The role of frontal neurons in pain and acupuncture analgesia

To study the role of frontal neurons in pain and acupuncture analgesia, experiments were carried out on 4 monkeys (Macaca mulatta) during performing two tasks, the Heat Discrimination Response GO/NO-GO Task (Task I) and the Heat Delayed Discrimination GO/NO-GO Task (Task II). After a criterion of 90% of correct performances over three successive days had been reached, activity of single neurons was recorded from the frontal cortex. Of 276 Task I-related neurons, 211 responded to noxious and/or innocuous heat stimuli. Of 73 Task II-related neurons, 59 responded to noxious and/or innocuous heat stimuli. During acupuncture, the nociceptive reaction time for lever release to avoid painful stimuli was increased, the percentage of correct performances was lowered, and the neuronal responses to noxious and/or innocuous stimuli were suppressed. These neurons were mainly located in a circumscribed area medial to the superior ramus of the arcuate sulcus in frontal cortex, including the prefrontal and premotor areas. It is suggested that neurons in this area may be related to the discrimination of noxious and innocuous stimuli. The suppression of the activity of these neurons during acupuncture may be involved in the whole mechanism of acupuncture to facilitate the production of analgesia.     

高效液相色谱-柱后固定化酶反应器-电化学检测法分析大鼠脑微透析液中的乙酰胆碱和胆碱

 建立了用高效液相色谱分离－柱后固定化酶反应器酶解－电化学检测器检测酶解最终产物Ｈ２Ｏ２的方法，分析了麻醉和自由活动大鼠脑微透析液中乙酰胆碱（ＡＣｈ）和胆碱（Ｃｈ）的含量。至少在０．２～１００μｍｏｌ／Ｌ范围内ＡＣｈ和Ｃｈ的浓度与其响应的线性关系良好，它们的检测极限都可达５０ｆｍｏｌ。对高效液相色谱结合固定化酶反应器的分析方法作了简要的讨论。     

微柱高效液相色谱法测定大鼠脑微透析液中 的乙酰胆碱和胆碱

用所建立的微柱高效液相色谱分离、柱后固定化酶反应器酶解、电化学检测器检测的方法, 对用不同浓度异氟醚麻醉前后的大鼠脑微透析液中的乙酰胆碱和胆碱的浓度进行了测定。在 乙酰胆碱和胆碱的浓度为10～ 2 000 nmol/L 时,其浓度分别与各自相应峰高的线性 关系良好,两者的检测限(以信噪比为3计)均可达5 nmol/L 。微透析液中乙酰胆碱和胆碱的含量在大鼠清醒时最高,随着异氟醚吸入浓度的增加,乙酰胆碱和胆碱含量明显降低。在大鼠海马和脑皮层中乙酰胆碱浓度的降低与大鼠吸入异氟醚的浓度相关( P ＜     

脑部磁共振结合静息态fMRI方法在卒中患者认知障碍中的应用

本研究主要探讨脑部磁共振结合静息态fMRI方法在卒中患者认知障碍中的应用。本研究将符合卒中认知障碍患者25例设为患者组,将25例健康者设为对照组。采用脑部静息态核磁共振对实验者进行扫描,以脑部双侧海马为种子点进行功能连接分析,并通过软件分析两组实验者静息态双侧海马差异。结果发现,与对照组相比,患者组右侧海马与左右侧额中回、左侧额下回等的连接减少;右侧海马与左侧颞上中回、双侧丘脑的连接增加;左侧海马与左侧颞额中回、右侧颞额上回、右侧额下回的连接减少;左侧海马与右侧顶下回连接增加。总之,本研究发现卒中患者海马脑区与颞叶皮层、额叶皮层之间的连接减少,可能是由卒中患者脑损害导致。     

Participation of the Cholinergic System in the Development of Polycystic Ovary Syndrome

In rats with polycystic ovary syndrome (PCOS) induced by injection of estradiol valerate (EV), unilateral or bilateral section of the vagus nerve restores ovulatory function in 75% of animals, suggesting that the vagus nerve participates in the development of PCOS. Since the vagus nerve is a mixed nerve through which mainly cholinergic-type information passes, the objective of the present study was to analyze whether acetylcholine (ACh) is involved in the development of PCOS. Ten-day-old rats were injected with 2.0 mg EV, and at 60 days of age, they were microinjected on the day of diestrus in the bursa of the left or right ovary with 100 or 700 mg/kg of ovarian weight atropine, a blocker of muscarinic receptors, and sacrificed for histopathological examination after the surgery. Animals with PCOS microinjected with 100 mg of atropine showed a lack of ovulation, lower serum concentrations of progesterone and testosterone, and cysts. Histology of the ovaries of animals microinjected with 700 mg of atropine showed corpus luteum and follicles at different stages of development, which was accompanied by a lower concentration of progesterone and testosterone. These results allow us to suggest that in animals with PCOS, ACh, which passes through parasympathetic innervation, is an important component in the persistence and development of the pathophysiology.     

