脑缺血对癫痫敏感性和脑内CCK及NPY的影响

本实验采用改良栓线法制备 SD大鼠右侧大脑中动脉缺血再灌注模型 ,腹腔注射阈下剂量 (3 5 mg/kg,2 d)戊四唑(PTZ)制备慢性癫痫点燃模型。通过观察大鼠的行为检测其癫痫敏感性的改变 ;用免疫细胞化学方法观察 PTZ点燃脑缺血大鼠的相应脑区的神经肽 Y(NPY)和胆囊收缩素 (CCK)的免疫反应阳性神经元数量和免疫反应活性的变化。结果表明 :脑缺血后大鼠癫痫敏感性明显增强 ,PTZ点燃大鼠缺血侧齿状回颗粒细胞区、海马门部和额叶皮质等处出现大量 CCK阳性神经元且免疫反应强度明显增强 ,而 NPY阳性神经元则大量脱失且免疫反应强度也明显减弱。本研究提示 ,大鼠右侧大脑中动脉缺血再灌后出现的癫痫敏感性增高以及 PTZ点燃大鼠缺血侧额叶皮质、背侧海马齿状回颗粒细胞层和海马门部 CCK中间神经元代偿性增生和大量 NPY中间神经元丢失可能参与癫痫敏感性增高的形成机制。     

Focal,but not global,cerebral ischaemia causes loss of myenteric neurons and upregulation of vasoactive intestinal peptide in mouse ileum

Reduced blood flow to the brain induces cerebral ischaemia, potentially causing central injury and peripheral complications including gastrointestinal (GI) dysfunction. The pathophysiology behind GI symptoms is suspected to be neuropathy in the enteric nervous system (ENS), which is essential in regulating GI function. This study investigates if enteric neuropathy occurs after cerebral ischaemia, by analysing neuronal survival and relative numbers of vasoactive intestinal peptide (VIP) and neuronal nitric oxide synthase (nNOS) expressing neurons in mouse ileum after three types of cerebral ischaemia. Focal cerebral ischaemia, modelled by permanent middle cerebral artery occlusion (pMCAO) and global cerebral ischaemia, modelled with either transient occlusion of both common carotid arteries followed by reperfusion (GCIR) or chronic cerebral hypoperfusion (CCH) was performed on C56BL/6 mice. Sham‐operated mice for each ischaemia model served as control. Ileum was collected after 1–17 weeks, depending on model, and analysed using morphometry and immunocytochemistry. For each group, intestinal mucosa and muscle layer thicknesses, neuronal numbers and relative proportions of neurons immunoreactive (IR) for nNOS or VIP were estimated. No alterations in mucosa or muscle layer thicknesses were noted in any of the groups. Loss of myenteric neurons and an increased number of VIP‐IR submucous neurons were found in mouse ileum 7 days after pMCAO. None of the global ischaemia models showed any alterations in neuronal survival or relative numbers of VIP‐ and nNOS‐IR neurons. We conclude that focal cerebral ischaemia and global cerebral ischaemia influence enteric neuronal survival differently. This is suggested to reflect differences in peripheral neuro‐immune responses.     

Co-existence of necrosis and apoptosis in rat hippocampus following transient forebrain ischemia

Morphological changes of CA1 and CA3 pyramidal neurons in rat hippocampus at different intervals following transient forebrain ischemia were examined to determine the nature of post-ischemic cell death in these regions. In the CA1 region, swelling of small dendrites occurred at approximately 24 h reperfusion. At approximately 48 h reperfusion, swelling was found in large dendrites of many CA1 neurons and the mitochondria and endoplasmic reticulum (ER) were dilated. A small portion of neurons showed chromatin aggregation and nuclear indentation without swelling signs. At approximately 60 h reperfusion, swelling of somata was evident in many neurons. Large dense chromatin clumps with round or ovoid contour were found in other neurons. At 72 and 96 h after ischemia, many large vacuoles and glias with active phagocytosis were observed. At 7 days after ischemia, the tissue was compact and many glias were found in the region. Most of the CA3 neurons had normal appearance after ischemia. A total of 5-10% CA3 neurons exhibited shrinking nuclei and chromatin aggregation at approximately 24 h reperfusion. The number of these neurons decreased overtime and disappeared at 72 h after ischemia. These results demonstrate the co-existence of necrosis and apoptosis in the CA1 region after transient forebrain ischemia. Most CA3 neurons remained intact after ischemia while a small portion of them showed apoptotic cell death.     

