Poly(ADP-ribose) polymerase inhibitors attenuate necrotic but not apoptotic neuronal death in experimental models of cerebral ischemia

An excessive activation of poly(ADP-ribose) polymerase (PARP) has been proposed to play a key role in post-ischemic neuronal death. We examined the neuroprotective effects of the PARP inhibitors benzamide, 6(5H)-phenanthridinone, and 3,4-dihydro-5-[4-1(1-piperidinyl)buthoxy]-1(2H)-isoquinolinone in three rodent models of cerebral ischemia. Increasing concentrations of the three PARP inhibitors attenuated neuronal injury induced by 60 min oxygen-glucose deprivation (OGD) in mixed cortical cell cultures, but were unable to reduce CA1 pyramidal cell loss in organotypic hippocampal slices exposed to 30 min OGD or in gerbils following 5 min bilateral carotid occlusion. We then examined the necrotic and apoptotic features of OGD-induced neurodegeneration in cortical cells and hippocampal slices using biochemical and morphological approaches. Cortical cells exposed to OGD released lactate dehydrogenase into the medium and displayed ultrastructural features of necrotic cell death, whereas no caspase-3 activation nor morphological characteristics of apoptosis were observed at any time point after OGD. In contrast, a marked increase in caspase-3 activity was observed in organotypic hippocampal slices after OGD, together with fluorescence and electron microscope evidence of apoptotic neuronal death in the CA1 subregion. Moreover, the caspase inhibitor Z-VAD-FMK reduced OGD-induced CA1 pyramidal cell loss. These findings suggest that PARP overactivation may be an important mechanism leading to post-ischemic neurodegeneration of the necrotic but not of the apoptotic type.     

Perineuronal Nets Characterized by Vital Labelling, Confocal and Electron Microscopy in Organotypic Slice Cultures of Rat Parietal Cortex and Hippocampus

Perineuronal nets (PNs) of the extracellular matrix have been shown to develop in organotypic slice cultures largely corresponding with regional patterns known from in vivo experiments. In the present study, we use vital labelling to investigate aspects of the cell type-dependent development of PNs associated with nonpyramidal neurons and pyramidal cells in the parietal cortex and hippocampus. Frontal sections were cut from brains of 3-5-day-old rats and were cultured for 3-5 weeks. PNs were sequentially labelled using biotinylated Wisteria floribunda agglutinin and chromogen-tagged streptavidin either in living slice cultures, examined by confocal microscopy in vitro, or in cultures examined by confocal and electron microscopy after fixation. Nonpyramidal and pyramidal cells were characterized by immunoreaction for parvalbumin and the ionotropic glutamate receptor subunits 2/3. Vital labelling and examination of fixed slices correspondingly revealed that large numbers of PNs developed around cortical and hippocampal interneurons under depolarizing conditions induced by elevated external potassium concentration. After culture in standard medium, PNs were mainly found in association with subpopulations of pyramidal cells in the parietal cortex. PNs showed ultrastructural characteristics resembling those known from perfusion-fixed brain. A zone of labelled extracellular matrix aggregates was found in close proximity to the neuronal cell surface, surrounding presynaptic boutons and preterminal axons. The results show that characteristic features of PNs are retained after vital labelling in slice cultures. Moreover, our findings suggest that the cell type-specific development of PNs is regulated by patterns of intrinsic activity mediated by intra-cortical and -hippocampal synaptic contacts on potentially net-associated neurons.     