嗅觉记忆相关蛋白的蛋白质组学研究

应用双向凝胶电泳结合质谱鉴定和数据库检索, 分析比较了C57BL/6J小鼠在嗅觉训练和记忆测试后嗅球蛋白表达的差异, 探讨了与嗅觉记忆相关的蛋白质. C57BL/6J小鼠经嗅觉训练后, 可对相应的气味保持记忆能力, 其嗅球蛋白表达存在明显差异, 5种蛋白与嗅觉记忆形成显著相关. 上述蛋白功能涉及神经发育, 信号转导及细胞骨架和核苷酸代谢, 其中神经发育和信号转导相关蛋白表达上调, 而细胞骨架和核苷酸代谢相关蛋白表达水平降低. 这些与嗅觉记忆形成相关的蛋白深化了对嗅觉记忆机制的认识, 为研究和治疗认知相关疾病提供了新靶标.     

Repeated electroconvulsive seizure induces c-Myc down-regulation and Bad inactivation in the rat frontal cortex

Repeated electroconvulsive seizure (ECS), a model for electroconvulsive therapy (ECT), exerts neuroprotective and proliferative effects in the brain. This trophic action of ECS requires inhibition of apoptotic activity, in addition to activation of survival signals. c-Myc plays an important role in apoptosis of neurons, in cooperation with the Bcl-2 family proteins, and its activity and stability are regulated by phosphorylation and ubiquitination. We examined c-Myc and related proteins responsible for apoptosis after repeated ECS. In the rat frontal cortex, repeated ECS for 10 days reduced the total amount of c-Myc, while increasing phosphorylation of c-Myc at Thr58, which reportedly induces degradation of c-Myc. As expected, ubiquitination of both phosphorylated and total c-Myc increased after 10 days ECS, suggesting that ECS may reduce c-Myc protein level via ubiquitination-proteasomal degradation. Bcl-2 family proteins, caspase, and poly(ADP-ribose) polymerase (PARP) were investigated to determine the consequence of down-regulating c-Myc. Protein levels of Bcl-2, Bcl-X(L), Bax, and Bad showed no change, and cleavage of caspase-3 and PARP were not induced. However, phosphorylation of Bad at Ser-155 and binding of Bad to 14-3-3 increased without binding to Bcl-X(L) after repeated ECS, implying that repeated ECS sequesters apoptotic Bad and frees pro-survival Bcl-XL. Taken together, c-Myc down-regulation via ubiquitination-proteasomal degradation and Bad inactivation by binding to 14-3-3 may be anti-apoptotic mechanisms elicited by repeated ECS in the rat frontal cortex. This finding further supports the trophic effect of ECS blocking apoptosis as a possible therapeutic effect of ECT.     

The influence of cholesterol on the generation of radicals in mixed reverse micelles of cationic surfactants with hydroperoxides

Cationic surfactants (S+) and acetylcholine (ACh), the most important neurotransmitter playing an essential role in the neuromuscular and cognitive activity of living beings, form mixed reverse micelles with hydroperoxides (ROOH) in organic media, where ROOH decay into free radicals is catalytically accelerated. Adding cholesterol (Chol, 30 mol.%) to pyridinium (СРВ) and cetyltimethylammonioum (СТАВ) bromides, reduces the radical generation rate in ROOH catalytic decay several times. However, a higher radical initiation rate is observed in the case of less ordered and larger ACh–ROOH reverse micelles. A Chol additive does not essentially affect the size of СТАВ and СРВ micelles with hydroperoxides but results in their decrease in the case of ACh–ROOH.

