Prolonged exposure to hypobaric hypoxia transiently reduces GABAA receptor number in mice cerebral cortex

The central nervous system is severely affected by hypoxic conditions, which produce alterations in neural cytoarchitecture and neurotransmission, resulting in a variety of neuropathological conditions such as convulsive states, neurobehavioral impairment and motor CNS alterations. Some of the neuropathologies observed in hypobaric hypoxia, corresponding to high altitude conditions, have been correlated with a loss of balance between excitatory and inhibitory neurotransmission, produced by alterations in glutamatergic and GABAergic receptors. In the present work, we have studied the effect of chronic hypobaric hypoxia (506 hPa, 18 h/day×21 days) applied to adult male mice on GABA_A receptors from cerebral cortex, to determine whether hypoxic exposure may irreversibly affect central inhibitory neurotransmission. Saturation curves for [³H]GABA specifically bound to GABA_A receptors in isolated synaptic membranes showed a 30% decrease in maximal binding capacity after hypoxic exposure (B_max control, 4.70±0.19, hypoxic, 3.33±0.10 pmol/mg protein), with no effect on GABA binding sites affinity (K_d control: 159.3±13.3 nM, hypoxic: 164.2±15.1 nM). Decreased B_max values were observed up to the 10th post-hypoxic day, returning to control values by the 15th post-hypoxic day. Pharmacological properties of GABA_A receptor were also affected by hypoxic exposure, with a 45 to 51% increase in the maximal effect by positive allosteric modulators (pentobarbital and 5α-pregnan-3α-ol-20-one). We conclude that long-term hypoxia produces a significant but reversible reduction on GABA binding to GABA_A receptor sites in cerebral cortex, which may reflect an adaptive response to this sustained pathophysiological state.     

Ultrastructural concomitants of hypoxia-induced angiogenesis

Prolonged hypoxia results in structural and functional adaptive responses to improve tissue oxygen delivery. Structural changes within the brain include vascular proliferation and elongation. The aim of the current study was to investigate whether ultrastructural changes in capillary walls also occur as part of the adaptive response. Adult rats were exposed to 2 or 3 weeks of moderate hypobaric hypoxia at 0.5 atmospheres and their cerebral microvasculature examined using quantitative ultrastructural methods. We found that hypoxic rats had an 18% increase in their brain capillary diameter but no change in endothelial wall thickness, basement membrane thickness, or coverage of the endothelial wall by pericytes. The increased diameter of cerebral capillaries may play an important role in decreasing the resistance to capillary perfusion which is brought about by the increased erythrocyte fraction in the blood of hypoxic rats. Ultrastructural features relevant to the blood-brain barrier were maintained in hypoxic rats. Pericytes, that are thought to form a second line of defense in the blood-brain barrier, maintained their numerical and size relationships to the endothelial cells. Endothelial junctions were unchanged and endothelial vesicles were somewhat lower in density than normal at 2 weeks of hypoxia, but had regained their normal density by 3 weeks. Mitochondria of the brain capillary endothelial cells maintained normal numerical and volume densities in hypoxia, but the mitochondria of the surrounding neuropil were decreased significantly by about 30%. Received: 2 July 1996 / Revised, accepted: 6 December 1996     

Quantitative capillary topography and blood flow in the cerebral cortex of cats: an in vivo microscopic study

In 50 anesthetized cats the microcirculation in intermediate and deeper layers of the cerebral cortex was visualized in vivo by microtransillumination, and documented by high-speed microcinephotography. The viability of the preparation was verified in a series of experiment s demonstrating spontaneous vasomotion and responsiveness to chemical stimulation of pial arterioles and small arteries. Stereological methods for quantitative analysis of projected images of capillaries in a comparatively large tissue volume were employed to determine morphometric and topographical parameters of the asymmetric, highly tortuous intracortical capillary network. Capillary diameters (5.1 ± 0.84μm), radii of curvature (median 57 μm), total capillary lengths per tissue volume939 ± 338.2 mm/cu.mm, capillary volume fractions (2.1 ± 0.51%), total capillary surface areas per tissue volume (15.3 ± 4.85 sq.mm/cu.mm), and intercapillary distances (median 24.2 μm) showed significant interregional differences. The frequency distribution of the lenghts of capillary segments (median 108 μm) was best described by a Weibull distribution. On the average 90% of all capillaries were continously perfused. Capillary red cell flow (median velocity 1500 μm/sec) was predominantly unidirectional and conspicuously irregular. The variance of capillary red cell velocities (CRCVs) was significantly correlated (τ = 0.48) with capillary tortuosity. An extreme value distribution best describes the observed frequency distribution of CRCVs. Flow irregularities represented both twice noise and a significant stochastic periodicity at frequencies between 40 and 90 Hz.     