海马脑片盲法膜片钳全细胞记录技术

本文较为详细地介绍了海马脑片盲法膜片钳全细胞记录技术,对其关键步骤和需要注意的问题进行了重点说明,同时对CA1区锥体神经元突触活动的特点,电压门控性Ca^2 通道以及谷氨酸（glutamate,Glu)γ－氨基丁酸（GABA）受体通道电流性质等进行了观察和分析,实验结果为采用海马脑片盲法膜片钳全细胞记录技术研究海马神经元离子通道动力学性质和中枢神经系统药物对突触活动的影响提供了可靠的依据。     

REGIONAL TRANSPORT OF TRYPTOPHAN IN RAT BRAIN

Abstract— Tryptophan uptake was studied in brain slices and synaptosomes prepared from regions known to vary in the numbers of serotoninergic cell bodies and nerve endings that they contain. The rate of tryptophan uptake was highest in hypothalamus for both types of preparation. Differences among the regions were much more pronounced in isolated nerve endings (synaptosomes). Loading with tryptophan did not affect the uptake into tissue slices. Tryptophan accumulation in hypothalamus synaptosomes was reduced after intraventricular injection of 5,7–dihydroxytryptamine whereas no change was observed in synaptosomes prepared from cerebellum under the same conditions; accumulation by synaptosomes prepared from the hypothalamic and hippocampal regions was reduced after raphe lesions.     

Indomethacin enhances learning and memory potential by interacting with CaMKII

The present study examined the effect of indomethacin (IM), a cyclooxygenase inhibitor, on learning and memory functions. IM activated Ca(2+) /calmodulin-dependent protein kinase II (CaMKII) in cultured rat hippocampal neurons. IM (100 μM) significantly increased the rate of spontaneous AMPA receptor-mediated miniature excitatory postsynaptic currents elicited from CA1 pyramidal neurons of rat hippocampal slices, without affecting the amplitude, and enhanced extracellular high K(+) (20 mM)-induced glutamate release from rat hippocampal slices, indicating that IM stimulates presynaptic glutamate release. Those IM effects were clearly inhibited by the CaMKII inhibitor KN-93. IM persistently facilitated synaptic transmission monitored from the CA1 region of rat hippocampal slices in a concentration (1-100 μM)-dependent manner that was also abolished by KN-93. In the water maze test, IM (1 mg/kg, i.p.) enhanced spatial learning and memory ability for normal rats, and ameliorated scopolamine-induced spatial learning and memory impairment or age-related spatial learning and memory deterioration for senescence-accelerated mouse-prone 8 mice. In the test to learn 15 numbers consisting of three patterns of five digit number for healthy human subjects, oral intake with IM (25 mg/kg) significantly raised the scores of correct number arrangements that subjects memorized 5 min and 3 days after the test. The results of the present study indicate that IM could enhance learning and memory potential by facilitating hippocampal synaptic transmission as a result from stimulating presynaptic glutamate release under the control of CaMKII.     

Apoptosis inhibition in ischemic brain by intraperitoneal PTD-BIR3-RING (XIAP)

Anti-apoptotic treatment is a promising strategy for neuroprotection against various brain injuries resulting from ischemia or neuron degeneration. X-linked inhibitor of apoptosis protein (XIAP) is regarded as the most effective apoptosis inhibitor, in which C-terminal structure BIR3-RING mainly inhibits caspase-9-dependent apoptosis. In the present study, we fused XIAP (BIR3-RING) to the protein transduction domain (PTD) of antennapedia homeodomain of Drosophila (Antp HD), and then used the oxygen glucose deprivation (OGD)-induced hippocampal slices injury in vitro, and the rat transient middle cerebral artery ischemia (tMCAO) models in vivo, to explore the anti-apoptotic effect of this recombinant protein. The results showed that the PTD could efficiently mediate the transduction of BIR3-RING into the hippocampal slices and rat brains. PTD-BIR3-RING could decrease OGD-induced cell death in brain slices (p < 0.05). Intraperitoneal injection of PTD-BIR3-RING could attenuate terminal deoynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) positive cells and decrease cleaved caspase-3 in the ischemic bounder zone compared with the control animals (p < 0.05). Further studies showed that ischemia-induced neurological outcomes were improved in rats with PTD-BIR3-RING treatment (p < 0.05). These results demonstrate that PTD-BIR3-RING could attenuate cell death in OGD hippocampal slices and decrease cell apoptosis in tMCAO brains through inhibiting of caspase-3 cleavage, suggesting that PTD-mediated protein transduction provides a novel and effective approach for the therapies of brain diseases such as cerebral ischemia.     