     

DEPTH MEASUREMENTS AND HISTOPATHOLOGICAL CHARACTERIZATION OF PHOTODYNAMIC THERAPY GENERATED NORMAL BRAIN NECROSIS AS A FUNCTION OF INCIDENT OPTICAL ENERGY DOSE

The response of normal brain to photodynamic therapy (PDT) was investigated in 62 Fisher rats. The animals were injected i.p. with Photofrin II (12.5 mg/kg). Forty-eight hours following injection, an area of dura 5 mm in diameter over the frontal cortex was photoactivated with red light (632 +/- 2 nm) at 100 mW cm-2, with no contributing thermal increases, at optical energy doses ranging from 1-140 J cm-2 from an argon-pumped dye laser. Appropriate controls were also prepared. Brain tissue samples for histological analysis were taken 24 h following PDT treatment. Maximum lesion depth perpendicular to the pial brain surface, was measured using an eyepiece micrometer. Lesions of increasing depth were generated as the incident optical energy dose was increased. Fitting the depth of necrosis to a natural log dependence of incident optical dose yielded a slope of 0.83 mm/ln J cm-2 (r2 = 0.99). The intercept of 1.47 J cm-2 indicated the energy dose below which no normal tissue damage would occur at the incident laser intensity of 100 mW cm-2. The smallest lesions consisted almost exclusively of isolated neuronal injury and neuropil vacuolation, suggestive of an early ischemic lesion. Damage at the upper energy levels (35-140 J cm-2) consisted of complete coagulative necrosis identical to that induced by an arterial occlusion. The existence of viable tissue alongside neurons in various stages of necrosis at low energy levels (less than 35 J cm-2) is suggestive of reversible injury and possibly clinically relevant treatment levels.     

Subsynaptic Distribution,Lipid Raft Targeting and G Protein-Dependent Signalling of the Type 1 Cannabinoid Receptor in Synaptosomes from the Mouse Hippocampus and Frontal Cortex

Numerous studies have investigated the roles of the type 1 cannabinoid receptor (CB1) in glutamatergic and GABAergic neurons. Here, we used the cell-type-specific CB1 rescue model in mice to gain insight into the organizational principles of plasma membrane targeting and Gαi/o protein signalling of the CB1 receptor at excitatory and inhibitory terminals of the frontal cortex and hippocampus. By applying biochemical fractionation techniques and Western blot analyses to synaptosomal membranes, we explored the subsynaptic distribution (pre-, post-, and extra-synaptic) and CB1 receptor compartmentalization into lipid and non-lipid raft plasma membrane microdomains and the signalling properties. These data infer that the plasma membrane partitioning of the CB1 receptor and its functional coupling to Gαi/o proteins are not biased towards the cell type of CB1 receptor rescue. The extent of the canonical Gαi/o protein-dependent CB1 receptor signalling correlated with the abundance of CB1 receptor in the respective cell type (glutamatergic versus GABAergic neurons) both in frontal cortical and hippocampal synaptosomes. In summary, our results provide an updated view of the functional coupling of the CB1 receptor to Gαi/o proteins at excitatory and inhibitory terminals and substantiate the utility of the CB1 rescue model in studying endocannabinoid physiology at the subcellular level.     

Explosion hazards of ion exchange resin mixed with perchloric acid

On January 21, 2003, an explosion occurred while ion exchange resin (IER) was being used to separate impurities from uranium solution. To clarify the cause of the accident and go/no-go criteria of the explosion, elemental analysis of the IER, DSC analysis, and SIKAREX analysis (a screening tool for runaway reactions) were performed. Finally, experiments on the same scale as the accident were conducted in an explosion chamber. When HClO₄ was added to IER-NO₃, the IER violently exploded without any heating nor metal ions such as uranium. It was confirmed that the accident was caused by an incorrect procedure in the chemical process. From the standpoint of explosion safety, IER-NO₃ in particular should be kept away from perchloric acid in the laboratory.     

