Essential role of adenosine, adenosine A1 receptors, and ATP-sensitive K+ channels in cerebral ischemic preconditioning.

Preconditioning with sublethal ischemia protects against neuronal damage after subsequent lethal ischemic insults in hippocampal neurons. A pharmacological approach using agonists and antagonists at the adenosine A1 receptor as well as openers and blockers of ATP-sensitive K+ channels has been combined with an analysis of neuronal death and gene expression of subunits of glutamate and gamma-aminobutyric acid receptors, HSP70, c-fos, c-jun, and growth factors. It indicates that the mechanism of ischemic tolerance involves a cascade of events including liberation of adenosine, stimulation of adenosine A1 receptors, and, via these receptors, opening of sulfonylurea-sensitive ATP-sensitive K+ channels.

Pax-6 is essential for lens-specific expression of zeta-crystallin.

Pax-6 is essential for normal eye development and has been implicated as a "master gene" for lens formation in embryogenesis. Guinea pig zeta-crystallin, a taxon-specific enzyme crystallin, achieves high expression specifically in lens through use of an alternative promoter. Here we show that Pax-6 binds a site in this promoter, which is essential for lens-specific expression. Lens and lens-derived cells exhibit a tissue-specific pattern of alternative splicing of Pax-6 transcripts and Pax-6 is expressed in adult lenses and cells that support zeta-crystallin expression. These results suggest that zeta-crystallin is a natural target gene for Pax-6 and that this Pax family member has a direct role in the continuing expression of tissue-specific genes.

Intersubunit contacts made by tryptophan 120 with biotin are essential for both strong biotin binding and biotin-induced tighter subunit association of streptavidin.

In natural streptavidin, tryptophan 120 of each subunit makes contacts with the biotin bound by an adjacent subunit through the dimer-dimer interface. To understand quantitatively the role of tryptophan 120 and its intersubunit communication in the properties of streptavidin, a streptavidin mutant in which tryptophan 120 is converted to phenylalanine was produced and characterized. The streptavidin mutant forms a tetrameric molecule and binds one biotin per subunit, as does natural streptavidin, indicating that the mutation of tryptophan 120 to phenylalanine has no significant effect on the basic properties of streptavidin. However, its biotin-binding affinity was reduced substantially, to approximately 10(8) M-1, indicating that the contact made by tryptophan 120 to biotin has a considerable contribution to the extremely tight biotin binding by streptavidin. The mutant retained bound biotin over a wide pH range or with the addition of urea up to 6 M at neutral pH. However, bound biotin was efficiently released by the addition of excess free biotin due, presumably, to exchange reactions. Electrophoretic analysis revealed that the intersubunit contact made by tryptophan 120 to biotin through the dimer-dimer interface is the major interaction responsible for the biotin-induced, tighter subunit association of streptavidin. In addition, the mutant has weaker subunit association than natural streptavidin even in the absence of biotin, indicating that tryptophan 120 also contributes to the subunit association of tetramers in the absence of biotin.

Non-clathrin-coat protein alpha is a conserved subunit of coatomer and in Saccharomyces cerevisiae is essential for growth.

To complete the molecular characterization of coatomer, the preformed cytosolic complex that is involved in the formation of biosynthetic transport vesicles, we have cloned and characterized the gene for non-clathrin-coat protein alpha (alpha-COP) from Saccharomyces cerevisiae. The derived protein, molecular weight of 135,500, contains four WD-40 repeated motifs (Trp/Asp-containing motifs of approximately 40 amino acids). Disruption of the yeast alpha-COP gene is lethal. Comparison of the DNA-derived primary structure with peptides from bovine alpha-COP shows a striking homology. alpha-COP is localized to coated transport vesicles and coated buds of Golgi membranes derived from CHO cells.

Thrombopoietin, the Mp1 ligand, is essential for full megakaryocyte development.

The development of megakaryocytes (MKs) from their marrow precursors is one of the least understood aspects of hematopoiesis. Current models suggest that early-acting MK colony-stimulating factors, such as interleukin (IL) 3 or c-kit ligand, are required for expansion of hematopoietic progenitors into cells capable of responding to late-acting MK potentiators, including IL-6 and IL-11. Recently, the Mp1 ligand, or thrombopoietin (Tpo), has been shown to display both MK colony-stimulating factor and potentiator activities, at potencies far greater than that of other cytokines. In light of these findings, we tested the hypothesis that Tpo is absolutely necessary for MK development. In this report we demonstrate that neutralizing the biological activity of Tpo eliminates MK formation in response to c-kit ligand, IL-6, and IL-11, alone and in combination, but that these reagents only partially reduce MK formation in the presence of combinations of cytokines including IL-3. However, despite the capacity of IL-3 to support the proliferation and initial stages of MK differentiation, elimination of Tpo prevents the full maturation of IL-3-induced MK. These data indicate that two populations of MK progenitors can be identified: one that is responsive to IL-3 but can fully develop only in the presence of Tpo and a second that is dependent on Tpo for both proliferation and differentiation. Thus, our results strongly suggest that Tpo is the primary regulator of MK development and platelet production.

