A Novel Approach for Studying the Physiology and Pathophysiology of Myelinated and Non-Myelinated Axons in the CNS White Matter

Advances in brain connectomics set the need for detailed knowledge of functional properties of myelinated and non-myelinated (if present) axons in specific white matter pathways. The corpus callosum (CC), a major white matter structure interconnecting brain hemispheres, is extensively used for studying CNS axonal function. Unlike another widely used CNS white matter preparation, the optic nerve where all axons are myelinated, the CC contains also a large population of non-myelinated axons, making it particularly useful for studying both types of axons. Electrophysiological studies of optic nerve use suction electrodes on nerve ends to stimulate and record compound action potentials (CAPs) that adequately represent its axonal population, whereas CC studies use microelectrodes (MEs), recording from a limited area within the CC. Here we introduce a novel robust isolated "whole" CC preparation comparable to optic nerve. Unlike ME recordings where the CC CAP peaks representing myelinated and non-myelinated axons vary broadly in size, "whole" CC CAPs show stable reproducible ratios of these two main peaks, and also reveal a third peak, suggesting a distinct group of smaller caliber non-myelinated axons. We provide detailed characterization of "whole" CC CAPs and conduction velocities of myelinated and non-myelinated axons along the rostro-caudal axis of CC body and show advantages of this preparation for comparing axonal function in wild type and dysmyelinated shiverer mice, studying the effects of temperature dependence, bath-applied drugs and ischemia modeled by oxygen-glucose deprivation. Due to the isolation from gray matter, our approach allows for studying CC axonal function without possible "contamination" by reverberating signals from gray matter. Our analysis of "whole" CC CAPs revealed higher complexity of myelinated and non-myelinated axonal populations, not noticed earlier. This preparation may have a broad range of applications as a robust model for studying myelinated and non-myelinated axons of the CNS in various experimental models.     

Effect of ischemia in vivo and oxygen-glucose deprivation in vitro on NOS pools in the spinal cord: comparative study

1. This study was performed to compare both the Ca(2+)-dependent nitric oxide synthase (NOS) activity and the neuronal nitric oxide synthase immunoreactivity (nNOS-IR) in the rabbit lumbosacral spinal cord after 15 min abdominal aorta occlusion (ischemia in vivo) and oxygen-glucose deprivation of the spinal cord slices for 45 and 60 min (ischemia in vitro). All ischemic periods were followed by 15, 30 and 60 min reoxygenation in vitro. 2. Catalytic nitric oxide synthase activity was determined by the conversion of (L)-[(14)C]arginine to (L)-[(14)C]citrulline. Neuronal nitric oxide synthase immunoreactivity in the spinal cord was detected by incubation of sections with polyclonal sheep-nNOS-primary antibody and biotinylated anti-sheep secondary antibody. 3. Our results show that ischemia in vivo and the oxygen-glucose deprivation of spinal cord slices in vitro result in a time-dependent loss of constitutive NOS activity with a partial restoration of enzyme activity during 15 and 45 min ischemia followed by 30 min of reoxygenation. A significant decrease of enzyme activity was found during 60 min ischemia alone, which persisted up to 1 h of oxygen-glucose restoration. The upregulation of neuronal nitric oxide synthase was observed in the ventral horn motoneurons after all ischemic periods. The remarkable changes in optical density of neuronal nitric oxide synthase immunoreactive motoneurons were observed after 45 and 60 min ischemia in vitro followed by 30 and 60 min reoxygenation. 4. Our results suggest that the oxygen-glucose deprivation followed by reoxygenation in the spinal cord is adequately sensitive to monitor ischemia/reperfusion changes. It seems that 15 min ischemia in vivo and 45 min ischemia in vitro cause reversible changes, while the decline of Ca(2+)-dependent nitric oxide synthase activity after 60 min ischemic insult suggests irreversible alterations.     