INDUCTION OF LEAF UNROLLING BY PHYTOCHROME and ACETYLCHOLINE IN ETIOLATED WHEAT SEEDLINGS

Abstract-Phytochrome regulates the unrolling of primary leaf sections from 8-day-old dark-grown wheat ( Triticum aeslivum L. cv. Arminda) seedlings. Red light (R)-stimulated unrolling of leaf sections pretreated in 1 m M ethylene-bis-(β-aminoethylether)- N,N,N',N' -tetraacetic acid (EGTA) if 1 m M CaCl₂ was added during a 30 min treatment period including and following irradiation. Nifedipine at 1 μ M (a Ca²+-channel antagonist) applied 10 min before R prevented the R stimulation of leaf unrolling. The Ca²+-channel agonist Bay K-8644 (1 μ M ) and acetylcholine (ACh, 1 mY M ) stimulated unrolling of leaf sections prewashed in EGTA in darkness, if 1 m M CaCl₂ was present in the medium during a 30 min treatment period. Acetylcholine also induced leaf unrolling in the absence of Ca²+ when 100 μ M NaCl was present in the medium. Apart from ACh, only carbamylcholine out of the choline derivatives tested was active in induction of leaf unrolling in the presence of 1 m M Ca²+. The ACh receptor antagonists, atropine (10 μ M ) AND D-tubocurarine (10 μ M ), nullified the ACh-induced Ca²+- and Na+-dependent leaf unrolling, respectively. Muscarine and nicotine, agonists of ACh, at 1 μ M stimulated leaf unrolling in the presence of Ca²+ and Na+, respectively. The ACh-induced Ca²+-dependent leaf unrolling was reduced by 1 μ M Nifedipine, 10 μ M Li+ and 10 μ M "calmodulin" inhibitor, trifluoperazine (TFP), whereas only TFP was active in the reduction of the Na+-dependent ACh-induced leaf unrolling response. It is proposed that leaf unrolling of dark-grown primary wheat leaves can be regulated by phytochrome and by activation of two different types of ACh receptors.     

Effect of ginseng saponins on the survival of cerebral cortex neurons in cell cultures

The effects of nerve growth factor (NGF) and saponins isolated from Panax ginseng C.A. Mayer on the survival of chick and rat embryonic cerebral cortex neurons were examined. Ginsenoside Rg1 (GRg1) exerted a survival-promoting effect on both chick and rat cerebral cortex neurons in cell cultures. Ginsenoside Rb1 (GRb1) also had an effect in the rat and displayed some influence in the chick. NGF alone exerted no effect on both neurons, although it did potentiate the GRb1 effect on chick embryonic cerebral cortex neurons, but did not alter the GRb1 effect on rat embryonic cerebral cortex neurons. NGF did not alter the survival-promoting effect of GRg1 on either chick or rat embryonic cerebral cortex neurons. The other saponins alone or with NGF exerted no effect on the survival of cerebral cortex neurons in either the chick or rat.     

A new exposure system for the in vitro detection of GHz field effects on neuronal networks

The possible effects of high-frequency electromagnetic fields (EMF) on biological systems are a subject of public concern and scientific discussion. It is generally accepted that the absorption of part of the field energy may cause a temperature rise in biological tissue. Nevertheless, our setup aims to detect possible athermal effects on the electric activity of neuronal in vitro networks. Such networks were formed by primary neurons derived from the murine frontal cortex and cultivated on micro-sensor chips. The action potentials of the neurons were detected in real time by an integrated, electrically passive microelectrode array. For EMF exposure, the chips were introduced into a rectangular wave-guide that could be operated in the propagating or standing wave modes. The drive signals were either continuous waves (1.9-2.2 GHz) or a generic mobile phone signal (UMTS-standard) of up to approximately 8 W. An on-chip sensor allowed the temperature progression to be recorded. In addition, ISFETs and Clark-like electrodes were integrated for the on-chip detection of pH and O(2), respectively.     

Increased expression of the receptor for advanced glycation end products in neurons and astrocytes in a triple transgenic mouse model of Alzheimer's disease

The receptor for advanced glycation end products (RAGE) has been reported to have a pivotal role in the pathogenesis of Alzheimer''s disease (AD). This study investigated RAGE levels in the hippocampus and cortex of a triple transgenic mouse model of AD (3xTg-AD) using western blotting and immunohistochemical double-labeling to assess cellular localization. Analysis of western blots showed that there were no differences in the hippocampal and cortical RAGE levels in 10-month-old adult 3xTg-AD mice, but significant increases in RAGE expression were found in the 22- to 24-month-old aged 3xTg-AD mice compared with those of age-matched controls. RAGE-positive immunoreactivity was observed primarily in neurons of aged 3xTg-AD mice with very little labeling in non-neuronal cells, with the notable exception of RAGE presence in astrocytes in the hippocampal area CA1. In addition, RAGE signals were co-localized with the intracellular amyloid precursor protein (APP)/amyloid beta (Aβ) but not with the extracellular APP/Aβ. In aged 3xTg-AD mice, expression of human tau was observed in the hippocampal area CA1 and co-localized with RAGE signals. The increased presence of RAGE in the 3xTg-AD animal model showing critical aspects of AD neuropathology indicates that RAGE may contribute to cellular dysfunction in the AD brain.     