Evidence that dendritic cells infiltrate atherosclerotic lesions in apolipoprotein E-deficient mice

Earlier we reported that atherosclerotic lesions of apoE-deficient mice contained cells which stained positively with anti-S-100 antibody and that cells exhibiting the ultrastructural features of dendritic cells were present in the aortic lesions. These observations suggested that dendritic cells might be involved in mouse atherosclerosis. By employing DEC-205 and MIDC-8 antibodies specific for dendritic cells, the present study has established that dendritic cells indeed accumulate in atherosclerotic lesions of apoE-deficient mice. Finding dendritic cells infiltrating atherosclerotic lesions in apoE-deficient mice offers the possibility of investigating the migratory routes of dendritic cells and their involvement in T-cell activation.     

Plasmin potentiates synaptic N-methyl-D-aspartate receptor function in hippocampal neurons through activation of protease-activated receptor-1

Protease-activated receptor-1 (PAR1) is activated by a number of serine proteases, including plasmin. Both PAR1 and plasminogen, the precursor of plasmin, are expressed in the central nervous system. In this study we examined the effects of plasmin in astrocyte and neuronal cultures as well as in hippocampal slices. We find that plasmin evokes an increase in both phosphoinositide hydrolysis (EC(50) 64 nm) and Fura-2/AM fluorescence (195 +/- 6.7% above base line, EC(50) 65 nm) in cortical cultured murine astrocytes. Plasmin also activates extracellular signal-regulated kinase (ERK1/2) within cultured astrocytes. The plasmin-induced rise in intracellular Ca(2+) concentration ([Ca(2+)](i)) and the increase in phospho-ERK1/2 levels were diminished in PAR1(-/-) astrocytes and were blocked by 1 microm BMS-200261, a selective PAR1 antagonist. However, plasmin had no detectable effect on ERK1/2 or [Ca(2+)](i) signaling in primary cultured hippocampal neurons or in CA1 pyramidal cells in hippocampal slices. Plasmin (100-200 nm) application potentiated the N-methyl-D-aspartate (NMDA) receptor-dependent component of miniature excitatory postsynaptic currents recorded from CA1 pyramidal neurons but had no effect on alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate- or gamma-aminobutyric acid receptor-mediated synaptic currents. Plasmin also increased NMDA-induced whole cell receptor currents recorded from CA1 pyramidal cells (2.5 +/- 0.3-fold potentiation over control). This effect was blocked by BMS-200261 (1 microm; 1.02 +/- 0.09-fold potentiation over control). These data suggest that plasmin may serve as an endogenous PAR1 activator that can increase [Ca(2+)](i) in astrocytes and potentiate NMDA receptor synaptic currents in CA1 pyramidal neurons.     

Glutamine from Glial Cells Is Essential for the Maintenance of the Nerve Terminal Pool of Glutamate: Immunogold Evidence from Hippocampal Slice Cultures

Abstract: The immunogold labeling for glutamate and glutamine was studied at the electron microscopic level in hippocampal slice cultures following inhibition of l -glutamine synthetase [ l -glutamate:ammonia ligase (ADP-forming); EC 6.3.1.2]. In control cultures, glutamate-like immunoreactivity was highest in terminals, intermediate in pyramidal cell bodies, and low in glial cells. Glutamine-like immunoreactivity was high in glial cells, intermediate in pyramidal cell bodies, and low in terminals. After inhibition of glutamine synthetase with l -methionine sulfoximine, glutamate-like immunoreactivity was reduced by 52% in terminals and increased nearly fourfold in glia. Glutamine-like immunoreactivity was reduced by 66% in glia following l -methionine sulfoximine, but changed little in other compartments. In cultures that were treated with both l -methionine sulfoximine and glutamine (1.0 m M ), glutamate-like immunoreactivity was maintained at control levels in terminals, whereas in glia glutamate-like immunoreactivity was increased and glutamine-like immunoreactivity was decreased to a similar extent as in cultures treated with l -methionine sulfoximine alone. We conclude that (a) glutamate accumulates in glia when the flux through glutamine synthetase is blocked, emphasizing the importance of this pathway for the handling of glutamate; and (b) glutamine is necessary for the maintenance of a normal level of glutamate in terminals, and neither reuptake nor de novo synthesis through pathways other than the glutaminase reaction is sufficient.     

