Differences in kainate receptor involvement in hippocampal mossy fibre long-term potentiation depending on slice orientation

Long-term potentiation (LTP) is a well-established experimental model used to investigate the synaptic basis of learning and memory. LTP at mossy fibre - CA3 synapses in the hippocampus is unusual because it is normally N-methyl-d-aspartate (NMDA) receptor-independent. Instead it seems that the trigger for mossy fibre LTP involves kainate receptors (KARs). Although it is generally accepted that pre-synaptic KARs play an essential role in frequency facilitation and LTP, their subunit composition remains a matter of significant controversy. We have reported previously that both frequency facilitation and LTP can be blocked by selective antagonism of GluK1 (formerly GluR5/Glu(K5))-containing KARs, but other groups have failed to reproduce this effect. Moreover, data from receptor knockout and mRNA expression studies argue against a major role of GluK1, supporting a more central role for GluK2 (formerly GluR6/Glu(K6)). A potential reason underlying the controversy in the pharmacological experiments may reside in differences in the preparations used. Here we show differences in pharmacological sensitivity of synaptic plasticity at mossy fibre - CA3 synapses depend critically on slice orientation. In transverse slices, LTP of fEPSPs was invariably resistant to GluK1-selective antagonists whereas in parasagittal slices LTP was consistently blocked by GluK1-selective antagonists. In addition, there were pronounced differences in the magnitude of frequency facilitation and the sensitivity to the mGlu2/3 receptor agonist DCG-IV. Using anterograde labelling of granule cells we show that slices of both orientations possess intact mossy fibres and both large and small presynaptic boutons. Transverse slices have denser fibre tracts but a smaller proportion of giant mossy fibre boutons. These results further demonstrate a considerable heterogeneity in the functional properties of the mossy fibre projection.     

Salt sensitivity in chickpea (Cicer arietinum L.): ions in reproductive tissues and yield components in contrasting genotypes

The reproductive phase in chickpea (Cicer arietinum L.) is affected by salinity, but little is known about the underlying cause. We investigated whether high concentrations of Na⁺ and Cl^– in the reproductive structures influence reproductive processes. Chickpea genotypes contrasting in tolerance were subjected to 0, 35 or 50 mm NaCl applied to soil in pots. Flower production and abortion, pod number, percentage of empty pods, seed number and size were evaluated. The concentrations of Na⁺, K⁺ and Cl^– were measured in various plant tissues and, using X‐ray microanalysis, in specific cells of developing reproductive structures. Genotypic variation in reproductive success measured as seed yield in saline conditions was associated with better maintenance of flower production and higher numbers of filled pods (and thus seed number), whereas seed size decreased in all genotypes. Despite the variation in reproductive success, the accumulation of Na⁺ and Cl^– in the early reproductive tissues of developing pods did not differ between a tolerant (Genesis836) and a sensitive (Rupali) genotype. Similarly, salinity tolerance was not associated with the accumulation of salt ions in leaves at the time of reproduction or in seeds at maturity.     

重组肾上腺髓质素表达载体的构建及在哺乳动物细胞中的表达

为探索通过体内表达肾上腺髓质素（adrenomedullin,AM）治疗高血压和慢性心衰的可能性,本实验构建了重组AM真核表达载体,并在无内源笥AM表达的K562细胞株上进行了体外表达实验。实验中采用RT－PCR技术扩增AM cDNA片段,并将扩增的cDNA片段插入pcDNA3.1真核表达质粒,构建成含AM cDNA的重组质料pcDNA3.1AM。用脂质体介导将该质粒转染培养的人白血病细腻K562株,在转染的细胞中,用RT－PCR检测证实有AM mRNA存在;用班点免疫分析方法检测转染细胞的培养液上清,证实有AM多肽存在,表明本实验中构建的重组pcDNA3.1AM载体能够在哺乳类细胞中表达AM。     

A Survey of Heavy Metal Levels in Eels (Anguilla anguilla) from some Rivers in East Anglia,England: the Use of Eels as Pollution Indicators

A survey was made of mercury, cadmium and lead in the liver and flesh of 517 eels from eleven rivers in East Anglia. Eels from some sites showed high metal levels, indicating sources of metal contamination. Eel liver is a suitable indicator of inorganic pollution and the flesh is a suitable indicator of organic pollution.     