Simultaneous electroencephalography and functional magnetic resonance imaging of primary and secondary somatosensory cortex in humans after electrical stimulation

The arcuate nucleus gives rise to approximately 80% of the neuropeptide Y (NPY)-immunoreactive (IR) innervation to thyrotropin-releasing hormone (TRH) neurons in the hypothalamic paraventricular nucleus (PVN). However, the source of the remaining 20% is unknown. Since medullary adrenergic neurons synthesize NPY and also innervate the hypophysiotropic TRH neurons, we raised the possibility that adrenergic neurons contribute to the NPY-ergic innervation of TRH neurons in the PVN. Triple-labeling immunofluorescence was performed to study the colocalization of NPY and phenylethanolamine N-methyltransferase (PNMT)--the key enzyme of adrenaline synthesis--in axons in association with hypophysiotropic TRH neurons. NPY-immunoreactivity was observed in 74% of PNMT-IR axon varicosities apposed to proTRH-IR neurons, comprising 26% of all NPY-IR axons in contact with proTRH perikarya and dendrites in the PVN. We conclude that at least two distinct populations of NPY neurons innervate hypophysiotropic TRH neurons, the NPY neurons of the hypothalamic arcuate nucleus and the medullary adrenergic neurons that co-contain NPY.     

Calcium binding proteins and neuropeptides as molecular markers of GABAergic interneurons in the cat visual cortex

Summary In the cat visual cortex, almost all parvalbumin-positive cells are GABAergic, and about 80% of the calbindin D-28K-positive neurons are also GABA-immunoreactive. About 37% of the GABAergic neurons contain parvalbumin and a smaller fraction (about 18%) contains calbindin. Furthermore, parvalbumin and calbindin are localized in two separate neuronal populations in the cat visual cortex, suggesting that two GABAergic populations can be distinguished, one containing parvalbumin and one containing calbindin. Double staining for parvalbumin and neuropeptides (CCK, SRIF and NPY), revealed no double-labeled cells, with the exception of a few SRIF- and parvalbumin-positive neurons. These results show that cortical GABAergic cells can be differentiated on basis of their calcium binding protein and neuropeptide immunoreactivity.     

Intimate associations between the neuropeptide Y system and the galanin-immunoreactive neurons in the human diencephalon

Galanin and neuropeptide Y (NPY) are among the most abundant neuropeptides in the hypothalamus. The role of NPY and galanin in the regulation of the secretory activity of the anterior pituitary has been well established. In addition, the two peptides interact with a number of neurons synthesizing the releasing and inhibiting hormones and a large number of other neuropeptides. The aim of the present studies was to explore if, as in rodents, NPY innervates galanin-immunoreactive (IR) neurons in the human diencephalon. Due to the long post mortem period and subsequent lack of optimal preservation of the cell membranes in the brain, electron microscopy could not be employed to show the presence of NPY-IR synapses on galanin-IR neurons. Therefore, we used light microscopic double label immunocytochemistry and high magnification microscopy with oil immersion to identify putative juxtapositions between NPY and galanin. Our studies show that similarly to rats, numerous NPY-IR nerve terminals surrounded galanin-IR neurons in the human hypothalamus. Among the hypothalamic regions, the infundibulum (infundibular or arcuate nucleus) contained the largest number of galanin-IR neurons heavily surrounded with NPY-IR nerve terminals. These en passant-type intimate associations between NPY-IR and galanin-IR neuronal elements may be functional synapses and may provide the morphological basis for the NPY-mediated galanin release. Consequently, NPY–galanin communication may mediate effects of NPY on neuronal systems innervated by galanin, and therefore may play a pivotal role in the regulation of reproduction, growth, energy and metabolism.     