Metabotropic Glutamate Receptors Protect Oligodendrocytes from Acute Ischemia in the Mouse Optic Nerve

Studies by Bruce Ransom and colleagues have made a major contribution to show that white matter is susceptible to ischemia/hypoxia. White matter contains axons and the glia that support them, notably myelinating oligodendrocytes, which are highly vulnerable to ischemic-hypoxic damage. Previous studies have shown that metabotropic GluRs (mGluRs) are cytoprotective for oligodendrocyte precursor cells and immature oligodendrocytes, but their potential role in adult white matter was unresolved. Here, we report that group 1 mGluR1/5 and group 2 mGluR3 subunits are expressed in optic nerves from mice aged postnatal day (P)8–12 and P30–35. We demonstrate that activation of group 1 mGluR protects oligodendrocytes against oxygen-glucose deprivation (OGD) in developing and young adult optic nerves. In contrast, group 2 mGluR are shown to be protective for oligodendrocytes against OGD in postnatal but not young adult optic nerves. The cytoprotective effect of group 1 mGluR requires activation of PKC, whilst group 2 mGluR are dependent on negatively regulating adenylyl cyclase and cAMP. Our results identify a role for mGluR in limiting injury of oligodendrocytes in developing and young adult white matter, which may be useful for protecting oligodendrocytes in neuropathologies involving excitoxicity and ischemia/hypoxia.     

Effect of Ischemia In vivo and Oxygen-Glucose Deprivation In vitro on NOS Pools in the Spinal Cord: Comparative Study

1. This study was performed to compare both the Ca²⁺-dependent nitric oxide synthase (NOS) activity and the neuronal nitric oxide synthase immunoreactivity (nNOS-IR) in the rabbit lumbosacral spinal cord after 15 min abdominal aorta occlusion (ischemia in vivo) and oxygen-glucose deprivation of the spinal cord slices for 45 and 60 min (ischemia in vitro). All ischemic periods were followed by 15, 30 and 60 min reoxygenation in vitro.2. Catalytic nitric oxide synthase activity was determined by the conversion of _L-[¹⁴C]arginine to _L-[¹⁴C]citrulline. Neuronal nitric oxide synthase immunoreactivity in the spinal cord was detected by incubation of sections with polyclonal sheep-nNOS-primary antibody and biotinylated anti-sheep secondary antibody.3. Our results show that ischemia in vivo and the oxygen-glucose deprivation of spinal cord slices in vitro result in a time-dependent loss of constitutive NOS activity with a partial restoration of enzyme activity during 15 and 45 min ischemia followed by 30 min of reoxygenation. A significant decrease of enzyme activity was found during 60 min ischemia alone, which persisted up to 1 h of oxygen-glucose restoration. The upregulation of neuronal nitric oxide synthase was observed in the ventral horn motoneurons after all ischemic periods. The remarkable changes in optical density of neuronal nitric oxide synthase immunoreactive motoneurons were observed after 45 and 60 min ischemia in vitro followed by 30 and 60 min reoxygenation.4. Our results suggest that the oxygen-glucose deprivation followed by reoxygenation in the spinal cord is adequately sensitive to monitor ischemia/reperfusion changes. It seems that 15 min ischemia in vivo and 45 min ischemia in vitro cause reversible changes, while the decline of Ca²⁺-dependent nitric oxide synthase activity after 60 min ischemic insult suggests irreversible alterations. Abbreviations: ACSF, artificial cerebrospinal fluid; ATP, adenosine triphosphate; DAB, diaminobenzidine-tetrahydrochloride; DTT, dithiothreitol; EDTA, ethylenediaminetetraacetic acid; eNOS, endothelial nitric oxide synthase; FAD, flavin adenine dinucleotide; H₄B, tetrahydrobiopterin; iNOS, inducible nitric oxide synthase; NADPH, nicotinamide adenine dinucleotide phosphate; NMDA, N-methyl-_D-aspartate; NO, nitric oxide; NOS, nitric oxide synthase; nNOS, neuronal nitric oxide synthase; NOS-IR, nitric oxide synthase immunoreactivity; PBS, phosphate-buffered saline; PTFE, polytetrafluoroethylene     

Isolation and characterization of axolemma-enriched fractions from discrete areas of bovine CNS

