Effect of various stabilizing agents on Imperata cylindrica grass pollen allergen extract

BACKGROUND: Allergen extracts are unstable, heat labile or susceptible to proteases. Stability of allergen extracts is important for proper diagnosis and therapy of allergic disorders. OBJECTIVE: The present study was undertaken to determine the preservation and stabilization conditions of Imperata cylindrica (Ic) grass pollen extract. METHODS: The Ic extract was kept with 0.1 mepsilon-aminocaproic acid (EACA), 0.75 m sucrose, 5% glycerol, 0.03% human serum albumin (HSA) or 0.4% phenol for different time periods. The extracts were stored for 3, 6 and 12 months each at 4 degrees C, 4 degrees C with daily exposure to room temperature (RT) for 1 h, and RT. The quality of extracts was analysed by SDS-PAGE, Western blot, ELISA, ELISA inhibition and skin test. RESULTS: Extracts kept with EACA and sucrose retained most of the protein bands followed by glycerol as determined by SDS-PAGE and Western blot during all storage periods and conditions in comparison with standard extracts. The extracts kept with HSA, phenol and without preservative (WP) showed protein degradation below 33 kDa after 3 months storage at all conditions. However, a 67-kDa allergen was stable in these extracts. EACA extract required 75 to 120 ng of protein for 50% inhibition in IgE binding under different conditions, whereas standard extract required 70 ng for the same. ELISA also demonstrated high allergenic reactivity of EACA extract. ID test on allergy patients with EACA extract demonstrated same allergenic potency as that of standard extract. CONCLUSION: EACA is the best preservative/stabilizing agent of Ic pollen extract, followed by sucrose and glycerol. Ic extract kept with phenol, HSA and without preservative showed degradation within 3 months. EACA preserved extract is equally potent as that of standard extract up to 1 year's storage.     

Cluster headache: phospholipid metabolism in polymorphonuclear cells

Our group has previously reported significant changes in the incorporation of precursors into glycerophospholipids, particularly phosphatidylserine, in polymorphonuclear cells obtained from the peripheral blood of cluster headache patients, when compared with controls. The potential of these results led to further work using both the previous methodology and a modified isolation technique to obtain polymorphonuclear cells in as pure a state as possible. Neither the new results obtained using the original technique, nor the results with high purity polymorphonuclear cells from controls and cluster headache patients, confirm the marked changes in precursor uptake into glycerophospholipids originally reported.     

The effect of methamphetamine on serotonin and its metabolite in the suprachiasmatic nucleus: A microdialysis study

Summary The suprachiasmatic nucleus (SCN) has been identified as a major circadian pacemaker. Methamphetamine has been shown to modify the behavior of circadian rhythms. We detected extracellular serotonin (5-HT) and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) in the SCN in freely moving rats, using a microdialysis method, to investigate biochemical effects of methamphetamine in the SCN. Methamphetamine infusion into the SCN dose-dependently increased extracellular 5-HT and decreased extracellular 5-HIAA.     

Quisqualate lesions of rat NBM: Selective effects on working memory in a double Y-maze

Some authors have reported that quisqualic acid lesions of the nucleus basalis magnocellularis (NBM), although producing large cortical cholinergic losses, have little effect on memory. The purpose of the present study was to investigate the effects of quisqualic acid lesions of the NBM on working and reference memory in a double Y-maze. Each trial started with placement into one of the two end arms of the first Y-maze, and the correct response was to go down the stem (reference memory). Access was then given to the second Y-maze, the correct response being conditional upon the side of the first Y-maze from which that trial had begun (working memory). Rats were trained to an 88% correct criterion and were then given either bilateral quisqualic acid (60 nM, 0.5 microliters) or sham lesions (0.9% saline, 0.5 microliters) of the NBM. One week postsurgery, rats were tested on the double Y-maze task with delays of 0, 5 or 30 seconds being introduced prior to both the working and reference memory choice. NBM lesions produced a 63.2 +/- 6.2% decrease of cortical choline acetyltransferase (ChAT) compared to unoperated controls. Delays affected only the working memory of the sham group. Rats with lesions showed a significant impairment of working memory at all delays, but no change in reference memory. Results indicate that quisqualic acid lesions of the NBM that produce significant reductions in cortical ChAT selectively impair working memory.     

Quantitative autoradiographic study of L-glutamate binding sites in normal and atrophic human cerebellum.

In the present work the distribution of L-glutamate binding sites in the different layers of human cerebellum of normal individuals and of seven patients who died with olivopontocerebellar atrophy (OPCA) was examined with the technique of quantitative autoradiography. Specific L-[3H]glutamate binding was higher in the molecular than in the granule cell layer of normal cerebellar tissue. A significant decrease of L-[3H]glutamate specific binding was observed in the molecular layer of all OPCA tissues. In the granule cell layer L-[3H]glutamate binding was decreased only in two patients who suffered from late-onset sporadic OPCA and in one patient who suffered from a form of OPCA inherited in a dominant manner. Quisqualate-sensitive binding sites were the most abundant binding sites in the molecular layer of normal cerebella, whereas N-methyl-D-aspartic acid (NMDA)-sensitive binding sites were the most abundant type in the granule cell layer. A significant decrease of quisqualate-sensitive and an increase in NMDA-sensitive binding sites were observed in the molecular layer of OPCA cerebellar tissues. No significant changes were observed in the granule cell layer of these tissues.     