Mineralocorticoid and glucocorticoid receptor expressions in astrocytes and microglia in the gerbil hippocampal CA1 region after ischemic insult

In the present study, we observed expression and changes of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) in the gerbil hippocampal CA1 region, but not in the CA2/3 region, after 5 min of transient forebrain ischemia. In blood, corticosterone levels were increased biphasically at 30 min and 12 h after ischemia/reperfusion, and thereafter its levels were decreased. In the sham-operated group, MR and GR immunoreactivities were weakly detected in the CA1 region. By 3 days after ischemia, MR and GR were not significantly altered in the CA1 region: at 12 h after ischemia, GR was expressed in a few neurons in the CA1 region, whereas MR was not expressed in any neurons after ischemic insult. From 4 days after ischemia, MR and GR immunoreactivities were detected in astrocytes and microglia in the CA1 region, and at 7 days after ischemia, MR and GR immunoreactivities peaked in the hippocampal CA1 region. At this time, 55% of astrocytes and 30% of microglia showed MR immunoreactivity, and 20% of astrocytes and 40% of microglia showed GR immunoreactivity. Western blot analyses showed that the pattern of changes in MR and GR protein levels was similar to the immunohistochemical changes observed after transient forebrain ischemia. From 4 days after ischemia, MR and GR protein levels were increased time-dependently after ischemia. In conclusion, enhanced MR and GR expressions in astrocytes and microglia were detected in the hippocampal CA1 region 4-7 days after ischemia/reperfusion. At this time, GR immunoreactivity was abundant in microglia, whereas MR immunoreactivity was prominent in astrocytes. The specific distribution of corticosteroid receptors in the astrocytes and microglia may be associated with the differences of MR and GR functions against ischemic damage.     

Morphological evidence of sensory neurons in the root of the rat tongue

In our previous studies, a large number of substance P (SP)-immunoreactive (IR) nerve fibers were detected in the rat tongue and their number increased after inflammation, suggesting that these fibers might be involved in the axon reflex. Therefore, in this study, we have examined the different neuropeptide-containing nerve elements by light, electron, and confocal laser microscopy. SP, vasoactive intestinal polypeptide (VIP), and neuropeptide Y (NPY) IR varicose fibers were numerous compared with other ones. Small groups of ganglia with perikarya IR for SP, VIP, NPY, galanin, and somatostatin were observed. The SP-IR nerve cell bodies were mainly located in the tunica propria just below the epithelial lining. Double-labeling immunohistochemistry showed that the intrinsic SP-IR neurons did not colocalize VIP. The SP containing nerve terminals were observed in and below the epithelium as well as in very close contact to or making real synapses with other neurons in the intralingual ganglion. Our data confirmed the possibility of intrinsic sensory neurons, which might be the afferent branch of the intralingual reflex arch, while the VIP- and NPY-IR neurons located in the salivary glands, around the blood vessels, and in the muscle layer might constitute the efferent site of this reflex.     

Target-specific projections of intrinsic ganglionic neurons with different chemical codes in the canine larynx.