Role of the adenylate cyclase system in cholinergic modulation of synaptic transmission in the hippocampus

Involvement of the adenylate cyclase system in cholinergic modulation of synaptic transmission was investigated in area CA1 in rat hippocampal slices. Microiontophoretic application of acetylcholine as well as addition of carbachol to the superfusate or of tolbutamide (a cAMP-dependent protein kinase inhibitor) depressed transmission at synapses formed by Schaffer collaterals and commissural fibers with dendrites of pyramidal cells belonging to hippocampal area CA1. Both numbers of free quanta of neurotransmitter and the likelihood of transmitter release decreased following carbachol action. Atropine suppressed the inhibitory action of carbachol on synaptic transmission. Dibutyryl cAMP and forskolin increased the amplitude of synaptic potentials and suppressed, either partially or in full, the inhibitory effects of cholinomimetics on synaptic potentials. It was concluded that cholinomimetics and activators of the adenylate cyclase system exert opposing effects on neurotransmission at synapses formed between Schaffer collaterals/commissural fibers and dendrites of pyramidal neurons belonging to hippocampal area CA1.Institute of Biophysics, Academy of Sciences of the USSR, Pushchino. Translated from Neirofiziologiya, Vol. 21, No. 4, pp. 435–442, July–August, 1989.     

Effects of interleukin-10 on the epileptiform activity development in the hippocampus induced by brief hypoxia, bicuculline and electrical kindling

The comparative effects of antiinflammatory cytokine interleukin-10 on the epileptiform activity development in CA1 hippocampal neurons were studied in different functional models of epileptogenesis that are not accompanied the visible morphological disturbances in the brain cells: --in vitro hypoxic model in the rat hippocampal slices; 2--in vitro disinhibitory model with using GABAA antagonist, bicuculline, in the rat hippocampal slices; 3--partial hippocampal kindling model in freely moving rats. Interleukin-10 (1 ng/ml) depressed the posthypoxic hyperexcitability in CA1 pyramidal neurons of the rat hippocampal slices through a decrease of the effectiveness of hypoxia to depresses the functional neuronal activity in the rat hippocampal slices during hypoxic episode. On the other hand, interleukin-10 (1 ng/ml) did not affect an initiation of epileptiform activity in CA1 pyramidal neurons of the rat hippocampal slices induced by bicuculline. Interleukin-10 (1 ng/5 microl) applied to the dorsal hippocampus in awake rats depressed an initiation of focal seizures ("ictal"-like components of afterdischarges) induced by hippocampal kindling during the first six hours after an application. However, this cytokine did not affect neither the duration of "interictal"-like component of afterdischarges nor motor seizure development. Thus, our findings showed that antiinflammatory cytokine interleukin-10, in addition to its antihypoxic action, exert the neuroprotective effect on the initiation of "ictal"-like, but not "interictal"-like, epileptiform discharges.     

New roles for the gamma rhythm: population tuning and preprocessing for the Beta rhythm

Gamma (30–80 Hz) and beta (12–30 Hz) oscillations such as those displayed by in vitro hippocampal (CA1) slice preparations and by in vivo neocortical EEGs often occur successively, with a spontaneous transition between them. In the gamma rhythm, pyramidal cells fire together with the interneurons, while in the beta rhythm, pyramidal cells fire on a subset of cycles of the interneurons. It is shown that gamma and beta rhythms have different properties with respect to creation of cell assemblies. In the presence of heterogeneous inputs to the pyramidal cells, the gamma rhythm creates an assembly of firing pyramidal cells from cells whose drive exceeds a threshold. During the gamma to beta transition, a slow outward potassium current is activated, and as a result the cell assembly vanishes. The slow currents make each of the pyramidal cells fire with a beta rhythm, but the field potential of the network still displays a gamma rhythm. Hebbian changes of connections among the pyramidal cells give rise to a beta rhythm, and the cell assemblies are recovered with a temporal separation between cells firing in different cycles. We present experimental evidence showing that such a separation can occur in hippocampal slices.     

