP_SWE /PS 1A246E (APP , amyloid precursor protein; PS 1, presenilin 1) transgenic mice and controls from normal mice were subjected to oxygen and glucose deprivation (OGD ), an in vitro model of ischemia. Cell death was increased in the APP_SWE /PS 1A246E line compared to the control. Increasing extracellular calcium concentration ([Ca²⁺]) exacerbated cell death in the mutant but not in the control cells. In order to explore cellular Ca²⁺ homeostasis, the cells were challenged with ATP or thapsigargin and [Ca²⁺] was measured by fluorescence microscopy. Changes in cytosolic Ca²⁺ concentration ([Ca²⁺]_c) were potentiated in the APP_SWE /PS 1A246E transgenic line. Mitochondrial function was also altered in the APP_SWE /PS 1A246E astroglioma cells; mitochondrial membrane potential and production of reactive oxygen species were increased, while mitochondrial basal respiratory rate and ATP production were decreased compared to control astroglioma cells. These results suggest that AD mutations in astrocytes make them more sensitive to ischemia; Ca²⁺ dysregulation and mitochondrial dysfunction may contribute to this increased vulnerability. Our results also highlight the role of astrocyte dyshomeostasis in the pathophysiology of neurodegenerative brain disorders.