高原低压低氧诱导大鼠脑皮层神经元中脑红蛋白的表达

目的 观察SD大鼠在急慢性高原缺氧环境下脑皮层神经元中脑红蛋白(NGB)的表达及变化. 方法 将70只SD大鼠按照随机数字表法分为2组:正常对照组(10只)和实验组(60只),正常对照组置于海拔2295m 的西宁市内饲养观察,实验组运至海拨4600m的青海大学可可西里高原医学研究基地制作急慢性高原缺氧模型;依据高原暴露时间不同,将实验组又再细分为缺氧12h组、24h组、48h组、72h组、1周组、1月组,每组均10只.血氧饱和度仪测定大鼠外周血氧饱和度,实时定量PCR和Western blot检测高原低压低氧暴露不同时间大鼠脑皮层神经元中NGBmRNA和蛋白的表达含量. 结果 高原低压低氧暴露12h时SD大鼠外周血氧饱和度最低,随着暴露时间的延长,外周血氧饱和度开始缓慢升高,但始终低于对照组,差异有统计学意义(P＜0.05).高原低压低氧暴露24h后NGB mRNA的表达出现首个高峰,此后逐渐下降,至1周时恢复正常,而后缓慢升高,至1月时保持在一个较高水平.Western blot结果显示同样的变化趋势. 结论 在急慢性高原缺氧环境下,NGB表达的增高增强了SD大鼠对低压低氧的耐受性,提示NGB表达变化可能是脑组织对脑缺氧的一种内源性神经保护反应.     

Postnatal hypobaric hypoxia in rats impairs water maze learning and the morphology of neurones and macroglia in cortex and hippocampus

Newborn rats were exposed to intermittent hypobaric hypoxia from birth until the age of 19 days. Spatial memory was tested in a Morris water maze from postnatal day (P) 23 to P32 and from P100 to P109. From P24 to P27 and on days P100 and P101, the escape latencies of hypoxic animals were longer than those of controls. At P24, the number of neuronal bodies increased in cortical layer II of the somatosensory, motor, and auditory areas, and in layer V of the motor area, but the number of neuronal bodies throughout the whole cortical thickness was unchanged. Decreases in the immunostaining density for neurofilaments (anti-NF 160), astrocytes (anti-GFAP), and oligodendrocytes (RIP) were found in the hippocampus, and the typical parallel organisation of neuronal and macroglial processes was lost. Decreases in immunostaining for neurofilaments and oligodendrocytes were also found in the somatosensory cortex and motor cortex. In adult hypoxic rats, at P114-P240, the number of neuronal bodies and the immunostaining density for neurofilaments, astrocytes, and oligodendrocytes in the examined areas were similar to adult controls; however, in the hippocampus we found hypertrophy of fine astrocytic processes and a decreased number of oligodendrocytic processes. We conclude that the neonatal brain damage induced by hypobaric hypoxia impairs spatial memory in infant as well as adult rats. Hypobaric hypoxia delays the maturation of neurones and substantially affects macroglia in the cortex and hippocampus.     

Effect of up-regulation of NMDA receptors on cerebral O2 consumption and blood flow in rat

We tested the hypothesis that cerebrocortical blood flow and O₂ consumption would be proportional to an up-regulated number of functional N-methyl- -aspartate (NMDA) receptors. Previous work had shown a relationship between cerebral metabolism and NMDA receptor activity. We increased the specific binding to NMDA receptors in the cerebral cortex, from 2.2 ± 0.9 to 4.5 ± 0.8 (density units) in male Long-Evans rats by daily giving two intraperitoneal injections (30 mg/kg) of CGS-19755, an NMDA receptor inhibitor, for 7 consecutive days (discontinued for 20 h before experiment). Twelve up-regulated (CGS treated) and 12 control rats were used in this study. Under isoflurane anesthesia and after topical stimulation of the right cerebral cortex with 10⁻² M NMDA, the blood flow (¹⁴C-iodoantipyrine method) increased from 98 ± 11 ml/min/100 g in the unstimulated cortex of the control rats to 161 ± 37 ml/min/100 g in the stimulated cortex. The unstimulated value for blood flow (95 ± 7 ml/min/100 g) did not change in the upregulated group but it doubled (194 ± 69 ml/min/100 g) in the stimulated, upregulated cortex. Similarly, O₂ consumption (cryomicrospectrophotometrically determined) in normal rats increased 46%, from 9.3 ± 1 ml/min/100 g to 13.6 ± 4 after NMDA stimulation. While in the upregulated animals, O₂ consumption increased 103% from 7.9 ± 0.6 to 16 ± 6.5 after NMDA stimulation. In conclusion, NMDA receptor upregulation does not alter basal cerebrocortical blood flow or O₂ consumption but in the NMDA-stimulated cortex, the blood flow and O₂ consumption increase is dependent on the number of NMDA receptors present.     