Cellular Targets of Nitric Oxide in the Hippocampus

In the hippocampus, as in many other CNS areas, nitric oxide (NO) participates in synaptic plasticity, manifested as changes in pre- and/or postsynaptic function. While it is known that these changes are brought about by cGMP following activation of guanylyl cyclase-coupled NO receptors attempts to locate cGMP by immunocytochemistry in hippocampal slices in response to NO have failed to detect the cGMP elevation where expected, i.e. in the pyramidal neurones. Instead, astrocytes, unidentified varicose fibres and GABA-ergic nerve terminals are reported to be the prominent NO targets, raising the possibility that NO acts indirectly via other cells. We have re-investigated the distribution of cGMP generated in response to endogenous and exogenous NO in hippocampal slices using immunohistochemistry and new conditions designed to optimise cGMP accumulation and, hence, its detectability. The conditions included use of tissue from the developing rat hippocampus, a potent inhibitor of phosphodiesterase-2, and an allosteric enhancer of the NO-receptive guanylyl cyclase. Under these conditions, cGMP was formed in response to endogenous NO and was found in a population of pyramidal cell somata in area CA3 and subiculum as well as in structures described previously. The additional presence of exogenous NO resulted in hippocampal cGMP reaching the highest level recorded for brain tissue (1700 pmol/mg protein) and in cGMP immunolabelling throughout the pyramidal cell layer. Populations of axons and interneurones were also stained. According with these results, immunohistochemistry for the common NO receptor β1-subunit indicated widespread expression. A similar staining pattern for the α1-subunit with an antibody used previously in the hippocampus and elsewhere, however, proved to be artefactual. The results indicate that the targets of NO in the hippocampus are more varied and extensive than previous evidence had suggested and, in particular, that the pyramidal neurones participating in NO-dependent synaptic plasticity are direct NO targets.     

Tributyltin induces oxidative stress and neuronal injury by inhibiting glutathione S-transferase in rat organotypic hippocampal slice cultures

Tributyltin (TBT) has been used as a heat stabilizer, agricultural pesticide and antifouling agents on ships, boats and fish-farming nets; however, the neurotoxicity of TBT has recently become a concern. TBT is suggested to stimulate the generation of reactive oxygen species (ROS) inside cells. The aim of this study was to determine the mechanism of neuronal oxidative injury induced by TBT using rat organotypic hippocampal slice cultures. The treatment of rat hippocampal slices with TBT induced ROS production, lipid peroxidation and cell death. Pretreatment with antioxidants such as superoxide dismutase, catalase or trolox, suppressed the above phenomena induced by TBT, indicating that TBT elicits oxidative stress in hippocampal slices, which causes neuronal cell death. TBT dose-dependently inhibited glutathione S-transferase (GST), but not glutathione peroxidase or glutathione reductase in the cytosol of rat hippocampus. The treatment of hippocampal slices with TBT decreased the GST activity. Pretreatment with reduced glutathione attenuated the reduction of GST activity and cell death induced by TBT, indicating that the decrease in GST activity by TBT is involved in hippocampal cell death. When hippocampal slices were treated with sulforaphane, the expression and activity of GST were increased. Notably, TBT-induced oxidative stress and cell death were significantly suppressed by pretreatment with sulforaphane. These results indicate that GST inhibition could contribute, at least in part, to the neuronal cell death induced by TBT in hippocampal slices. This study is the first report to show the link between neuronal oxidative injury and the GST inhibition elicited by TBT.     