The distribution and origin of peptide-containing intrinsic nerve fibers within the larynx were examined by immunohistochemistry and denervation experiments in the dog. In the normal larynx, a dense network of vasoactive intestinal polypeptide (VIP)-immunoreactive (IR) fibers was seen around the acini of submucosal glands. VIP-, substance P (SP)-, or calcitonin gene-related peptide (CGRP)-IR fibers were seen in the walls of submucosal arteries, and VIP-, neuropeptide Y (NPY)-, or enkephalin (ENK)-IR fibers were seen around the arteries in the muscle tissue. Most of these peptide-IR fibers remained after bilateral denervation of the superior and inferior laryngeal nerves. Several small intrinsic ganglia were found along the peripheral branches of the laryngeal nerves. About 97% of the ganglionic neurons were VIP-IR; of these, 44% were immunoreactive to VIP alone, 22% to VIP and NPY, 13% to VIP and SP, 7% to VIP and ENK, and 14% to VIP, NPY and SP. These results reveal that the exocrine glands and blood vessels are innervated by the intrinsic ganglionic neurons and that subpopulations of ganglionic neurons with different chemical codes innervate specific target organs in the canine larynx.     

Neuroprotection and neurogenesis: modulation of cornus ammonis 1 neuronal survival after transient forebrain ischemia by prior fimbria-fornix deafferentation

Severe transient forebrain ischemia causes selective neuronal death in the hippocampal cornus ammonis 1 region. We tested the hypothesis that fimbria-fornix deafferentation can provide long-term protection to cornus ammonis 1 neurons and modulate neurogenesis following ischemia. Fimbria-fornix lesion or sham-fimbria-fornix lesion was performed on Wistar rats 13 days prior to 10 min forebrain ischemia or sham ischemia. Temperature was regulated and rats survived for 7, 14 or 28 days. Immunofluorescent bromodeoxyuridine and neuron specific nuclear protein staining and immunochemistry terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling staining were performed. At 7 days after ischemia, 73%+/-14% of cornus ammonis 1 neurons were damaged, while deafferentation reduced the injury to 36%+/-17% of cornus ammonis 1 neurons. This protection persisted for at least 28 days. Ischemia significantly increased the number of bromodeoxyuridine-positive cells (85-90 cells/section in stroke group vs. 6 to 11 cells/section in normal or sham stroke group), with very few terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling-stained cells adjacent to the hippocampal cornus ammonis 1. Fimbria-fornix lesioning followed by ischemia increased the percentage of new neurons 13-fold over ischemia alone and 6.5-fold over sham lesion plus ischemia. The results indicate that fimbria-fornix deafferentation provides long-term neuroprotection in cornus ammonis 1 following forebrain ischemia and promotes neurogenesis after ischemic insults.     

Bicyclol attenuates oxidative stress and neuronal damage following transient forebrain ischemia in mouse cortex and hippocampus

To assess its potential neuroprotective effect against ischemia/reperfusion (IR) injury in mice, bicyclol was administered intragastrically once a day for 3 days. After 6 h of bicyclol pretreatment on the third day, forebrain ischemia was induced for 1 h by bilateral occlusion of the carotid arteries. After different times of reperfusion, the histopathological changes and the levels of mitochondria-generated reactive oxygen species (ROS), malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in the cortex and hippocampus were measured. We found that extensive neuronal death occurred in the cortex and the CA1 area of the hippocampus at day 7 after IR and that bicyclol significantly attenuated IR-induced neuronal death in a dose-dependent manner. We also found that pretreatment with bicyclol dose dependently decreased the generation of ROS and the MDA content and reduced the compensatory increase in SOD activity in the cortex and hippocampus at 4 h of reperfusion. These results suggest that bicyclol protects the mouse brain against cerebral IR injury by attenuating oxidative stress and lipid peroxidation.     

Effect of carvedilol on neuronal survival and poly(ADP-ribose) polymerase activity in hippocampus after transient forebrain ischemia

Carvedilol a beta-adrenoreceptor antagonist with potent antioxidant properties raises high expectations in therapy of ischemia. In this study the effect of carvedilol on neuronal survival after transient forebrain ischemia in gerbils was investigated. The role of poly(ADP-ribose) polymerase (PARP-1) in this process was evaluated. Our data indicated that carvedilol administered subcutaneously in a dose of 7 or 70 mg/kg b.w. directly after 5 min of transient forebrain ischemia protects significant population of neurons in hippocampal area CA1, but has no effect after induction of prolonged 10 min ischemia. Carvedilol significantly decreased PARP activity in hippocampus that was markedly increased after both 15 min and 4 days of reperfusion following 5 min of ischemia. Moreover, carvedilol prevented NAD+ depletion after ischemic-reperfusion insult. These results indicated that carvedilol protects neurons against death and suggested that suppression of PARP activity during reperfusion could be involved in this process.     