Mild hypoxia preconditioning prevents impairment of passive avoidance learning and suppression of brain NGFI-A expression induced by severe hypoxia

The aim of this work was to study effects of mild preconditioning hypobaric hypoxia (380 Torr for 2 h, repeated 3 or 6 times spaced at 24 h) on brain NGFI-A immunoreactivity and passive avoidance (PA) behavior in rats exposed to severe hypoxia (160 Torr for 3 h). Severe hypobaric hypoxia produced extensive neuronal loss in hippocampal CA1, while the preceding hypoxic preconditioning had clear protective effect on neuronal viability of vulnerable hippocampal cells. Besides, the hypoxic preconditioning prevented impairment of acquisition and retention of PA caused by severe hypoxia. The six-trial hypobaric preconditioning was more effective in protection against PA learning deficits in severe hypoxia exposed rats than the three-trial preconditioning. The preconditioning up-regulated severe hypoxia-suppressed neocortical and hippocampal expression of NGFI-A, suggesting a possible role for NGFI-A in the neuroprotective mechanisms activated by hypoxic preconditioning.     

Correlation of local changes in cerebral blood flow,capillary density,and cytochrome oxidase during development

Although elevations in cerebral metabolic demand during development may induce angiogenesis, the correlation among ontogenic changes in local cerebral blood flow, cytochrome oxidase activity (an index of oxidative capacity) and capillary density have not been examined previously. We measured these parameters in selected regions of the brains of anesthetized rabbits of various ages. Increases in all three parameters occurred postnatally within the cerebral cortex and striatum, whereas in the medulla, values at birth were similar to those in adults. In newborns, the pattern of distribution of blood flow within the parietal cortex was such that levels were maximal in the outer layers and declined in deeper layers. This distribution correlated closely with that of capillary density, whereas cytochrome oxidase activity was maximal at levels deeper in the cortex. By postnatal day 17, the distribution for all three parameters was similar to that of cytochrome oxidase activity in young animals. A regression analysis of the regional values demonstrated a positive correlation between capillary density and blood flow in young (≤ postnatal day 8) and old (≥ postnatal day 17) animals. In contrast, cytochrome oxidase activity and capillary density were poorly correlated in young animals but positively correlated in older animals, with the slopes being markedly different (P < 0.005). The results suggest that early in postnatal development, the pattern of cytochrome oxidase activity is relatively mature compared with that of capillary density. By postnatal day 17, microvascular anatomy is closely associated to oxidative capacity, likely reflecting a steady state regulation of capillary density to metabolic requirements. © 1994 Wiley-Liss, Inc.     

Effect of nitroprusside on regional cerebral cyclic GMP, blood flow and O2 consumption in rat