Myelinated axons were isolated by flotation from bovine pons, middle cerebellar peduncle, cervical spinal cord and three regions of the subcortical white matter. The myelinated axons were osmotically and mechanically shocked, followed by fractionation on a linear 15% sucrose to 45% sucrose density gradient. Axolemma-enriched fractions (AEF) found in the 28% to 32% sucrose region of the gradient from brainstem and cord white matter had high acetylcholinesterase (AChE) while little or nil AChE activity was found in corresponding AEF derived from the subcortical white matter. Morphologically, the subcortical white matter from all regions contained a heterogeneous population of well-myelinated to thinly myelinated axons, while brainstem and cord regions contained a more homogeneous population of well-myelinated axons. Histochemical analysis of AChE localized this enzyme to axonal elements. The AEF derived from any white matter source had similar polypeptide compositions. AEF derived from subcortical white matter contained two-fold more myelin basic protein and a three-fold greater content of 2 3 cyclic nucleotide 3 phosphodiesterase (CNP) compared with AEF derived from well myelinated white matter. We conclude that the purity of the AEF is related to the degree of myelination of the white matter from which the AEF is derived. Homogeneously well myelinated white matter (pons, cerebellar peduncle, cervical spinal cord) yields the highest purity AEF, as judged by the low CNP and myelin basic protein content and highest enrichment in AChE specific activity.     

Mechanisms underlying cell death in ischemia-like damage to the rat spinal cord in vitro

New spinal cord injury (SCI) cases are frequently due to non-traumatic causes, including vascular disorders. To develop mechanism-based neuroprotective strategies for acute SCI requires full understanding of the early pathophysiological changes to prevent disability and paralysis. The aim of our study was to identify the molecular and cellular mechanisms of cell death triggered by a pathological medium (PM) mimicking ischemia in the rat spinal cord in vitro. We previously showed that extracellular Mg²⁺ (1 mM) worsened PM-induced damage and inhibited locomotor function. The present study indicated that 1 h of PM+Mg²⁺ application induced delayed pyknosis chiefly in the spinal white matter via overactivation of poly (ADP-ribose) polymerase 1 (PARP1), suggesting cell death mediated by the process of parthanatos that was largely suppressed by pharmacological block of PARP-1. Gray matter damage was less intense and concentrated in dorsal horn neurons and motoneurons that became immunoreactive for the mitochondrial apoptosis-inducing factor (the intracellular effector of parthanatos) translocated into the nucleus to induce chromatin condensation and DNA fragmentation. Immunoreactivity to TRPM ion channels believed to be involved in ischemic brain damage was also investigated. TRPM2 channel expression was enhanced 24 h later in dorsal horn and motoneurons, whereas TRPM7 channel expression concomitantly decreased. Conversely, TRPM7 expression was found earlier (3 h) in white matter cells, whereas TRPM2 remained undetectable. Simulating acute ischemic-like damage in vitro in the presence of Mg²⁺ showed how, during the first 24 h, this divalent cation unveiled differential vulnerability of white matter cells and motoneurons, with distinct changes in their TRPM expression.     

Neuroprotective effect of the stearic acid against oxidative stress via phosphatidylinositol 3-kinase pathway

Stearic acid is a long-chain saturated fatty acid consisting of 18 carbon atoms without double bonds. In the present study, we reported the neuroprotective effects and mechanism of stearic acid on cortical or hippocampal slices insulted by oxygen-glucose deprivation, NMDA or hydrogen peroxide (H(2)O(2)) in vitro. Different types of models of brain slice injury in vitro were developed by 10 min of oxygen/glucose deprivation, 0.5 mM NMDA or 2 mM H(2)O(2), respectively. After 30 min of preincubation with stearic acid (3-30 microM), cortical or hippocampal slices were subjected to oxygen-glucose deprivation, NMDA or H(2)O(2). Then the tissue activities were evaluated by using the 2,3,5-triphenyltetrazolium chloride (TTC) method. Population spikes were recorded in randomly selected hippocampal slices. Stearic acid (3-30 microM) dose-dependently protected brain slices from oxygen-glucose deprivation, NMDA and H(2)O(2) insults. Its neuroprotective effect against H(2)O(2) insults can be completely blocked by wortmannin (inhibitor of PI3K) and partially blocked by H7 (inhibitor of PKC) or genistein (inhibitor of TPK). Treatment of cortical or hippocampal slices with 30 microM stearic acid resulted in a significant increase in PI3K activity at 5, 10, 30 and 60 min. These observations reveal that stearic acid can protect cortical or hippocampal slices against injury induced by oxygen-glucose deprivation, NMDA or H(2)O(2), and its neuroprotective effects are via phosphatidylinositol 3-kinase dependent mechanism.     