Effects of Muscle Electrical Activity on the Transmission of Developing Neuromuscular Junction

Miniature endplate potentials (MEPPS) caused by the spontaneous release of ACh from the growth cone of cholinergic neurons, are recorded by the whole-cell patch-clamp technique on a large number of 1-day cultured myoballs which have contact neurites of co-cultured neurons. Both muscle cell and neuron are dissociated from the 1-day-old (about stage 20) Xenopus embryo. Frequency and/or amplitude of MEPPs can obviously increase after the repetitive high-level depolarization caused by the stimuli on muscle cells. No detectable changes of single ACh receptor channel property are observed by using the single-channel recording technique. These results suggest that the mechanism of the increase of MEPPs after electrical activity of postsynaptic muscle cells probably involve some alteration of presynaptic membrane.     

Antidementia Effects of Alternanthera philoxeroides in Ovariectomized Mice Supported by NMR-Based Metabolomic Analysis

The crude ethanol extract of the whole plant of Alternanthera philoxeroides (Mart.) Griseb was investigated for its potential as antidementia, induced by estrogen deprivation, based on in vitro antioxidant activity, β-amyloid aggregation inhibition and cholinesterase inhibitory activity, as well as in vivo Morris water maze task (MWMT), novel object recognition task (NORT), and Y-maze task. To better understand the effect of the extract, oxidative stress-induced brain membrane damage through lipid peroxidation in the whole brain was also investigated. Additionally, expressions of neuroinflammatory cytokines (IL-1β, IL-6 and TNF-α) and estrogen receptor-mediated facilitation genes such as PI3K and AKT mRNA in the hippocampus and frontal cortex were also evaluated. These effects were confirmed by the determination of its serum metabolites by NMR metabolomic analysis. Both the crude extract of A. philoxeroides and its flavone constituents were found to inhibit β-amyloid (Aβ) aggregation.     

Aged Brains Express Less Melanocortin Receptors,Which Correlates with Age-Related Decline of Cognitive Functions

Brain G-protein coupled receptors have been hypothesized to be potential targets for maintaining or restoring cognitive function in normal aged individuals or in patients with neurodegenerative disease. A number of recent reports suggest that activation of melanocortin receptors (MCRs) in the brain can significantly improve cognitive functions of normal rodents and of different rodent models of the Alzheimer’s disease. However, the potential impact of normative aging on the expression of MCRs and their potential roles for modulating cognitive function remains to be elucidated. In the present study, we first investigated the expression of these receptors in six different brain regions of young (6 months) and aged (23 months) rats following assessment of their cognitive status. Correlation analysis was further performed to reveal potential contributions of MCR subtypes to spatial learning and memory. Our results revealed statistically significant correlations between the expression of several MCR subtypes in the frontal cortex/hypothalamus and the hippocampus regions and the rats’ performance in spatial learning and memory only in the aged rats. These findings support the hypothesis that aging has a direct impact on the expression and function of MCRs, establishing MCRs as potential drug targets to alleviate aging-induced decline of cognitive function.     

Distribution of lipids in human brain

The enormous abundance of lipid molecules in the central nervous system (CNS) suggests that their role is not limited to be structural and energetic components of cells. Over the last decades, some lipids in the CNS have been identified as intracellular signalers, while others are known to act as neuromodulators of neurotransmission through binding to specific receptors. Neurotransmitters of lipidic nature, currently known as neurolipids, are synthesized during the metabolism of phospholipid precursors present in cell membranes. Therefore, the anatomical identification of each of the different lipid species in human CNS by imaging mass spectrometry (IMS), in association with other biochemical techniques with spatial resolution, can increase our knowledge on the precise metabolic routes that synthesize these neurolipids and their localization. The present study shows the lipid distribution obtained by MALDI-TOF IMS in human frontal cortex, hippocampus, and striatal area, together with functional autoradiography of cannabinoid and LPA receptors. The combined application of these methods to postmortem human brain samples may be envisioned as critical to further understand neurological diseases, in general, and particularly, the neurodegeneration that accompanies Alzheimer’s disease.