Alcohol is a potent stimulant of immature neuronal networks: implications for fetal alcohol spectrum disorder

Ethanol consumption during development affects the maturation of hippocampal circuits by mechanisms that are not fully understood. Ethanol acts as a depressant in the mature CNS and it has been assumed that this also applies to immature neurons. We investigated whether ethanol targets the neuronal network activity that is involved in the refinement of developing hippocampal synapses. This activity appears during the growth spurt period in the form of giant depolarizing potentials (GDPs). GDPs are generated by the excitatory actions of GABA and glutamate via a positive feedback circuit involving pyramidal neurons and interneurons. We found that ethanol potently increases GDP frequency in the CA3 hippocampal region of slices from neonatal rats. It also increased the frequency of GDP-driven Ca2+ transients in pyramidal neurons and increased the frequency of GABA(A) receptor-mediated spontaneous postsynaptic currents in CA3 pyramidal cells and interneurons. The ethanol-induced potentiation of GABAergic activity is probably the result of increased quantal GABA release at interneuronal synapses but not enhanced neuronal excitability. These findings demonstrate that ethanol is a potent stimulant of developing neuronal circuits, which might contribute to the abnormal hippocampal development associated with fetal alcohol syndrome and alcohol-related neurodevelopmental disorders.     

The effects of long-term exposure of magnetic field via 900-MHz GSM radiation on some biochemical parameters and brain histology in rats

The aim of this study is to determine the effects of magnetic field via cell phones on some blood parameters and neurons in the brain of rats. Animals have been classified into three groups: control, Magnetic Field (MF), and F2 groups. Throughout this study, cell phones were placed on the wall of the cages. Rats were exposed to the effects of cell phones during prenatal and postnatal periods until they were 80 days old. During the study, the exposure procedure of rats was that the phone was in standby mode for a whole day and in talking mode for 30 min per day. The waves of cell phones caused an increased blood glucose level from 96.52 ± 5.64 mg/dl to 132.14 ± 5.93 mg/dl and an increased serum protein level from 131.14 ± 6.19 mg/dl to 319.29 ± 6.73 mg/dl compared to control. Statistically, significant differences wasn't observed in the blood cholesterol concentration between the groups compared to the control. Weekly weight gain decreased in all groups compared to the control. MF exposure decreased pyramidal neuron numbers 51.15% and increased ischemic neuron numbers 73% at cortex region of brain. In addition, vascular dilatations have increased clearly in group F2.Whereas the procedure of MF did not have any effects on hippocampal pyramidal cell numbers, magnetic fields increased the amount of ischemic neurons three-fold compared to the control. In conclusion, MF affected some biochemical parameters, especially the cortex region of the brain.     

Imaging synaptic inhibition in transgenic mice expressing the chloride indicator, Clomeleon

We describe here a molecular genetic approach for imaging synaptic inhibition. The thy-1 promoter was used to express high levels of Clomeleon, a ratiometric fluorescent indicator for chloride ions, in discrete populations of neurons in the brains of transgenic mice. Clomeleon was functional after chronic expression and provided non-invasive readouts of intracellular chloride concentration ([Cl(-)](i)) in brain slices, allowing us to quantify age-dependent declines in resting [Cl(-)](i) during neuronal development. Activation of hippocampal interneurons caused [Cl(-)](i) to rise transiently in individual postsynaptic pyramidal neurons. [Cl(-)](i) increased in direct proportion to the amount of inhibitory transmission, with peak changes as large as 4 mM. Integrating responses over populations of pyramidal neurons allowed sensitive detection of synaptic inhibition. Thus, Clomeleon imaging permits non-invasive, spatiotemporally resolved recordings of [Cl(-)](i) in a large variety of neurons, opening up new opportunities for imaging synaptic inhibition and other forms of chloride signaling.     

Leptin receptor long-form splice-variant protein expression in neuron cell bodies of the brain and co-localization with neuropeptide Y mRNA in the arcuate nucleus.