White matter injury following systemic endotoxemia or asphyxia in the fetal sheep

White matter injury is the most frequently observed brain lesion in preterm infants. The etiology remains unclear, however, both cerebral hypoperfusion and intrauterine infections have been suggested as risk factors. We compared the neuropathological outcome, including the effect on oligodendrocytes, astrocytes, and microglia, following either systemic asphyxia or endotoxemia in fetal sheep at midgestation. Fetal sheep were subjected to either 25 minutes of umbilical cord occlusion or systemic endotoxemia by administration of Escherichia coli lipopolysaccharide (LPS O111:B4, 100 ng/kg, IV). Periventricular white matter lesions were observed in 2 of 6 asphyxiated fetuses, whereas the remaining animals showed diffuse injury throughout the subcortical white matter and neuronal necrosis in subcortical regions, including the striatum and hippocampus. LPS-treatment resulted in focal inflammatory infiltrates and cystic lesions in periventricular white matter in 2 of 5 animals, but with no neuron specific injury. Both experimental paradigms resulted in microglia activation in the white matter, damaged astrocytes, and loss of oligodendrocytes. These results show that the white matter at midgestation is sensitive to injury following both systemic asphyxia and endotoxemia. Asphyxia induced lesions in both white and subcortical grey matter in association with microglia activation, and endotoxemia resulted in selective white matter damage and inflammation.     

Transient glucose and amino acid deprivation induces delayed preconditioning in cultured rat cortical neurons

Several studies have demonstrated that glucose deprivation, combined either with anoxia or with the inhibition of oxidative phosphorylation, leads to the development of ischemic tolerance in neurons. The aim of our experiments was to investigate whether similar effects could be achieved by transient energy deprivation without either anoxia or the inhibition of the electron transfer chain. Preconditioning was carried out by incubating primary rat cortical neuronal cultures for 3, 6 or 9 h in a glucose- and amino acid-free balanced salt solution supplemented with B27 in normoxic conditions. After 24 h, neuronal cultures were exposed to oxygen-glucose deprivation, glutamate or hydrogen peroxide. Cell viability was measured 24 h after the lethal insults. Potential mechanisms that can influence free radical production were also examined. Energy deprivation protected neuronal cells against lethal stimuli (e.g. cell survival after oxygen-glucose deprivation was 33.1 +/- 0.52% in the untreated group and 80.1 +/- 1.27% in the 9-h energy deprivation group), reduced mitochondrial membrane potential, decreased free radical formation, attenuated the intracellular free calcium surge upon glutamate receptor stimulation, and resulted in an elevated level of GSH. Our findings show that transient energy deprivation induces delayed preconditioning and prevents oxidative injuries and neuronal cell death.     

Diversity of metabolic shift in response to oxygen deprivation in Corynebacterium glutamicum and its close relatives

Oxygen-deprived Corynebacterium glutamicum R cells remain metabolically active, producing considerable amounts of organic acids even when not actively growing. We compared the proficiencies of C. glutamicum and close relatives grown under aerobic conditions to metabolize glucose when deprived of oxygen. Eight strains that readily consumed glucose without cell growth subsequently produced organic acids. Among these, the glucose consumption rates of the two C. glutamicum strains (>40 mM/h) and Corynebacterium efficiens (>12 mM/h) were an order of magnitude higher than those of the other five strains. The resultant organic acid yields of these three strains (>86%) consequently exceeded those of the other five (<60%). This difference is probably rooted in the comparatively inferior activities of glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase, and malate dehydrogenase observed in the five strains. Moreover, under oxygen deprivation, phosphoenolpyruvate carboxylase (PEPC) activity of C. efficiens was elevated tenfold, but its lack of fumarase activity meant that no succinic acid could be produced. The metabolic shift occasioned by addition of the PEPC substrate sodium bicarbonate resulted in a doubling of the glucose consumption rate of the two C. glutamicum strains but not that of the other six close relatives.     

