Radiation-induced alterations in synaptic neurotransmission of dentate granule cells depend on the dose and species of charged particles.

The evaluation of potential health risks associated with neuronal exposure to space radiation is critical for future long duration space travel. The purpose of this study was to evaluate and compare the effects of low-dose proton and high-energy charged particle (HZE) radiation on electrophysiological parameters of the granule cells in the dentate gyrus (DG) of the hippocampus and its associated functional consequences. We examined excitatory and inhibitory neurotransmission in DG granule cells (DGCs) in dorsal hippocampal slices from male C57BL/6 mice at 3 months after whole body irradiation with accelerated proton, silicon or iron particles. Multielectrode arrays were used to investigate evoked field synaptic potentials, an extracellular measurement of synaptic excitability in the perforant path to DG synaptic pathway. Whole-cell patch clamp recordings were used to measure miniature excitatory postsynaptic currents (mEPSCs) and miniature inhibitory postsynaptic currents (mIPSCs) in DGCs. Exposure to proton radiation increased synaptic excitability and produced dose-dependent decreases in amplitude and charge transfer of mIPSCs, without affecting the expression of γ-aminobutyric acid type A receptor α2, ß3 and γ2 subunits determined by Western blotting. Exposure to silicon radiation had no significant effects on synaptic excitability, mEPSCs or mIPSCs of DGCs. Exposure to iron radiation had no effect on synaptic excitability and mIPSCs, but significantly increased mEPSC frequency at 1 Gy, without changes in mEPSC kinetics, suggesting a presynaptic mechanism. Overall, the data suggest that proton and HZE exposure results in radiation dose- and species-dependent long-lasting alterations in synaptic neurotransmission, which could cause radiation-induced impairment of hippocampal-dependent cognitive functions.

Exposure to 56Fe-particle radiation accelerates electrophysiological alterations in the hippocampus of APP23 transgenic mice.

Abstract An unavoidable complication of space travel is exposure to high-charge, high-energy (HZE) particles. In animal studies, exposure of the CNS to HZE-particle radiation leads to neurological alterations similar to those seen in aging or Alzheimer's disease. In this study we examined whether HZE-particle radiation accelerated the age-related neuronal dysfunction that was previously described in transgenic mice overexpressing human amyloid precursor protein (APP). These APP23 transgenic mice exhibit age-related behavioral abnormalities and deficits in synaptic transmission. We exposed 7-week-old APP23 transgenic males to brain-only (56)Fe-particle radiation (600 MeV/nucleon; 1, 2, 4 Gy) and recorded synaptic transmission in hippocampal slices at 2, 6, 9, 14 and 18-24 months. We stimulated Schaeffer collaterals and recorded field excitatory postsynaptic potentials (fEPSP) and population spikes (PS) in CA1 neurons. Radiation accelerated the onset of age-related fEPSP decrements recorded at the PS threshold from 14 months of age to 9 months and reduced synaptic efficacy. At 9 months, radiation also reduced PS amplitudes. At 6 months, we observed a temporary deficit in paired-pulse inhibition of the PS at 2 Gy. Radiation did not significantly affect survival of APP23 transgenic mice. We conclude that irradiation of the brain with HZE particles accelerates Alzheimer's disease-related neurological deficits.

Radiation exposure prior to ischemia decreases lesion volume, brain edema and cell death.

PURPOSE: To investigate the neuronal response to ischemic injury following exposure to whole brain proton irradiation. METHODS: Brain only proton irradiation (8 Gy, 250 MeV) was performed ten days prior to middle cerebral artery occlusion (MCAO) in 1 year old male Sprague Dawley rats. MCAO was induced in two animal groups: proton irradiated (MCAO + Rad) and MCAO only. Magnetic resonance imaging (MRI) and quantitative analysis were performed prior to and 2 days after irradiation, and then 2, 14 and 28 days after MCAO. After the last imaging time point animals were sacrificed and TUNEL staining was performed on 4% paraformaldehyde - fixed brain sections. RESULTS: Neuroimaging demonstrated a reduction in ischemic lesion volume in the MCAO + Rad group compared with MCAO alone. Neurological deficits did not differ between ischemia groups. Interestingly, there was a 34% decrease in the number of TUNEL-positive cells in MCAO + Rad brains compared to MCAO alone. CONCLUSION: Our results suggest that radiation treatment reduces brain edema, ischemic lesion volume and peri-ischemic apoptosis. The underlying mechanisms are currently unknown and additional studies will elucidate the significance of these results.

Decreased brain levels of 2',3'-cyclic nucleotide-3'-phosphodiesterase in Down syndrome and Alzheimer's disease.