This investigation was conducted to test whether topical nitroprusside (NP), a cytosolic guanylate cyclase activator, would increase the level of cyclic GMP and alter O₂ consumption or blood flow in the cerebral cortex of rats. Male Long-Evans rats were used in a control (n = 9), low dose NP (n = 13, 10⁻³ M) or high dose NP (n = 12, 10⁻² M) group. Nitroprusside or saline was topically applied to the right side of the cerebral cortex and the left side was used as a control. The cyclic GMP level was determined in five rats in each group using a radioimmunoassay. In the o ther rats in each group, regional cerebral blood flow was measured by [¹⁴C]iodoantipyrine and regional arterial and venous O₂ saturations were determined microspectrophotometrically. Nitroprusside significantly increased the cyclic GMP level from 21.4 ± 12.0 pmol/g (contralateral cortex) to 52.2 ± 36.7 pmol/g (NP treated cortex) in low dose nitroprusside group and from 19.9 ± 22.6 pmol/g (contralateral cortex) to 58.5 ± 15.1 pmol/g (NP treated cortex) in high dose nitroprusside group. High dose nitroprusside significantly increased cerebral blood flow from 80 ± 11 ml · min⁻¹ · 100 g (contralateral cortex) to 114 ± 11 ml · min⁻¹ · 100 g (NP treated cortex). However, there was no significant difference in O₂ extraction and O₂ consumption between the NP treated cortex and contralateral cortex in either the low or the high dose NP groups. In the high dose NP group, the O₂ extraction was 8.0 ± 1.3 ml O₂ · 100 ml⁻¹ in the treated cortex and 8.8 ± 1.5 ml O₂ · 100 ml⁻¹ in the contralateral cortex, while the O₂ consumptions in the NP treated cortex and contralateral cortex were 8.1 ± 1.3 ml O₂ · min⁻¹ · 100 g⁻¹ and 7.3 ± 1.2, respectively. Thus, NP increased the cyclic GMP level without a significant change in O₂ consumption in the cerebral cortex. Our data suggested that O₂ consumption in the cerebral cortex was not affected by the increased level of cyclic GMP.     

Protective effect of MgSO4 infusion on NMDA receptor binding characteristics during cerebral cortical hypoxia in the newborn piglet

This study tests the hypothesis that magnesium, a selective non-competitive antagonist of the NMDA receptor, will attenuate hypoxia-induced alteration in NMDA receptors and preserve MK-801 binding characteristics during cerebral hypoxia in vivo. Anesthetized, ventilated and instrumented newborn piglets were divided into three groups: normoxic controls were compared to untreated hypoxic and Mg²⁺-treated hypoxic piglets. Cerebral hypoxia was induced by lowering the FiO₂ to 5–7% and confirmed biochemically by a decrease in the levels of phosphocreatine (82% lower than control). The Mg²⁺-treated group received MgSO₄ 600 mg/kg over 30 min followed by 300 mg/kg administered during 60 min of hypoxia. Plasma Mg²⁺ concentrations increased from1.6 ± 0.1mg/dl to17.7 ± 3.3mg/dl.³H-MK-801 binding was used as an index of NMDA receptor modification. TheB_max in control, hypoxic and Mg²⁺-treated hypoxic piglets was1.09 ± 0.17, 0.70 ± 0.25and0.96 ± 0.14pmoles/mg protein, respectively. TheK_d for the same groups were10.02 ± 2.04, 4.88 ± 1.43and8.71 ± 2.23nM, respectively. TheB_max andK_d in the hypoxic group were significantly lower compared to the control and Mg²⁺-treated hypoxic groups, indicating a preservation of NMDA receptor number and affinity for MK-801 during hypoxia with Mg²⁺. The activity of Na⁺, K⁺ ATPase, a marker of neuronal membrane function, was lower in the hypoxic group compared to the control and Mg²⁺-treated hypoxic groups. These findings show that MgSO₄ prevents the hypoxia-induced modification of the NMDA receptor and attenuates neuronal membrane dysfunction. We suggest that the administration of Mg²⁺ prior to and during hypoxia may be neuroprotective in vivo, possibly by reducing the NMDA receptor-mediated influx of calcium.     

Corticosterone synthesis inhibitor metyrapone ameliorates chronic hypobaric hypoxia induced memory impairment in rat

Chronic exposure to hypobaric hypoxia causes oxidative stress and neurodegeneration leading to memory impairment. The present study aimed at investigating the role of corticosterone in hypoxia induced neurodegeneration and effect of metyrapone, a corticosterone synthesis inhibitor that reduces the stress induced elevation of corticosterone without affecting the basal level, in ameliorating chronic hypobaric hypoxia induced cognitive decline. Rats were exposed to simulated altitude of 25,000 ft for 0, 3, 7, 14 and 21 days to determine the temporal alterations in corticosterone and its receptors following exposure to hypobaric hypoxia. Our results showed an elevation of corticosterone in plasma and hippocampal tissue following 7 days of exposure, which declined on prolonged hypoxic exposure for 21 days. A concomitant increase in ROS and lipid peroxidation was observed along with depletion of intracellular antioxidants. Glucocorticoid and mineralocorticoid receptors were upregulated on 3 and 7 days of hypoxic exposure. Though expression of Glut1 and Glut3 were upregulated on 3 days of hypoxic exposure, sharp decline in Glut1 expression following 7 days of hypoxic exposure leads to reduced neuronal glucose uptake. Administration of metyrapone from 3rd to 7th day of hypoxic exposure to suppress hypoxia induced increase in corticosterone levels resulted in reduced oxidative damage, neurodegeneration and improvement of intracellular energy status. The metyrapone treated hypoxic animals performed better in the Morris Water Maze. Further, administration of exogenous corticosterone along with metyrapone during hypoxic exposure blunted the neuroprotective effect of metyrapone indicating a role for corticosterone in mediating hypobaric hypoxia induced neurodegeneration and memory impairment.     

