Excitatory Amino Acid Receptors in the Ventral Tegmental Area Regulate Dopamine Release in the Ventral Striatum

Abstract: The role of excitatory amino acid (EAA) receptors located in the ventral tegmental area (VTA) in tonic and phasic regulation of dopamine release in the ventral striatum was investigated. Microdialysis in conscious rats was used to assess dopamine release primarily from the nucleus accumbens shell region of the ventral striatum while applying EAA antagonists or agonists to the VTA. Infusion of the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (25 and 100 µ M ) into the VTA did not affect dopamine release in the ventral striatum. In contrast, intra-VTA infusion of the NMDA receptor antagonist 2-amino-5-phosphopentanoic acid (100 and 500 µ M ) dose-dependently decreased the striatal release of dopamine. Intra-VTA application of the ionotropic EAA receptor agonists NMDA and AMPA dose-dependently (10 and 100 µ M ) increased dopamine efflux in the ventral striatum. However, infusion of 50 or 500 µ M trans -(±)-1-amino-1,3-cyclopentanedicarboxylic acid (ACPD), a metabotropic EAA receptor agonist, did not significantly affect these levels. These data suggest that NMDA receptors in the VTA exert a tonic excitatory influence on dopamine release in the ventral striatum. Furthermore, dopamine neurotransmission in this region may be enhanced by activation of NMDA and AMPA receptors, but not ACPD-sensitive metabotropic receptors, located in the VTA. These data further suggest that EAA regulation of dopamine release primarily occurs in the VTA as opposed to presynaptically at the terminal level.     

Isolation and partial characterization of a lectin from ground elder (Aegopodium podagraria) rhizomes

A lectin has been isolated from rhizomes of ground elder (Aegopodium podagraria) using a combination of affinity chromatography on erythrocyte membrane proteins immobilized on cross-linked agarose and hydroxyapatite, and ion-exchange chromatography. The molecular structure of the lectin was determined by gelfiltration, sucrose density-gradient centrifugation and gel electrophoresis under denaturing conditions. It has an unusually high M_r (about 480000) and is most probably an octamer composed of two distinct types of subunits with slightly different M_r (about 60000). Hapten inhibition assays indicated that the Aegopodium lectin is preferentially inhibited by N-acetylgalactosamine. Nevertheless, it does not agglutinate preferentially blood-group-A erythrocytes. The ground-elder lectin is a typical non-seed lectin, which occurs virtually exclusively in the underground rhizomes. In this organ it is an abundant protein as it represents up to 5% of the total protein content. The lectin content of the rhizome tissue varies strongly according to its particular location along the organ. In addition, the lectin content changes dramatically as a function of the seasons. The ground-elder lectin differs from all other plant lectins by its unusually high molecular weight. In addition, it is the first lectin to be isolated from a species of the family Apiaceae.Abbreviations APA Aegopodium podagraria agglutinin - PBS phosphate-buffered saline - SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis     

Methods for simultaneous measurement of apoptosis and cell surface phenotype of epithelial cells in effusions by flow cytometry

We describe here a protocol for the detection of epithelial cells in effusions combined with quantification of apoptosis by flow cytometry (FCM). The procedure described consists of the following stages: culturing and induction of apoptosis by staurosporine in control ovarian carcinoma cell lines (SKOV-3 and OVCAR-8); preparation of effusion specimens and cell lines for staining; staining of cancer cells in effusions and cell lines for cell surface markers (Ber-EP4, EpCAM and CD45) and intracellular/nuclear markers of apoptosis (cleaved caspase-3 and caspase-8, and incorporated deoxyuridine triphosphates); and FCM analysis of stained cell lines and effusions. This protocol identifies a specific cell population in cytologically heterogeneous clinical specimens and applies two methods to measure different aspects of apoptosis in the cell population of interest. The cleaved caspase and deoxyuridine triphosphate incorporation FCM assays are run in parallel and require (including sample preparation, staining, instrument adjustment and data acquisition) 8 h. The culturing of cell lines requires 2-3 days and induction of apoptosis requires 16 h.     

Use of an inhibitor to identify members of the hormone-sensitive lipase family

Hormone-sensitive lipase (HSL) contributes importantly to the mobilization of fatty acids from the triacylglycerols stored in adipocytes, which provide the main source of energy in mammals. On the basis of amino acid sequence alignments and three-dimensional structures, this enzyme was previously found to be a suitable template for defining a family of serine carboxylester hydrolases. In this study, the HSL family members are characterized rather on the basis of their inhibition by 5-methoxy-3-(4-phenoxyphenyl)-3H-[1,3,4]oxadiazol-2-one (compound 7600). This compound inhibits mammalian HSL as well as other HSL family members, such as EST2 from the thermophilic eubacterium Alicyclobacillus acidocaldarius and AFEST from the hyperthermophilic archaeon Archaeoglobus fulgidus. Various carboxylester hydrolases that are not members of the HSL family were found not to be inhibited by compound 7600 under the same experimental conditions. These include nonlipolytic hydrolases such as Torpedo californica acetylcholinesterase and pig liver esterase, as well as lipolytic hydrolases such as human pancreatic lipase, dog gastric lipase, Thermomyces lanuginosus lipase, and Bacillus subtilis LipA. When vinyl esters were used as substrates, the residual activity of HSL, AFEST, and EST2 decreased with an increase in compound 7600 concentration in the incubation mixture. The inhibitor concentration at which the enzyme activity decreased to 50% after incubation for 5 min was 70, 20, and 15 nM with HSL, AFEST, and EST2, respectively. Treating EST2 and AFEST with the inhibitor resulted in an increase in the molecular mass, as established by performing matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis. This increase in the molecular mass, which corresponds approximately to the molecular mass of the inhibitor, indicates that a covalent enzyme-inhibitor complex has been formed. Surface-enhanced laser desorption ionization time-of-flight mass spectrometry analysis of a trypsin digest of AFEST treated with the inhibitor or not treated showed the occurrence of an increase in the molecular masses of the "GESAGG"-containing peptide, which is compatible with the formation of a covalent complex with the inhibitor.     