脑缺血在在体大鼠海马CA1锥体神经元上诱发出一种NMDA受体介导的新突触后电位

脑缺血在在体大鼠海马ＣＡ１锥体神经元上诱发出一种ＮＭＤＡ受体介导的新突触后电位高天明（ＧａｏＴｉａｎｍｉｎｇ）徐造成（ＸｕＺａｏｃｈｅｎｇ）（田纳西大学医学院神经病学系，盂菲斯，田纳西３８１０４美国；第一军医大学生理学教研室，广州５１０５１５）海马Ｃ...     

多巴胺耗竭对纹状体神经元在缺血再灌注损伤中的保护作用

为探讨全脑缺血再灌注损伤过程中多巴胺对纹状体内各类神经元的影响 ,本实验用 6 -OHDA损毁大鼠一侧黑质多巴胺能神经元以耗竭该侧纹状体内的多巴胺 ,再进行 4血管结扎造成全脑缺血模型 ,3 0 min后分别复灌 6、12、2 4h,行连续冰冻切片 ,以焦油紫染色进行形态学分析 ,用 calbindin-D2 8k及 parvalbum in两种钙结合蛋白抗体进行免疫组化反应。结果显示 :(1)焦油紫染色 :缺血 3 0 min复灌 6 h即可见多巴胺未耗竭侧纹状体内神经元损伤比耗竭侧者明显加重 ,复灌 12及 2 4h组双侧差别更为显著 ;图像分析测定单位面积内焦油紫染色细胞面积总和 ,各实验组两侧间均有显著性或极显著性差异 (P<0 .0 5 ,P<0 .0 1) ;(2 ) calbindin-D2 8k免疫组织化学染色 :各组内多巴胺未耗竭侧纹状体 calbindin-D2 8k阳性反应明显比耗竭侧减弱 ,阳性神经元数量减少、染色变浅 ,图象分析显示其光密度明显低于耗竭侧 ;(3 ) parvalbumin免疫组化反应 :同时间组双侧纹状体 parval-bumin阳性神经元的形态和数量无明显差别 (P>0 .0 5 ) ,但随复灌时间的延长 ,双侧阳性细胞数均有降低。以上结果提示 :脑缺血时大量释放的 DA是纹状体神经元缺血性损伤的重要因素之一 ;纹状体内的 calbindin-D2 8k投射神经元对脑缺血及 DA含量的变化敏感 ,?     

Zinc translocation accelerates infarction after mild transient focal ischemia

Excess release of chelatable zinc (Zn(2+)) from central synaptic vesicles may contribute to the pathogenesis of selective neuronal cell death following transient forebrain ischemia, but a role in neurodegeneration after focal ischemia has not been defined. Adult male Long-Evans rats subjected to middle cerebral artery occlusion (MCAO) for 30 min followed by reperfusion developed delayed cerebral infarction reaching completion 3 days after the insult. One day after the insult, many degenerating cerebral neurons exhibited increased intracellular Zn(2+), and some labeled with the antibody against activated caspase-3. I.c.v. administration of the Zn(2+) chelator, EDTA saturated with equimolar Ca(2+) (CaEDTA), 15 min prior to ischemia attenuated subsequent Zn(2+) translocation into cortical neurons, and reduced infarct volume measured 3 days after ischemia. Although the protective effect of CaEDTA at this endpoint was substantial (about 70% infarct reduction), it was lost when insult severity was increased (from 30 to 60 min MCAO), or when infarct volume was measured at a much later time point (14 days instead of 3 days after ischemia).These data suggest that toxic Zn(2+) translocation, from presynaptic terminals to post-synaptic cell bodies, may accelerate the development of cerebral infarction following mild transient focal ischemia.     