Prolonged exposure to hypobaric hypoxia transiently reduces GABA(A) receptor number in mice cerebral cortex

The central nervous system is severely affected by hypoxic conditions, which produce alterations in neural cytoarchitecture and neurotransmission, resulting in a variety of neuropathological conditions such as convulsive states, neurobehavioral impairment and motor CNS alterations. Some of the neuropathologies observed in hypobaric hypoxia, corresponding to high altitude conditions, have been correlated with a loss of balance between excitatory and inhibitory neurotransmission, produced by alterations in glutamatergic and GABAergic receptors. In the present work, we have studied the effect of chronic hypobaric hypoxia (506 hPa, 18 h/day x 21 days) applied to adult male mice on GABA(A) receptors from cerebral cortex, to determine whether hypoxic exposure may irreversibly affect central inhibitory neurotransmission. Saturation curves for [3H]GABA specifically bound to GABA(A) receptors in isolated synaptic membranes showed a 30% decrease in maximal binding capacity after hypoxic exposure (Bmax control, 4.70+/-0.19, hypoxic, 3.33+/-0.10 pmol/mg protein), with no effect on GABA binding sites affinity (Kd control: 159.3+/-13.3 nM, hypoxic: 164.2+/-15.1 nM). Decreased B(max) values were observed up to the 10th post-hypoxic day, returning to control values by the 15th post-hypoxic day. Pharmacological properties of GABA(A) receptor were also affected by hypoxic exposure, with a 45 to 51% increase in the maximal effect by positive allosteric modulators (pentobarbital and 5alpha-pregnan-3alpha-ol-20-one). We conclude that long-term hypoxia produces a significant but reversible reduction on GABA binding to GABA(A) receptor sites in cerebral cortex, which may reflect an adaptive response to this sustained pathophysiological state.     

Effects of Chinese herbal monomers on oxidative phosphorylation and membrane potential in cerebral mitochondria isolated from hypoxia-exposed rats in vitro

Mitochondrial dysfunction is the key pathogenic mechanism of cerebral injury induced by high-altitude hypoxia. Some Chinese herbal monomers may exert anti-hypoxic effects through enhancing the efficiency of oxidative phosphorylation. In this study, effects of 10 kinds of Chinese herbal monomers on mitochondrial respiration and membrane potential of cerebral mitochondria isolated from hypoxia-exposed rats in vitro were investigated to screen anti-hypoxic drugs. Rats were exposed to a low-pressure environment of 405.35 mm Hg (54.04 kPa) for 3 days to establish high-altitude hypoxic models. Cerebral mitochondria were isolated and treated with different concentrations of Chinese herbal monomers (sinomenine, silymarin, glycyrrhizic acid, baicalin, quercetin, ginkgolide B, saffron, piperine, ginsenoside Rg1 and oxymatrine) for 5 minutes in vitro. Mitochondrial oxygen consumption and membrane potential were measured using a Clark oxygen electrode and the rhodamine 123 fluorescence analysis method, respectively. Hypoxic exposure significantly decreased the state 3 respiratory rate, respiratory control rate and mitochondrial membrane potential, and significantly increased the state 4 respiratory rate. Treatment with saffron, ginsenoside Rg1 and oxymatrine increased the respiratory control rate in cerebral mitochondria isolated from hypoxia-exposed rats in dose-dependent manners in vitro, while ginsenoside Rg1, piperine and oxymatrine significantly increased the mitochondrial membrane potential in cerebral mitochondria from hypoxia-exposed rats. The Chinese herbal monomers saffron, ginsenoside Rg1, piperine and oxymatrine could thus improve cerebral mitochondrial disorders in oxidative phosphorylation induced by hypobaric hypoxia exposure in vitro.     