Pathological changes in the white matter after spinal contusion injury in the rat

It has been shown previously that after spinal cord injury, the loss of grey matter is relatively faster than loss of white matter suggesting interventions to save white matter tracts offer better therapeutic possibilities. Loss of white matter in and around the injury site is believed to be the main underlying cause for the subsequent loss of neurological functions. In this study we used a series of techniques, including estimations of the number of axons with pathology, immunohistochemistry and mapping of distribution of pathological axons, to better understand the temporal and spatial pathological events in white matter following contusion injury to the rat spinal cord. There was an initial rapid loss of axons with no detectable further loss beyond 1 week after injury. Immunoreactivity for CNPase indicated that changes to oligodendrocytes are rapid, extending to several millimetres away from injury site and preceding much of the axonal loss, giving early prediction of the final volume of white matter that survived. It seems that in juvenile rats the myelination of axons in white matter tracts continues for some time, which has an important bearing on interpretation of our, and previous, studies. The amount of myelin debris and axon pathology progressively decreased with time but could still be observed at 10 weeks after injury, especially at more distant rostral and caudal levels from the injury site. This study provides new methods to assess injuries to spinal cord and indicates that early interventions are needed for the successful sparing of white matter tracts following injury.     

Residual stress in the adult mouse brain

This work provides direct evidence that sustained tensile stress exists in white matter of the mature mouse brain. This finding has important implications for the mechanisms of brain development, as tension in neural axons has been hypothesized to drive cortical folding in the human brain. In addition, knowledge of residual stress is required to fully understand the mechanisms behind traumatic brain injury and changes in mechanical properties due to aging and disease. To estimate residual stress in the brain, we performed serial dissection experiments on 500-mum thick coronal slices from fresh adult mouse brains and developed finite element models for these experiments. Radial cuts were made either into cortical gray matter, or through the cortex and the underlying white matter tract composed of parallel neural axons. Cuts into cortical gray matter did not open, but cuts through both layers consistently opened at the point where the cut crossed the white matter. We infer that the cerebral white matter is under considerable tension in the circumferential direction in the coronal cerebral plane, parallel to most of the neural fibers, while the cerebral cortical gray matter is in compression. The models show that the observed deformation after cutting can be caused by more growth in the gray matter than in the white matter, with the estimated tensile stress in the white matter being on the order of 100–1,000 Pa.     

p38α MAP Kinase Mediates Hypoxia-Induced Motor Neuron Cell Death: A Potential Target of Minocycline’s Neuroprotective Action

Hypoxia-ischemia (HI) may play a significant role in motor neuron death associated with the pathology of spinal cord injury and, perhaps, amyotrophic lateral sclerosis. The present study employs an in vitro model of HI to investigate the role of a stress kinase pathway, i.e., p38 MAP kinase, in cell death signaling in a motor neuron cell line, i.e., NSC34, subjected to oxygen-glucose deprivation (OGD). Although the neurons were essentially tolerant to either hypoxia (0.2% O₂) or low glucose (1 mM) alone, more than 60% of them died in response to combined low oxygen and low-glucose exposure. Minocycline, a semi-synthetic tetracycline known for its neuroprotective effects in models of neurodegeneration, afforded substantial (∼50%) protection against hypoxic cell death, assessed by lactate dehydrogenase release and flow cytometry, while suppressing OGD-induced p38 MAP kinase activation. An inhibitor of p38 kinase, SB203580, as well as siRNA-mediated down-regulation of p38 kinase elicited an almost complete blockade of OGD-induced cell death. The use of p38 isoform-specific siRNAs further revealed preferential involvement of the α over the β isoform of p38 MAP kinase in hypoxic neuronal cell death in our model.     

Changes within maturing neurons limit axonal regeneration in the developing spinal cord

Embryonic birds and mammals display a remarkable ability to regenerate axons after spinal injury, but then lose this ability during a discrete developmental transition. To explain this transition, previous research has emphasized the emergence of myelin and other inhibitory factors in the environment of the spinal cord. However, research in other CNS tracts suggests an important role for neuron-intrinsic limitations to axon regeneration. Here we re-examine this issue quantitatively in the hindbrain-spinal projection of the embryonic chick. Using heterochronic cocultures we show that maturation of the spinal cord environment causes a 55% reduction in axon regeneration, while maturation of hindbrain neurons causes a 90% reduction. We further show that young neurons transplanted in vivo into older spinal cord can regenerate axons into myelinated white matter, while older axons regenerate poorly and have reduced growth cone motility on a variety of growth-permissive ligands in vitro, including laminin, L1, and N-cadherin. Finally, we use video analysis of living growth cones to directly document an age-dependent decline in the motility of brainstem axons. These data show that developmental changes in both the spinal cord environment and in brainstem neurons can reduce regeneration, but that the effect of the environment is only partial, while changes in neurons by themselves cause a nearly complete reduction in regeneration. We conclude that maturational events within neurons are a primary cause for the failure of axon regeneration in the spinal cord.     