Mise en évidence d’une flore oxfordienne dans le Sud-Est de la Tunisie : intérêts stratigraphique et paléoécologique

The Late Jurassic - Early Cretaceous series encountered in South-East Tunisia, formely considered globally as Purbecko-Wealdian in age, has the most complete palaeobotanical record of all North-Gondwana. Thanks to the stratigraphical results of the last years and to new field researches, it was possible, for the first time, to evidence an Oxfordian flora at the base of this series (Bir Miteur Formation). This flora, constituted of impressions and axes of Pteridophytes and Gymnosperms, is encountered within different types of plant-bearing levels. The study of deposit facies and sequential analysis of this Oxfordian formation allowed the identification of corresponding sedimentary environments and to infer their palaeoecological meaning. The discovery of exceptional specimens provided new informations about the general morpho-anatomy of Alstaettia, a very peculiar tree fern of a now extinct type. This Oxfordian flora is an interesting key-point in the evolution of North-Gondwanian flora at the Jurassic/Cretaceous transition.     

全球变化联网控制实验的创新性设计:以我国草地生态系统水热要素联网控制实验为例

全球变化已对陆地生态系统结构和功能产生深远影响,明确生态系统对全球变化的响应和适应机制是实现人类对生态系统服务可持续利用的前提.联网实验是理解区域乃至全球尺度生态系统结构功能对全球变化要素响应和适应的重要手段.科学的顶层设计有利于实现联网数据间融合、比对以及分析,进而支撑普适性生态学理论的发展.本文从全球变化联网控制实...     

Sequence context affects the rate of short insertions and deletions in flies and primates

Background  

Insertions and deletions (indels) are an important evolutionary force, making the evolutionary process more efficient and flexible by copying and removing genomic fragments of various lengths instead of rediscovering them by point mutations. As a mutational process, indels are known to be more active in specific sequences (like micro-satellites) but not much is known about the more general and mechanistic effect of sequence context on the insertion and deletion susceptibility of genomic loci.     

The survey of ectoparasites on two species of Triakids (Mustelus mustelus and M. punctulatus) sharks from Tunisian coasts

Between 2018 and 2020, 696 fish belonging to two species of sharks from the Family Triakidae (Mustelus mustelus and M. punctulatus) were collected from the coasts of Tunisia and inspected for parasites. Six copepod taxa (Perissopus dentatus Steenstrup & Lütken, 1861, Eudactylinella alba Wilson, 1932, Kroyeria lineata Van Beneden, 1853, Nesippus orientalis Heller, 1865 and Lernaeopoda galei Krøyer, 1837, Kroyeria sp.), four isopod species (Anilocra physodes (Linnaeus, 1758), Emetha audouini (H. Milne Edwards, 1840), Ceratothoa parallela (Otto, 1828) and Ceratothoa oestroides (Risso, 1816)) and two monogenean species (Erpocotyle sp1. And Erpocotyle sp2) were collected. A large number of global host records was reported, including the occurrence of E. audouini on M. mustelus and of Erpocotyle sp.2 on M. punctulatus. The study of the diversity of parasites per host species revealed that M. mustelus had a higher parasitic richness compared to M. punctulatus. In this study, it was provided for the first records on ectoparasites on Triakidae sharks from Tunisian coasts and their infection indices.     

Identification of an intronic splicing regulatory element involved in auto‐regulation of alternative splicing of SCL33 pre‐mRNA

In Arabidopsis, pre‐mRNAs of serine/arginine‐rich (SR) proteins undergo extensive alternative splicing (AS). However, little is known about the cis‐elements and trans‐acting proteins involved in regulating AS. Using a splicing reporter (GFP–intron–GFP), consisting of the GFP coding sequence interrupted by an alternatively spliced intron of SCL33, we investigated whether cis‐elements within this intron are sufficient for AS, and which SR proteins are necessary for regulated AS. Expression of the splicing reporter in protoplasts faithfully produced all splice variants from the intron, suggesting that cis‐elements required for AS reside within the intron. To determine which SR proteins are responsible for AS, the splicing pattern of the GFP–intron–GFP reporter was investigated in protoplasts of three single and three double mutants of SR genes. These analyses revealed that SCL33 and a closely related paralog, SCL30a, are functionally redundant in generating specific splice variants from this intron. Furthermore, SCL33 protein bound to a conserved sequence in this intron, indicating auto‐regulation of AS. Mutations in four GAAG repeats within the conserved region impaired generation of the same splice variants that are affected in the scl33 scl30a double mutant. In conclusion, we have identified the first intronic cis‐element involved in AS of a plant SR gene, and elucidated a mechanism for auto‐regulation of AS of this intron.