Comparative distributions of dopamine D-1 and D-2 receptors in the cerebral cortex of rats, cats, and monkeys

The distributions and laminar densities of cerebral cortical dopamine D-1 and D-2 receptors were studied in rats, cats, and monkeys. Distributions were determined by using alternate, adjacent tissue sections processed for D-1 and D-2 receptor subtypes and compared to an adjacent, nearly adjacent, or similar sections stained for Nissl substance. [3H]-SCH 23390 and [3H]-spiroperidol (in the presence of 100 nM mianserin) were used to label the D-1 and D-2 receptors, respectively. The regional distribution and laminar density of dopamine receptors were determined by in vitro quantitative autoradiography and video densitometry of selected isocortical and peri-allocortical regions. Granular (prefrontal, primary somatosensory, and primary visual), agranular (primary motor and anterior cingulate), and limbic (entorhinal and perirhinal) cortices were examined. Where possible, homologous areas among the species were compared. The D-1 receptor was present in all regions and laminae of the cerebral cortex of rats, cats, and monkeys. The regional densities for the D-1 receptor were higher in the cat and monkey than in the rat. The rat D-1 receptor displayed a relatively homogeneous laminar pattern in most regions except that the deeper laminae (V and VI) contained more receptors than the superficial layers. The cats and monkeys, however, had distinctly heterogeneous laminar patterns in all regions of cortex that varied from one region to another and were quite different from that seen in the rat. The cats and monkeys had highest densities of the D-1 receptor in layers I and II and lowest densities in layers III and IV, whereas layers V and VI were intermediate. The density of D-1 receptors was greater than the density of D-2 receptors in all regions and laminae of cerebral cortex of the cat and monkey and greater in most regions and laminae of the rat cerebral cortex. The D-2 receptor was also distributed in all regions of the cerebral cortex of rats, cats, and monkeys. The D-2 receptor was very homogeneous in its regional distribution and laminar pattern compared to the D-1 receptor in all 3 species. The D-2 receptor was denser in the superficial layers (I and II) of the cortex than in the deeper layers in the rats, but more homogeneous in the different laminae of the cat and monkey cerebral cortex. The rat cortical D-2 receptor exceeded the D-1 receptor in restricted laminae of selective regions.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of nitric oxide synthase inhibition on high affinity Ca-ATPase during hypoxia in cerebral cortical neuronal nuclei of newborn piglets

Previous studies have shown that during hypoxia, neuronal nuclear high affinity Ca²⁺-ATPase activity is increased in the cerebral cortex of newborn piglets. The present study tests the hypothesis that pretreatment with N-nitro- -arginine (NNLA) will prevent the hypoxia-induced increase in high affinity Ca²⁺-ATPase activity in cortical neuronal nuclear membrane of newborn piglets. We also tested the hypothesis that nitration is a mechanism of elevation of the high affinity Ca²⁺-ATPase activity during hypoxia. Studies were performed in five normoxic, five hypoxic, and six NNLA-pretreated (40 mg/kg) hypoxic newborn piglets. Cerebral cortical neuronal nuclei were isolated and the high affinity Ca²⁺-ATPase activity was determined. Further, normoxic samples were aliquoted into two sub-groups for in vitro nitration with 0.5 mM peroxynitrite and subsequent determination of the high affinity Ca²⁺-ATPase activity. The activity increased from 309±40 nmol Pi/mg protein/h in the normoxic group to 520±108 nmol Pi/mg protein/h in the hypoxic group (P<0.05). In the NNLA-pretreated group, the activity was 442±53 nmol Pi/mg protein/h (P<0.05), which is 25% lower than in the hypoxic group. In the nitrated group the enzyme activity increased to 554±59 nmol Pi/mg protein/h (P<0.05). Thus peroxynitrite-induced nitration in vitro increased the high affinity Ca²⁺-ATPase activity and NNLA administration in vivo partially prevented the hypoxia-induced increase in neuronal nuclear high affinity Ca²⁺-ATPase activity. We conclude that the hypoxia-induced increase in nuclear membrane high affinity Ca²⁺-ATPase activity is NO-mediated and that nitration of the enzyme is a mechanism of its modification.     