Pharmacological inhibition of AMP-activated protein kinase provides neuroprotection in stroke

The restoration of energy balance during ischemia is critical to cellular survival; however, relatively little is known concerning the regulation of neuronal metabolic pathways in response to central nervous system ischemia. AMP-activated protein kinase (AMPK), a master sensor of energy balance in peripheral tissues, is phosphorylated and activated when energy balance is low. We investigated whether AMPK might also modulate neuronal energy homeostasis during ischemia. We utilized two model systems of ischemia, middle cerebral artery occlusion in vivo and oxygen-glucose deprivation in vitro, to delineate changes in AMPK activity incurred from a metabolic stress. AMPK is highly expressed in cortical and hippocampal neurons under both normal and ischemic conditions. AMPK activity, as assessed by phosphorylation status, is increased following both middle cerebral artery occlusion and oxygen-glucose deprivation. Pharmacological inhibition of AMPK by either C75, a known modulator of neuronal ATP levels, or compound C reduced stroke damage. In contrast, activation of AMPK by 5-aminoimidazole-4-carboxamide ribonucleoside exacerbated damage. Mice deficient in neuronal nitric-oxide synthase demonstrated a decrease in both stroke damage and AMPK activation compared with wild type, suggesting a possible interaction between NO and AMPK activation in stroke. These data demonstrate a role for AMPK in the response of neurons during metabolic stress and suggest that in ischemia the activation of AMPK is deleterious. The ability to manipulate pharmacologically neuronal energy balance during ischemia represents an innovative approach to neuroprotection.     

Human umbilical cord blood cells protect oligodendrocytes from brain ischemia through Akt signal transduction

Human umbilical cord blood (HUCB) cells protect the brain against ischemic injury, yet the mechanism of protection remains unclear. Using both in vitro and in vivo paradigms, this study examined the role of Akt signaling and peroxiredoxin 4 expression in human umbilical cord blood cell-mediated protection of oligodendrocytes from ischemic conditions. As previously reported, the addition of HUCB cells to oligodendrocyte cultures prior to oxygen glucose deprivation significantly enhanced oligodendrocyte survival. The presence of human umbilical cord blood cells also increased Akt phosphorylation and elevated peroxiredoxin 4 expression in oligodendrocytes. Blocking either Akt or peroxiredoxin 4 activity with Akt Inhibitor IV or a peroxiredoxin 4-neutralizing antibody, respectively, negated the protective effects of human umbilical cord blood cells. In vivo, systemic administration of human umbilical cord blood cells 48 h after middle cerebral artery occlusion increased Akt phosphorylation and peroxiredoxin 4 protein expression while reducing proteolytic cleavage of caspase 3 in oligodendrocytes residing in the ipsilateral external capsule. Moreover, human umbilical cord blood cells protected striatal white matter bundles from degeneration following middle cerebral artery occlusion. These results suggest that the soluble factors released from human umbilical cord blood cells converge on Akt to elevate peroxiredoxin 4 levels, and these effects contribute to oligodendrocyte survival.     

Role of osmotic stress in ion accumulation and physiological responses of mycorrhizal white spruce (<Emphasis Type="Italic">Picea glauca</Emphasis>) and jack pine (<Emphasis Type="Italic">Pinus banksiana</Emphasis>) to soil fluoride and NaCl