Cell cycle-regulated nuclear import and export of Cdc47, a protein essential for initiation of DNA replication in budding yeast.

The CDC47 gene was isolated by complementation of a cdc47 temperature-sensitive mutant in Saccharomyces cerevisiae and was shown to encode a predicted polypeptide, Cdc47, of 845 aa. Cdc47 belongs to the Cdc46/Mcm family of proteins, previously shown to be essential for initiation of DNA replication. Using indirect immunofluorescence microscopy and subcellular fractionation techniques, we show that Cdc47 undergoes cell cycle-regulated changes in its subcellular localization. At mitosis, Cdc47 enters the nucleus, where it remains until soon after the initiation of DNA replication, when it is rapidly exported back into the cytoplasm. Cdc47 protein levels do not vary with the cell cycle, but expression of CDC47 and nascent synthesis of Cdc47 occur late in the cell cycle, coinciding with mitosis. Together, these results show that Cdc47 is not only imported into the nucleus at the end of mitosis but is also exported back into the cytoplasm at the beginning of S phase. The observation that Cdc47 is exported from the nucleus at the beginning of S phase has important implications for how initiation of DNA replication is controlled.

Dopamine signaling is essential in cognitive process and motor performance

La dopamina es uno de los principales neurotransmisores del sistema nervioso central y desempeña un papel esencial en diferentes funciones: neuroendocrinas, motivacionales/emocionales y, especialmente, motoras y cognitivas. Las funciones de la dopamina están media-das en gran medida por la estimulación de sus principales receptores D1 (D1R) y D2 (D2R). En esta tesis hemos estudiado el papel que ambos receptores desempeñan en los procesos de apren-dizaje y memoria, así como la regulación que ejercen sobre las neuronas estriatales TH-immunoreactivas (TH-ir) y su posible implicación en la respuesta motora. Para abordar este proyecto hemos utilizado ratones knock-out para el receptor D1 (Drd1a-/-) y D2 (Drd2-/-) ya que no existen compuestos farmacológicos capaces de diferenciar eficazmente entre receptores dopaminérgicos de la misma familia. Además, para el estudio de las neuronas TH-ir realizamos lesiones con 6-OHDA a ratones que posteriormente recibieron un tratamiento crónico con L-DOPA, siendo este el mecanismo más eficaz para inducir la expresión de las neuronas TH-ir objeto de nuestro estudio. Para completar todo ello realizamos test conductuales que evalúan respuesta motora, como el test del cilindro, y diferentes tipos de aprendizaje y me-moria para los cuales utilizamos test específicos. Entre estos test se encuentran: los laberintos de Barnes y Morris para memoria espacial, evitación activa/pasiva y condicionamiento del mie-do para el aprendizaje asociativo, y el reconocimiento de objetos para la memoria de reconoci-miento...

Essential Techniques for Basic Analytical Chemistry

Professor Dwight M. Smith demonstrates proper procedures for accurate preparation and handling of solutions for analytical chemistry, including the analytical balance, volumetric glassware, and avoiding contamination. This video provides an opportunity to learn from a master of the techniques.

Ethnobotanical study of the sages used in traditional Valencian medicine and as essential oil: characterization of an endemic Salvia and its contribution to local development

Seven wild and cultivated Salvia species and two Phlomis species, used traditionally in Valencian medicine to treat a variety of external and internal ailments, were studied. New ethnobotanical data are provided, obtained from semistructured interviews with 34 people in the Valencian area. A seasonal characterization of the essential oil of a wild sage, Salvia blancoana Webb & Heldr. subsp. mariolensis Figuerola, by GC-FID and GC-MS was carried out as a means to ensure quality control of endemic traditional species such as this one, which has been commercialized by local industries. A comparison with the essential oil of Salvia lavandulifolia Vahl subsp.lavandulifolia allowed inclusion of the wild sage within the commercial 'Spanish sage' oil.