Cerebral extracellular lactate concentration and blood flow during chemical stimulation of the nucleus tractus solitarii in anesthetized rats

The extracellular lactate concentration and blood flow in the cerebral cortex of urethane-anesthetized, paralyzed and artificially ventilated rats were monitored continuously and simultaneously using an enzyme electrode and a laser Doppler flowmeter (LDF), respectively, during chemical stimulation of the nucleus tractus solitarii (NTS) by microinjection of -glutamate (1.7 nmol 50 nl). Chemical stimulation of the NTS significantly decreased the arterial blood pressure (ABP) from 85 ± 17 to 68 ± 14 mmHg, heart rate from 418 ± 13 to 402 ± 19 beats · min⁻¹ and cerebral blood flow (CBF) by 17.9 ± 6.2% (P < 0.001). However, chemical stimulation of the NTS significantly increased the lactate concentration by 58.9 ± 17.3 μM (P < 0.001). Barostat maneuver, which held systemic ABP constant during chemical stimulation of the NTS attenuated the responses in CBF and lactate concentration by 30 and 27%, respectively. The onset of the increase in lactate concentration was delayed about 19 s after that of the CBF decrease. Circulatory lactate produced no significant change in the cerebral extracellular lactate concentration. These results indicate that chemical stimulation of the NTS induces an increase in extracellular lactate concentration in the cerebral cortex through a decrease in CBF via cerebral vasoconstriction.     

Stimulus duration-dependent contribution of Kca channel activation and cAMP to hypoxic cerebrovasodilation

Activation of calcium sensitive (K_ca) K channels and cAMP contribute to pial artery dilation observed during a 10-min exposure to hypoxia. Recent studies show that pial dilation during a 20- or 40-min hypoxic exposure was less than that observed during a 5- or 10-min exposure indicating that stimulus duration determines the nature of the vascular response to hypoxia. The present study was designed to determine if the stimulus duration modulates the contribution of K_ca channel activation and cAMP-dependent mechanisms to hypoxic pial artery dilation in piglets equipped with a closed cranial window. The K_ca channel antagonist iberiotoxin had no influence on pial dilation during 5 min of hypoxia (pO₂≈25 mmHg), decremented the dilation during 10- and 20-min exposure, but had no effect on the dilation during a 40-min exposure (33±1% vs. 32±3%, 33±1% vs. 25±1%, 23±1% vs. 19±1%, and 21±2% vs. 17±2% for 5-, 10-, 20-, and 40-min hypoxic dilations before and after iberiotoxin). NS1619, a K_ca channel agonist, induced pial dilation during hypoxia that was attenuated by 20- and 40-min but not by 5- and 10-min exposure durations. Similarly, the cAMP antagonist Rp 8-Bromo cAMPs had no influence on pial dilation during 5 min of hypoxia, decremented the dilation during a 10-min exposure, but had no effect on the dilation during a 20- or 40-min exposure (36±1% vs. 34±2%, 34±1% vs. 22±1%, 24±2% vs. 21±2%, and 21±2% vs. 19±2% for 5-, 10-, 20-, and 40-min hypoxic dilations before and after Rp 8-Bromo cAMPs). Additionally, CSF cAMP was unchanged during 5 min, elevated during 10 min, but such elevations were attenuated during 20- and 40-min hypoxic exposure. Pial vasodilation to a cAMP analogue during hypoxia was attenuated by 20- and 40-min but not by 5- and 10-min hypoxic exposure durations. These data show that K_ca channel activation and cAMP contribute to hypoxic pial artery dilation in a stimulus duration-dependent manner. These data suggest that diminished pial artery dilation during longer hypoxic exposure results from attenuated K_ca channel and cAMP-dependent mechanisms.     

Effect of hypothermia on kainic acid-induced limbic seizures: an electroencephalographic and C-deoxyglucose autoradiographic study

The effect of body temperature on kainic acid (KA)-induced limbic seizures was examined in Wistar rats. In rats undergoing limbic seizure induced by 1 μg intra-amygdaloid injection of KA, the post-injection latency of initial ictal discharges in the left amygdala was significantly longer in rats whose body temperature was lowered to 30°C (2.55±0.94 min at 37°C, 13.19±5.70 min at 30°C; p=0.0017). The post-injection latency of initial ictal discharges in the left hippocampus was also significantly longer under the same conditions (23.68±9.96 min at 37°C, 43.85±17.98 min at 30°C; p=0.0253). The number of limbic seizures occurring in the first 2 h post-injection was significantly lower in hypothermic rats (30.0±10.7 at 37°C, 8.71±2.69 at 30°C; p=0.0017), as was the total duration of limbic seizures over the same period (23.61±8.45 min at 37°C, 10.30±4.48 min at 30°C; p=0.0060). Local cerebral glucose utilization (LCGU), measured 2 h post-injection, was significantly lower in hypothermic rats, mainly in the limbic structures. ¹⁴C-deoxyglucose autoradiograms showed decreased radiation density not only in the left amygdala and bilateral hippocampus, but also in the cerebral cortex of hypothermic rats. The results of the present experiment demonstrate that the use of hypothermia, which has been shown to be effective in the treatment of acute cerebral ischemia and brain injury, may also be effective in the treatment of status epileptics.     