Reduced leptin (Ob protein) signaling is proposed to be a stimulus for the activation of neuropeptide Y (NPY) gene activity and increased expression of mRNA for the long form of the leptin receptor (Ob-Rb) in the hypothalamic arcuate nucleus. To determine if Ob-Rb protein is expressed in arcuate nucleus NPY neurons, we developed an affinity-purified polyclonal antibody against amino acids 956-1102 of human Ob-Rb. This antibody specifically recognizes the cytoplasmic tail of Ob-Rb and does not react with shorter leptin-receptor variants. Western immunoblots of Ob-Rb-transfected COS cells showed a single 150-kD band, and immunofluorescence revealed intense perinuclear staining in the cytoplasm. A 150-kD band was also present in Western immunoblots of hypothalamus. Immunocytochemical staining of brain slices revealed immunoreactive Ob-Rb protein concentrated in many neuronal cell bodies in the same regions of the forebrain that also express Ob-Rb mRNA. In the hypothalamus, Ob-Rb-positive cell bodies were abundant in the arcuate nucleus and ventromedial nucleus, with lesser numbers in the dorsomedial nucleus and paraventricular nucleus. Immunostaining was also detected in cell bodies of pyramidal cell neurons of the pyriform cortex and cerebral cortex, in neurons of the thalamus, and on the surface of ependymal cells lining the third ventricle. The choroid plexus, which expresses the short Ob-Ra form, was negative. Combined immunocytochemistry for Ob-Rb protein and fluorescence in situ hybridization for NPY mRNA identified arcuate nucleus neurons containing both NPY mRNA and Ob-Rb protein. The present finding of Ob-Rb protein in neurons that express NPY mRNA supports the hypothesis that arcuate nucleus NPY neurons are direct targets of leptin and play an important role in regulation of food intake and body weight.     

α-Synuclein and neuronal cell death

Mutations in the DJ-1 gene have been linked to autosomal recessive familial Parkinson's disease. To understand the function of DJ-1, we determined the DJ-1 expression in both zebrafish and post mortem human brains. We found that DJ-1 was expressed early during zebrafish development and throughout adulthood. Knock down (KD) of DJ-1 by injection of morpholino did not cause dramatic morphologic alterations during development, and no loss of dopaminergic neurons was observed in embryos lacking DJ-1. However, DJ-1 KD embryos were more susceptible to programmed cell death. While a slight reduction in staining for islet-1 positive neurons was observed in both DJ-1 KD and H₂O₂ treated embryos, the number of apoptotic cells was significantly increased in both KD and H₂O₂ treated embryos. Interestingly, DJ-1 expression was increased in brains of zebrafish under conditions of oxidative stress, indicating that DJ-1 is a part of stress-responsive machinery. Since oxidative stress is one of the major contributors to the development of Alzheimer's disease (AD), we also examined DJ-1 expression in AD brains. Using DJ-1 specific antibodies, we failed to detect a robust staining of DJ-1 in brain tissues from control subjects. However, DJ-1 immunoreactivity was detected in hippocampal pyramidal neurons and astrocytes of AD brains. Therefore, our results strongly suggest that DJ-1 expression is not necessary during zebrafish development but can be induced in zebrafish exposed to oxidative stress and is present in human AD brains.     

Changes in the neuronal excitability of rat hippocampal slices isolated after destruction of the medial septal area]

To determine if electrophysiological properties of hippocampal pathways are altered after medial septal area (MSA) destruction, extracellular recordings were made from hippocampal slices of rats 30 days following lesion and compared with those from unoperated controls. The preparation of slices, data accumulation and data analyses were done under the same conditions. The electrophysiological parameters of interest were the population spike (PS) and the field EPSP, produced in the CA1 pyramidal layer by stimulation of the Schaffer collaterals. The principal finding of this study was that neuronal excitability in slices from MSA-lesioned rats was altered. The most striking abnormalities were an epileptiform activity, which consisted of multiple PSs, and multiple seizure-like after discharges with a delayed onset to low stimulation intensities. In the CA1 region of the slices collected from lesioned rats the input-output curve of field EPSP versus PS showed a leftward shift as compared with their counterparts in normal slices. These changes may be related to relative reduction of inhibitory processes in interneuronal circuits of CA1 region.