Ischemic preconditioning prevents protein aggregation after transient cerebral ischemia

Transient cerebral ischemia leads to protein aggregation mainly in neurons destined to undergo delayed neuronal death after ischemia. This study utilized a rat transient cerebral ischemia model to investigate whether ischemic preconditioning is able to alleviate neuronal protein aggregation, thereby protecting neurons from ischemic neuronal damage. Ischemic preconditioning was introduced by a sublethal 3 min period of ischemia followed by 48 h of recovery. Brains from rats with either ischemic preconditioning or sham-surgery were then subjected to a subsequent 7 min period of ischemia followed by 30 min, 4, 24, 48 and 72 h of reperfusion. Protein aggregation and neuronal death were studied by electron and confocal microscopy, as well as by biochemical analyses. Seven minutes of cerebral ischemia alone induced severe protein aggregation after 4 h of reperfusion mainly in CA1 neurons destined to undergo delayed neuronal death (which took place after 72 h of reperfusion). Ischemic preconditioning reduced significantly protein aggregation and virtually eliminated neuronal death in CA1 neurons. Biochemical analyses revealed that ischemic preconditioning decreased accumulation of ubiquitin-conjugated proteins (ubi-proteins) and reduced free ubiquitin depletion after brain ischemia. Furthermore, ischemic preconditioning also reduced redistribution of heat shock cognate protein 70 and Hdj1 from cytosolic fraction to protein aggregate-containing fraction after brain ischemia. These results suggest that ischemic preconditioning decreases protein aggregation after brain ischemia.     

Associations between the human growth hormone-releasing hormone- and neuropeptide-Y-immunoreactive systems in the human diencephalon: a possible morphological substrate of the impact of stress on growth

Previous studies revealed that stress is a pivotal factor in the regulation of growth. Psychological harassment may result in psychosocial dwarfism with delayed puberty, short stature and depression. Growth hormone (GH) secretion is suppressed by stress, possibly via the attenuation of growth hormone-releasing hormone (GHRH) secretion. However, the morphological substrate of this phenomenon has not been elucidated yet. Since neuropeptide Y (NPY) levels in the plasma is increased by administration of various stressors, the common consensus is that NPY plays a crucial role in the stress response. In the present study, we examined the putative juxtapositions between the NPY- and GHRH-immunoreactive (IR) systems in the human hypothalamus using double-label immunohistochemistry. Our findings revealed that the majority of the GHRH-IR perikarya formed intimate associations with NPY-IR fiber varicosities. The majority of these juxtapositions were found in the infundibular nucleus/median eminence where NPY-IR fiber varicosities often covered a significant surface area of the GHRH neurons. Since the juxtapositions between the GHRH-IR perikarya and NPY-IR fiber varicosities may be functional synapses, they may represent the morphological substrate of stress-suppressed GH secretion. The large number of contacting elements indicates that NPY plays a pivotal role in GH release, and may be considered as a major factor in the attenuation of growth by stress in humans.     

Glial cell line-derived neurotrophic factor protects against delayed neuronal death after transient forebrain ischemia in rats