Lesions of the rostral ventrolateral medulla reduce the cerebrovascular response to hypoxia

Sympathoexcitatory neurons of the rostral ventrolateral medulla are tonically active and required for maintenance of resting levels of arterial pressure. They are also selectively excited by hypoxia and responsible for the associated sympathoexcitation. Since electrical or chemical stimulation of RVL will increase regional cerebral blood flow (rCBF) independently of changes in regional cerebral glucose utilization (rCGU) we investigated whether the RVL was also required to maintain resting levels of rCBF and also participated in the cerebrovascular vasodilation elicited by hypoxia. Rats were anesthetized (chloralose; 40 mg/kg, s.c.), paralyzed (tubocurarine) and ventilated (100% O₂). rCBF was measured in 10 dissected brain regions using [¹⁴C]iodoantipyrine; rCGU was measured by 2-deoxy-d-[¹⁴C]glucose. in controls (n = 6) rCBF ranged56 ± 5 in corpus callosum to101 ± 6ml/min× 100g in inferior colliculus. Hypoxic-hypoxia (PaO₂ - 36 ± 1mmHg, n = 6) increased rCBF in all structures maximally, at 204% of control, in occipital cortex. Hypercapnia (PaCO₂ = 63.5 ± 0.9, n =5) also increased rCBFP < 0.01) maximally to 199% of control in superior colliculus, Spinal cord transection with maintenance of arterial pressure did not affect resting rCBF and increased the vasodilation to hypoxia (PaO₂ = 39 ± 1mmHg, n = 5) from 2- to 3-fold in all structuresP < 0.01). Bilateral lesions within the RVL had no effect on resting rCBF or rCGU. However, they significantly reduced, in all areas by 50–69% (P < 0.01, n = 5), the cerebrovascular dilation elicited by hypoxia but not hypercapnia, Bilateral lesions in the spinal trigeminal nucleus (PaO₂ = 35 ± 1; n = 6), or transection of the IXth and Xth cranical nerves did not affect the rCBF response to hypoxia(PaO₂ = 41 ± 2; n = 6) (P > 0.05) indicating that the effect of RVL Lesions was not attributable to interference with arterial baro- or chemoreceptor reflexes. We conclude that neurons within RVL are not responsible for maintaining tonic levels of rCBF. However they contribute to the cerebrovascular vasodilation elicited by hypoxia but not hypercapnia. The cerebrovascular response to hypoxia appears reflexive and, in part, due to stimulation of oxygen-sensing neurons in RVL. In contrast, the vasodilation elicited by hypercapnia reflects local chemical signals in the cerebral microcirculation     

Delayed expression of c-fos protein in rat hippocampus and cerebral cortex following transient in vivo exposure to hypoxia

The time course of c-fos protein expression after hypoxia was examined in rat hippocampus and cerebral cortex using an immunohistochemical method. The rats were exposed to in vivo hypoxia for 30 min in a chamber containing 5% O₂ and 95% N₂. Immediately after the treatment, c-fos protein-like immunoreactivity was observed in the granule cell layer of the dentate gyrus. The change was transient, and the density of immunoreactive cells returned quickly to a control level 3 h after the exposure. However, the density of positive cells was again increased 1 day after hypoxia and reached the maximum 7 days after. In the cerebral cortex, on the other hand, no change was detected in the pattern of staining at any time, with an exception on 21 days after hypoxia. At this period, positively stained neurons were significantly increased in both density and intensity throughout the entire extent of the cerebral cortex including the cingulate gyrus. These results clearly indicate that hypoxia induces different patterns of c-fos protein expression among various regions of the brain. The biphasic pattern seen in the dentate gyrus as well as the delayed expression in the cerebral cortex may be related to delayed neuronal damages induced by hypoxia.