Acceptability of pandemic A(H1N1) influenza vaccination by Essential Community Workers in 2010 Alicante (Spain), perceived seriousness and sources of information

Objective. Describe acceptability of pandemic A(H1N1) influenza vaccination by Essential Community Workers (ECWs) from Alicante province (Spain) in January 2010. Evaluate the correlation with attitudes, beliefs, professional advice and information broadcasted by media. Method. In this cross-sectional study, face-to-face interviews were conducted with 742 ECWs to assess their attitudes towards vaccination against the pandemic influenza strain. A multivariable regression model was made to adjust the Odds Ratios (ORs). Results. Some ECWs reported having been vaccinated with seasonal vaccine, 21.5% (95%IC 18.6–24.9); only 15.4% (95%IC 12.8–18.4) with the pandemic one. ECWs vaccinated regularly against seasonal flu (OR 5.1; 95%IC 2.9–9.1), those who considered pandemic influenza as a severe or more serious disease than seasonal flu (OR 3.8; 95%IC 2.1–6.7) and those who never had doubts about vaccine safety (OR 3.7; 95%IC2.1–6.7) had a better acceptance of pandemic vaccine. Finally, 78.7% (95%IC 75.1–81.4) had doubts about pandemic vaccine's effectiveness. Conclusion. The vast amount of information provided by the media did not seem to be decisive to prevent doubts or to improve the acceptability of the vaccine in ECWs. Professional advice should be the focus of interest in future influenza vaccination campaigns. These results should be taken into account by health authorities.

Visual function alterations in essential tremor: A case report

Our purpose is to report alterations in contrast sensitivity function (CSF) and in the magno, parvo and koniocellular visual pathways by means of a multichannel perimeter in case of an essential tremor (ET). A complete evaluation of the visual function was performed in a 69-year old patient, including the analysis of the chromatic discrimination by the Fansworth–Munsell 100 hue test, the measurement of the CSF by the CSV-1000E test, and the detection of potential alteration patterns in the magno, parvo and koniocellular visual pathways by means of a multichannel perimeter. Visual acuity and intraocular pressure (IOP) were within the ranges of normality in both eyes. No abnormalities were detected in the fundoscopic examination and in the optical coherence tomography (OCT) exam. The results of the color vision examination were also within the ranges of normality. A significant decrease in the achromatic CSFs for right eye (RE) and left eye (LE) was detected for all spatial frequencies. The statistical global values provided by the multichannel perimeter confirms that there were significant absolute sensitivity losses compared to the normal pattern in RE. In the LE, only a statistically significant decrease in sensitivity was detected for the blue-yellow (BY) channel. The pattern standard deviation (PSD) values obtained in our patient indicated that there were significant localized losses compared to the normality pattern in the achromatic channel of the RE and in the red-green (RG) channel of the LE. Some color vision alterations may be present in ET that cannot be detected with conventional color vision tests, such as the FM 100 Hue.

The EU essential facilities doctrine. Research Papers in Law, 6/2006

Introduction. The essential facilities doctrine may be seen as the ‘extra weight’ which is put onto the balance, in order to give precedence to the maintenance of competition over the complete contractual freedom of undertakings controlling an important and unique facility. The main purpose of the doctrine is to impose upon such ‘dominant’ undertakings the duty to negotiate and/or give access to the facility, against a reasonable fee, to other undertakings, which cannot pursue their own activity (and therefore will perish) without access to such a facility. This very simple description of the content of the doctrine underlines its limitations: through the imposition of a duty to negotiate or contractual obligations, the rule tends to compensate for the weaknesses of the competitive structure of a market, which are due to the existence of some essential facility. In other words, the doctrine does not by itself provide a definitive solution to the lack of competition, but tends to contractually maintain or even create some competition.1 The doctrine of essential facilities originates in the US antitrust case law of the Circuit and District Courts, but has never been officially acknowledged by the Supreme Court. It has been further developed and hotly debated by scholars in the US, both from a legal and from an economic viewpoint. In the EU, the essential facilities doctrine was openly introduced by the Commission during the early 1990s, but has received only limited and indirect support by the Court of First Instance (the CFI) and the European Court of Justice (the ECJ). It also indirectly inspired the legislation concerning the deregulation of traditional ‘natural’ monopolies. The judicial origin of the doctrine, combined with the hesitant application by the appeal courts, both in the US and the EU, cast uncertainty not only on the precise scope of the doctrine, but also on the issue of its very existence. These questions receive a particular light within the EU context, where the doctrine is called upon to play a different role from its US counterpart. In order to address the above issues, we will first pretend that an EU essential facility doctrine does indeed exist and we shall try to identify the scope and content thereof, through its main applications (Section 1). Subsequently, we will try to answer the question whether such a doctrine should exist at all in the EU (Section 2).