To examine the mechanisms of earlier reported alleviation of fluoride injury in ectomycorrhizal plants by NaCl, jack pine (Pinus banksiana) and white spruce (Picea glauca) seedlings were subjected to 1 mM and 5 mM KF in the presence of either 60 mM NaCl or 10% polyethylene glycol 3350 (PEG) for 2 weeks. Before the treatments, seedlings had either been inoculated with the ectomycorrhizal fungus Suillus tomentosus or remained non-inoculated. The inoculation with S. tomentosus reduced Na uptake by shoots and roots of jack pine seedling and by roots of white spruce that were treated with 60 mM NaCl. Mycorrhizal associations also drastically decreased fluoride uptake by jack pine seedlings, but did not affect shoot fluoride concentrations in white spruce. When NaCl was replaced by PEG in the 5 mM KF treatment solution, shoot fluoride concentrations were reduced by more than twofold without corresponding reductions in transpiration rates in mycorrhizal and non-mycorrhizal white spruce seedlings. When fluoride was present in the treatment solution, Na concentrations were lower in shoots and roots of both jack pine and white spruce mycorrhizal and non-mycorrhizal seedlings. The results suggest that Suillus tomentosus may help alleviate the effects of soil fluoride and salinity in jack pine and that fluoride uptake in white spruce is sensitive to osmotic stress.     

Synthetic Oligodeoxynucleotides Containing Multiple Telemeric TTAGGG Motifs Suppress Inflammasome Activity in Macrophages Subjected to Oxygen and Glucose Deprivation and Reduce Ischemic Brain Injury in Stroke-Prone Spontaneously Hypertensive Rats

The immune system plays a fundamental role in both the development and pathobiology of stroke. Inflammasomes are multiprotein complexes that have come to be recognized as critical players in the inflammation that ultimately contributes to stroke severity. Inflammasomes recognize microbial and host-derived danger signals and activate caspase-1, which in turn controls the production of the pro-inflammatory cytokine IL-1β. We have shown that A151, a synthetic oligodeoxynucleotide containing multiple telemeric TTAGGG motifs, reduces IL-1β production by activated bone marrow derived macrophages that have been subjected to oxygen-glucose deprivation and LPS stimulation. Further, we demonstrate that A151 reduces the maturation of caspase-1 and IL-1β, the levels of both the iNOS and NLRP3 proteins, and the depolarization of mitochondrial membrane potential within such cells. In addition, we have demonstrated that A151 reduces ischemic brain damage and NLRP3 mRNA levels in SHR-SP rats that have undergone permanent middle cerebral artery occlusion. These findings clearly suggest that the modulation of inflammasome activity via A151 may contribute to a reduction in pro-inflammatory cytokine production by macrophages subjected to conditions that model brain ischemia and modulate ischemic brain damage in an animal model of stroke. Therefore, modulation of ischemic pathobiology by A151 may have a role in the development of novel stroke prevention and therapeutic strategies.     

Bisphenol A inhibits compound action potentials in the frog sciatic nerve in a manner independent of estrogen receptors

Although the endocrine disruptor bisphenol A (BPA) is reported to inhibit nerve conduction, the underlying mechanisms are unclear. Therefore, in the present study, we examined the effect of BPA on compound action potentials (CAPs) recorded from the frog sciatic nerve using the air-gap method. Treatment of the sciatic nerve with BPA (0.5 mM) for 20 min reduced the peak amplitude of the CAP by approximately 60% in a partially reversible manner. The reduction in the CAP peak amplitude was concentration-dependent, with a half-maximal inhibitory concentration (IC₅₀) value of 0.31 mM. This effect of BPA was unaffected by an estrogen-receptor antagonist, 4-hydroxytamoxifen, which by itself reduced CAP peak amplitude, with an IC₅₀ value of 0.26 mM (comparable to that of BPA). The natural estrogen 17β-estradiol, at the highest dissolvable concentration (0.05 mM), had an effect similar to that of BPA. The IC₅₀ value of BPA was comparable to those of some local anesthetics in inhibiting frog CAPs. Our findings suggest that BPA inhibits nerve conduction in a manner independent of estrogen receptors. This action of BPA may underlie, at least in part, the neurotoxicity of the compound.     

A potential protective effect of α-tocopherol on vascular complication in spinal cord reperfusion injury in rats

Background  

Paraplegia remains a potential complication of spinal cord ischemic reperfusion injury (IRI) in which oxidative stress induced cyclooxygenase activities may contribute to ischemic neuronal damage. Prolonged administration of vitamin E (α-TOL), as a potent biological antioxidant, may have a protective role in this oxidative inflammatory ischemic cascade to reduce the incidence of paraplegia. The present study was designed to evaluate the preventive value of α-TOL in IRI of spinal cord.