Cancer cells that survive checkpoint adaptation contain micronuclei that harbor damaged DNA

We have examined the relationship between checkpoint adaptation (mitosis with damaged DNA) and micronuclei. Micronuclei in cancer cells are linked to genomic change, and may induce chromothripsis (chromosome shattering). We measured the cytotoxicity of the cancer drug cisplatin in M059K (glioma fibroblasts, IC50 15 μM). Nearly 100% of M059K cells were positive for histone γH2AX staining after 48 h treatment with a cytotoxic concentration of cisplatin. The proportion of micronucleated cells, as confirmed by microscopy using DAPI and lamin A/C staining, increased from 24% to 48%, and the total micronuclei in surviving cells accumulated over time. Promoting entry into mitosis with a checkpoint inhibitor increased the number of micronuclei in cells whereas blocking checkpoint adaptation with a Cdk inhibitor reduced the number of micronuclei. Interestingly, some micronuclei underwent asynchronous DNA replication, relative to the main nuclei, as measured by deoxy-bromo-uracil (BrdU) staining. These micronuclei stained positive for histone γH2AX, which was linked to DNA replication, suggesting that micronuclei arise from checkpoint adaptation and that micronuclei may continue to damage DNA. By contrast the normal cell line WI-38 did not undergo checkpoint adaptation when treated with cisplatin and did not show changes in micronuclei number. These data reveal that the production of micronuclei by checkpoint adaptation is part of a process that contributes to genomic change.     

Alpha-synuclein overexpression negatively regulates insulin receptor substrate 1 by activating mTORC1/S6K1 signaling

Alpha-synuclein (α-Syn) is a major component of Lewy bodies, a pathological feature of Parkinson's and other neurodegenerative diseases collectively known as synucleinopathies. Among the possible mechanisms of α-Syn-mediated neurotoxicity is interference with cytoprotective pathways such as insulin signaling. Insulin receptor substrate (IRS)-1 is a docking protein linking IRs to downstream signaling pathways such as phosphatidylinositol 3-kinase/Akt and mammalian target of rapamycin (mTOR)/ribosomal protein S6 kinase (S6K)1; the latter exerts negative feedback control on insulin signaling, which is impaired in Alzheimer's disease. Our previous study found that α-Syn overexpression can inhibit protein phosphatase (PP)2A activity, which is involved in the protective mechanism of insulin signaling. In this study, we found an increase in IRS-1 phosphorylation at Ser636 and decrease in tyrosine phosphorylation, which accelerated IRS-1 turnover and reduced insulin-Akt signaling in α-Syn-overexpressing SK-N-SH cells and transgenic mice. The mTOR complex (C)1/S6K1 blocker rapamycin inhibited the phosphorylation of IRS-1 at Ser636 in cells overexpressing α-Syn, suggesting that mTORC1/S6K1 activation by α-Syn causes feedback inhibition of insulin signaling via suppression of IRS-1 function. α-Syn overexpression also inhibited PP2A activity, while the PP2A agonist C2 ceramide suppressed both S6K1 activation and IRS-1 Ser636 phosphorylation upon α-Syn overexpression. Thus, α-Syn overexpression negatively regulated IRS-1 via mTORC1/S6K1 signaling while activation of PP2A reverses this process. These results provide evidence for a link between α-Syn and IRS-1 that may represent a novel mechanism for α-Syn-associated pathogenesis.     

Contribution of estrogen receptors alpha and beta to the effects of estradiol in the brain

Clinical and experimental studies show a modulatory role of estrogens in the brain and suggest their beneficial action in mental and neurodegenerative diseases. The estrogen receptors ER and ERβ are present in the brain and their targeting could bring selectivity and reduced risk of cancer. Implication of ERs in the effect of estradiol on dopamine, opiate and glutamate neurotransmission is reviewed. The ER agonist, PPT, is shown as estradiol to modulate hippocampal NMDA receptors and AMPA receptors in cortex and striatum of ovariectomized rats whereas the ERβ agonist DPN is inactive. Striatal DPN activity suggests implication of ERβ in estradiol modulation of D2 receptors and transporters in ovariectomized rats and is supported by the lack of effect of estradiol in ERβ knockout (ERKOβ) mice. Both ER and ERβ agonists modulate striatal preproenkephalin (PPE) gene expression in ovariectomized rats. In male mice PPT protects against MPTP toxicity to striatal dopamine; this implicates Akt/GSK3β signaling and the apoptotic regulators Bcl2 and Bad. This suggests a role for ER in striatal dopamine neuroprotection. ERKO mice are more susceptible to MPTP toxicity and not protected by estradiol; differences in ERKOβ mice are subtler. These results suggest therapeutic potential for the brain of ER specific agonists.