Glial cell line-derived neurotrophic factor, a member of the transforming growth factor-beta superfamily, is a potent neurotrophic factor, which has a variety of biological activities that affect several types of neurons in both the central and peripheral nervous systems. In this study, we examined the effects of glial cell line-derived neurotrophic factor on delayed neuronal death in the hippocampal CA1 region of rats after transient forebrain ischemia. In the control rats pretreated with the vehicle, transient forebrain ischemia-induced delayed neuronal death in the hippocampal CA1 region was observed seven days after reperfusion. Pretreatment with glial cell line-derived neurotrophic factor (1.0 microg), which was directly microinjected into the right hippocampal CA1 region, gave significant protection against the delayed hippocampal neuronal death. On the contralateral side of the hippocampus, which was not injected with glial cell line-derived neurotrophic factor, delayed neuronal death similar to that seen in vehicle-treated control animals was observed. Intracerebroventricular glial cell line-derived neurotrophic factor (2.5 microg) injection also protected against delayed neuronal death. In addition, pretreatment with glial cell line-derived neurotrophic factor gave significant protection against apoptotic cell death induced by brain ischemia in the hippocampal CA1 region, as determined by in situ staining for DNA fragmentation. These findings suggest that glial cell line-derived neurotrophic factor plays an important role in delayed neuronal death induced by brain ischemia.     

Accumulation of 4-hydroxynonenal-modified proteins in hippocampal CA1 pyramidal neurons precedes delayed neuronal damage in the gerbil brain

It has been proposed that reactive oxygen species and lipid peroxidation have a role in the delayed neuronal death of pyramidal cells in the CA1 region. To explore the in situ localization and serial changes of 4-hydroxy-2-nonenal-modified proteins, which are major products of membrane peroxidation, we used immunohistochemistry of the gerbil hippocampus after transient forebrain ischemia with or without preconditioning ischemia. The normal gerbil hippocampus showed weak immunoreactivity for 4-hydroxy-2-nonenal-modified proteins in the cytoplasm of CA1 pyramidal cells. 4-hydroxy-2-nonenal immunoreactivity showed no marked changes after preconditioning ischemia. In the early period after ischemia and reperfusion, there was a transient increase of nuclear 4-hydroxy-2-nonenal immunoreactivity in CA1 pyramidal neurons. In contrast, cytoplasmic immunoreactivity transiently disappeared during same period and then increased markedly from 8h to seven days. One week after ischemia, 4-hydroxy-2-nonenal immunoreactivity was observed within reactive astrocytes in the CA1 region. Early nuclear accumulation of 4-hydroxy-2-nonenal in CA1 neurons may indicate a possible role in signal transduction between the nucleus and cytoplasm/mitochondria, while delayed accumulation of 4-hydroxy-2-nonenal-modified proteins in the cytoplasm may be related to mitochondrial damage.We conclude that 4-hydroxy-2-nonenal may be a key mediator of the oxidative stress-induced neuronal signaling pathway and may have an important role in modifying delayed neuronal death.     

DNA cleavage and proteolysis of microtubule-associated protein 2 after cerebral ischemia of different severity.

We report temporal profiles of cytoplasmic proteolysis and genomic DNA cleavage after cerebral ischemia of different severity in gerbils. Global forebrain ischemia by bilateral common carotid artery occlusion for 5 min with reperfusion, severe unilateral hemispheric ischemia by unilateral common carotid artery occlusion for 30 min with reperfusion, and complete ischemia by decapitation were used. The hippocampus was examined for proteolysis by using immunohistochemistry for microtubule-associated protein 2, DNA cleavage by using in situ nick-end labelling, and nuclear morphology by Hematoxylin staining. During evolution of delayed neuronal death after transient forebrain ischemia, loss of the immunoreaction for microtubule-associated protein 2 occurred almost in parallel with DNA cleavage in the CA1 region. In contrast, disappearance of the immunoreaction for microtubule-associated protein 2 was much faster than genomic DNA cleavage after unilateral hemispheric ischemia and reperfusion. The microtubule-associated protein 2 immunoreactivity was completely lost before development of changes in nuclear morphology or DNA cleavage after complete ischemia. The present study demonstrated the differences between necrosis and delayed neuronal death, but the nuclear morphology in the latter was not exactly the same as seen in apoptosis. Some elements of both necrotic and apoptotic machineries may work following transient ischemia, and the degree of ischemic insult may determine the character of cell death process.