In Down syndrome (DS) as well as in Alzheimer's disease (AD) oligodendroglial and myelin alterations have been reported. 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) and carbonic anhydrase II (CA II) are widely accepted as markers for oligodendroglia and myelin. However, only data on CNPase activity have been available in AD and DS brains so far. In our study we determined the protein levels of CNPase and CA II in DS, AD and in control post mortem brain samples in order to assess oligodendroglia and myelin alterations in both diseases. We used two dimensional electrophoresis to separate brain proteins that were subsequently identified by matrix assisted laser desorption and ionization mass-spectroscopy (MALDI-MS). Seven brain areas were investigated (frontal, temporal, occipital and parietal cortex, cerebellum, thalamus and caudate nucleus). In comparison to control brains we detected significantly decreased CNPase protein levels in frontal and temporal cortex of DS patients. The level of CA II protein in DS was unchanged in comparison to controls. In AD brains levels of CNPase were decreased in frontal cortex only. The level of CA II in all brain areas in AD group was comparable to controls. Changes of CNPase protein levels in DS and AD are in agreement with the previous finding of decreased CNPase activity in DS and AD brain. They probably reflect decreased oligodendroglial density and/or reduced myelination. These can be secondary to disturbances in axon/oligodendroglial communication due to neuronal loss present in both diseases. Alternatively, reduced CNPase levels in DS brains may be caused by impairment of glucose metabolism and/or alterations of thyroid functions.

The reduction of NADH ubiquinone oxidoreductase 24- and 75-kDa subunits in brains of patients with Down syndrome and Alzheimer's disease.

NADH: ubiquinone oxidoreductase (complex I), one of the most complicated multi-protein enzyme complexes, is important for energy metabolism because it is the initial enzyme of the mitochondrial respiratory chain. Deficiency of complex I is frequently found in various tissues of patients with neurodegenerative disease. Here we studied the protein levels of complex I 24- and 75-kDa subunits in several brain regions from patients with Down syndrome (DS) and Alzheimer's disease (AD). We determined protein levels of complex I 24-, 75-kDa subunits and mitochondrial marker proteins mitochondrial matrix protein P1 (hsp60) and aconitate hydratase from seven brain regions of patients with DS, AD and controls. Proteins were separated by two-dimensional (2-D) gel electrophoresis and identified by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS). Complex I 24-kDa subunit was significantly reduced in occipital cortex and thalamus in patients with DS and temporal and occipital cortices in patients with AD. Complex I 75-kDa subunit was significantly reduced in brain regions from patients with DS (temporal, occipital and caudate nucleus) and AD (parietal cortex). Reductions of two subunits of complex I may lead to the impairment of energy metabolism and result in neuronal cell death (apoptosis), a hallmark of both neurodegenerative disorders.

Differential expression of molecular chaperones in brain of patients with Down syndrome.

Heat shock proteins (HSPs) in their molecular capacity as chaperones have been reported to regulate the apoptotic pathway and also play a critical role in protein conformational diseases such as Alzheimer's disease (AD). As all Down syndrome (DS) brains display AD-like neuropathology, neuronal loss in DS was shown to be mediated by apoptosis. We decided to investigate the expression patterns of HSPs in seven brain regions of adults with DS using two-dimensional polyacrylamide gel electrophoresis (2-DE). Following 2-DE, approximately 120 protein spots were successfully identified by matrix-assisted laser desorption/ionization--mass spectrometry (MALDI-MS) followed by quantification of the identified proteins. We unambiguously identified and quantified nine different chaperone proteins. Accordingly, all but three chaperone proteins did exhibit a significant change in expression. HSP 70 RY, heat shock cognate (HSC) 71 and glucose-regulated protein (GRP) 75 showed a significant decrease (P < 0.05) in DS temporal cortex whereas HSP 70.1 and GRP 78 were significantly increased (P<0.05) in cerebellum. Whilst T-complex 1 (TCP-1) epsilon subunit showed a significant decrease (P< 0.05) in parietal cortex, a similar extent of increase (P<0.05) as that observed in cerebellum was obtained in parietal levels of GRP 78. Alpha-crystallin B, HSP 60 and GRP 94 did not show any detectable changes in expression patterns. This report presents the first approach to quantify nine different chaperones simultaneously at the protein level in different brain regions and provides evidence for aberrant chaperone expression patterns in DS. The relevance of this aberrant expression patterns are discussed in relation to the biochemical and neuropathological abnormalities in DS brain.

Decreased levels of complex III core protein 1 and complex V beta chain in brains from patients with Alzheimer's disease and Down syndrome.

Ubiquinol:cytochrome c oxidoreductase (complex III) and ATP synthase (complex V) are important enzymes in the mitochondrial electron transport chain. Defects in mitochondrial respiratory enzymes have been reported for several neurodegenerative diseases. In this study, we applied the proteomic approach to investigate protein levels of complex III core protein and complex V beta chain in brain regions of Alzheimer's disease (AD) and Down syndrome (DS) patients. Complex III core protein 1 was significantly reduced in the temporal cortex of AD patients. Complex V beta chain was significantly reduced in the frontal cortex of DS patients. We conclude that decreased mitochondrial respiratory enzymes could contribute to the impairment of energy metabolism observed in DS. These decreases could also cause the generation of reactive oxygen species and neuronal cell death (apoptosis) in DS as well as AD.