Effect of Vigabatrin on the Electroencephalogram in Rats

Vigabatrin (gamma-vinyl GABA; GVG) is a new antiepileptic drug (AED) that increases the level of the inhibitory transmitter, gamma-aminobutyric acid (GABA) in the brain. We evaluated the effect of GVG on the EEG of normal rats. GVG was administered intraperitoneally (i.p.) at a dose of 100 mg/kg once a day for 12 days. EEG was recorded at baseline, on the fourth day, at the end of the 12-day GVG period and 10 days after discontinuation of GVG. GVG increased the amplitude of delta (1-4 Hz) and theta (4-8 Hz) frequency bands and resulted in slowing of the peak frequency (Fp) and mean frequency (Fm) in both the frontal and occipital cortex, especially during waking-immobility. EEG changes normalized within 10 days after the last GVG injections. The results suggest that a relationship may exist between the EEG changes and increase in GABA levels with GVG.     

Protective effects of serotonin reuptake inhibitors, citalopram and clomipramine, against hippocampal CA1 neuronal damage following transient ischemia in the gerbil

To clarify the role of serotonin in cerebral ischemia, we examined the effects of selective serotonin reuptake inhibitors, citalopram and clomipramine, on ischemic neuronal damage in the gerbil. Pretreatment with citalopram (40 mg/kg i.p.) and clomipramine (20 mg/kg i.p.) protected against neuronal destruction of hippocampal CA1 pyramidal cells following 5 min of forebrain ischemia. Furthermore, microdialysis assays showed that a striking increase in extracellular excitatory amino acid levels during ischemia was significantly inhibited by pretreatment with citalopram and clomipramine. However, citalopram (40 mg/kg i.p.) did not alter the extracellular amino acid concentrations in normal gerbils. Thus, serotonin reuptake inhibitors have a protective effect against ischemic neuronal damage. Furthermore, the present result suggests that the protective effect is mediated through prevention of the accumulation of extracellular excitatory amino acids during and after ischemia.     

Regulation of neurotransmitter enzyme by quisqualate subtype glutamate receptors in cultured cerebellar and hippocampal neurons

The possible involvement of ionotropic and metabotropic quisqualate (QA) receptors in neuronal plasticity was studied in cultured glutamtergic cerebellar or hippocampal cells in terms of the specific activity of phosphate-activated glutaminase, an enzyme important in the synthesis of the putative neurotransmitter pool of glutamate. When cerebellar of hippocampal neurons were treated with QA, it elevated the specific activity of glutaminase in a dose-dependent manner. The half-maximal effect was obtained at about 0.1 μM, the maximum increase was at about 1 μM, but levels higher than 10 μM QA produced progressive reduction in glutaminase activity. In contrast, QA had little effects on the activities of lactate dehydrogenase and aspartate aminotransferase and the amount of protein, indicating that the increase in glutaminase was relatively specific. The QA-mediated increase in glutaminase was mimicked by the ionotropic QA receptor agonist -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA; EC₅₀, about 0.5 μM), but not by the metabotropic QA receptor agonist trans-(±)-1-aino-cyclopentyl-1,3,dicarboxyalte (t-ACPD; up to 0.5 mM). The specific ionotropic QA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) inhibited QA- and AMPA-mediated increases in glutaminase activity in a dose-dependent manner, whereas other glutamate receptor antagonists, -2-amino-5-phosphonovalerate, γ- -glutamyl aminomethyl sulphonic acid and γ- -glutamyl diethyl ester were ineffective. The elevation of neurotransmitter enzyme was Ca²⁺-dependent. The increase in Ca²⁺ influx essentially through the activation of L-type voltage-operated Ca²⁺ channels, and not the mobilization of internal Ca²⁺ stores, was responsible for these QA receptor-mediated long-term plastic changes in hippocampal and cerebellar neurons.     

锰超氧化物岐化酶模型配合物的研究

为探索天然超氧化物岐化酶（ＳＯＤ）岐化超氧离子的机理，及用于临床的可能性，我们合成了乳酸、柠檬酸和酒石酸的锰（Ⅱ）配合物作为Ｍｎ－ＳＯＤ的模拟物，用光照法测定了它们的活性，得到了有意义的结果：锰配合物作为Ｍｎ－ＳＯＤ的模拟物，有肯定的活性，其中以乳酸锰配合物活性最高。     

NMDA Receptor Activation Triggers Voltage Oscillations, Plateau Potentials and Burstinq in Neonatal Rat Lumbar Motoneurons ln Vitro

Whole-cell recordings of lumbar motoneurons in the intact neonatal rat spinal cord in vitro were undertaken to examine the effects of Kmethyl-D-aspartate (NMDA) receptor activation on membrane behaviour. Bath application of NMDA induced rhythmic voltage oscillations of 5.9 ± 2.1 mV (SD) at a frequency of 4.4 ± 1.5 Hz. Amplitude, but not frequency, of the voltage oscillations was membrane potential-dependent. Voltage oscillations could recruit action potentials and/or plateau potentials with or without superimposed bursting. Blockade of synaptic transmission with tetrodotoxin (TTX) sometimes resulted in a loss of oscillatory activity which could then be restored by increasing the NMDA concentration. After application of TTX, the trajectory of NMDA-induced oscillations was similar to the trajectory induced in the presence of intact synaptic networks, although the mean oscillation duration was longer and the oscillation frequency was slower (1.8 ± 1.1 Hz). Current ramps delivered after bath application of NMDA demonstrated bistable membrane properties which may underlie the plateau potentials. Injection of intracellular current pulses could initiate, entrain and terminate individual plateau potentials. The results suggest that membrane depolarization produced by oscillations may activate other intrinsic conductances which generate plateau potentials, thereby providing the neuron with enhanced voltage sensitivity, compared to that produced by NMDA receptor activation alone. These oscillatory events may have a role in the regulation of motor output in a variety of rhythmic behaviours including locomotion.