Effect of stobadine, U-74389G, trolox and melatonin on resistance of rat hippocampal slices to oxidative stress.

Reactive oxygen species have been suggested to participate in the impairment of nervous tissue by oxidative stress, induced by hypoxia (HYP) followed by reoxygenation (ROX). Although the mechanisms of such injury are rather complex, antioxidants might exert some protective action under such circumstances. This study tested the effect of a series of compounds interfering with the generation and action of reactive oxygen species on impairment of synaptic transmission in the CA1 region of rat hippocampal slices exposed to HYP followed by ROX in vitro. Shortlasting HYP (typically 4.5-7.5 min under the conditions used) resulted in fast decay of the amplitude of population spikes evoked in the CA1 neurons by stimulation of Schäffer collaterals. The impairment was mostly irreversible. However, in the presence of the antioxidants stobadine, 21-aminosteroid U-74389G, melatonin and trolox (with optimal concentrations of 10-30 micromol/l, 10 micromol/l, 30-100 micromol/l and 200 micromol/l, respectively), the irreversible damage of the transmission was significantly diminished. The decay of the synaptic transmission failure during HYP was also delayed by stobadine, U-74389G and melatonin. The results demonstrated that compounds with antioxidant activity may effectively protect nervous tissue during HYP and ROX.

Effects of stobadine, melatonin, and other antioxidants on hypoxia/reoxygenation-induced synaptic transmission failure in rat hippocampal slices.

In vitro reversible ischemia was simulated with rat hippocampal slices in order to test the neuroprotective activity of selected antioxidants with emphasis on the pyridoindole stobadine. Slices were exposed to hypoxia (HYP) combined with lowered D-glucose concentration to induce synaptic transmission (ST) failure, which turned out to be irreversible in approximately 80%-100% of slices during reoxygenation (ROX). The amplitude of population spikes (PoS) evoked trans-synaptically by electrical stimulation of Schäffer collaterals and recorded in CA1 neurons was the parameter of ST. Pretreatment of slices with stobadine dissolved in slice superfusion media (1 to 100 microM) improved ST recovery after 20-min tissue ROX. Stobadine decreased the number of irreversibly damaged slices and increased the average amplitude of PoS during tissue ROX. The concentration-response relationship of protective activity was bell-shaped, with maximum at 3-30 microM. Moreover, the half-time of PoS decay (t1/2) during HYP was significantly delayed in stobadine treated groups (10 to 100 microM). The neurohormone melatonin (30 to 100 microM) and 21-aminosteroid U-74389G (10 microM) revealed similar protective activity on ST recovery and on t1/2 during HYP. Trolox (200 microM) improved the PoS recovery, yet it had no effect on t1/2. The iron chelator deferoxamine (250 and 500 microM) had no protective effects at all. alpha-Tocopherol administered to animals orally (200 mg/kg for 10 days) only marginally improved the PoS recovery. Comparing the protective effect of compounds tested on PoS recovery, we assume the following rank order of potency: U-74389G > stobadine > melatonin >> trolox. Our findings suggest that stobadine as well as trolox, U-74389G and melatonin, antioxidants with remarkably different chemical structures, exerted neuroprotective activity, probably determined by antioxidative properties of these compounds. Moreover, stobadine, U-74389G, and melatonin were able to delay the early ST decay during HYP, which might indicate improved energetic state of neurons in the treated tissue. The study supports the notion about the neuroprotective activity of certain antioxidants.

Neuroprotection by the pyridoindole stobadine: a minireview.

The minireview summarizes data documenting that pyridoindole stobadine (STB) may protect nervous structures against oxidative stress. This was demonstrated by the impairment of synaptic transmission in hippocampal slices and sympathetic ganglia exposed to hypoxia/reoxygenation (H/R) in vitro as well as by survival of rats and dogs exposed to brain ischemia/reperfusion (I/R) in vivo. The STB effect was linked mostly to its free radical scavenging and antioxidant properties. STB seems to act primarily on phospholipids, thus protecting the integrity and function of somatic membranes in neurons as well as those in subcellular organelles, such as mitochondria and endoplasmic reticulum. STB prevented damage to Ca2+ sequestering systems in endoplasmic reticulum and synaptosomes induced by lipid peroxidation initiators. It was found to diminish changes in NMDA and adrenergic alpha1-receptors evoked in the brain by I/R or H/R. The compound prevented total thiols, participating in tissue antioxidative protection, from decreasing in brain under these conditions. It readily penetrates into both the hydrophilic and the hydrophobic compartments of the CNS. Data were obtained indicating that in I/R, protection of structures such as brain-blood vessels, endothelium, and/or erythrocytes may participate in the STB effect, besides the direct protection of nervous tissue. STB may be characterized as a potential protectant of the CNS in diseases in which oxidative injury may play an important role, for example, stroke, neurotrauma, chronic brain ischemia, or some neurodegenerative diseases. Its molecule could provide a useful model in the further search for novel compounds with even more pertinent pharmacological and pharmacokinetic profiles.