Das Projekt "Development of a yeast-based model system for expression of higher eukaryotic K-Channels and their pharmacological analysis" wird vom Umweltbundesamt gefördert und von Universität Bonn, Botanisches Institut und Botanischer Garten durchgeführt. Transmembrane ion channels regulate the movement of ions (particularly Na+, K+, Ca2+ and Cl-) across cellular membranes, and are critical to numerous aspects of neurobiology. Cells express a diverse array of ion-channel proteins that vary widely in their ion selectivity and in their modulation by ligands (such as neurotransmitters) or by membrane voltage. Potassium is the most abundant cellular cation and the imbalance of potassium across the cell membrane is responsible for the maintenance of the membrane potential. Activation of different K+ selective ion channels is essential to control the excitability of nerve and muscle cells. Considerable interest has been focused on the roles of potassium channels in shaping the physiological behaviours of both excitable and non-excitable cells. Pharmacological tools, such as inhibitors have been used to characterize individual classes of channels but for many potassium channels specific blockers are not available. Heterologous expression of ion channel proteins in yeast provides an alternative to animal testing for functional (pharmacological) analysis as well as providing a robust, cell-based system for rapid identification of new lead compounds. K+-channel modulators are valuable pharmacological tools with therapeutic potential.The cloning and characterization of the yeast K+ transport system, and most recently, of the outward rectifying K+channel enabled the generation of yeast mutants lacking those transporters and channels. This advance has made possible new approaches for the analysis of mammalian K+ selective channels by functional complementation of yeast mutants. The development of a yeast-based expression and screening system will play a key role in the development of in-vitro pharmacological tests for chemical and pharmacological agents.The development of a yeast screening systems provides useful tools both for academic and industrial applications in an EC wide strategy.
Das Projekt "Technical viability of a novel device for automated high-volume screening of teratogens based on simultaneous quantitative computer-assisted microscopical evaluation of clonal cell cycle progression and differentiation state" wird vom Umweltbundesamt gefördert und von Stiftung Tierärztliche Hochschule Hannover, Institut für Lebensmitteltoxikologie und Chemische Analytik durchgeführt. The developmental toxicity studies required by the regulatory authorities for registration of new pharmaceuticals and agrochemicals require a large number of animals and considerable effort from skilled technical personnel. The cost of testing a single chemical substance are approximately 250kECU, a factor which severely limits the number of novel agents and process derived by-products which can be evaluated. As a consequence the large number of novel compounds produced in research laboratories and those intermediates of step-wise procedures in the chemical industry, will never be tested by these in vivo procedures. In vitro methods provide an alterative strategy for assessment of developmental toxicity however the most promising methods are time consuming, involve animal use and require skilled personnel. Development of a simple, reliable and rapid screening system is required to facilitate the pharmaceutical and agro-chemical industries in evaluating embryotoxic potential in the large number of existing substances and the new materials which are added to the list each year. EU Directive 86/609 dictates that animals must not be used for testing if a validated, non-animal test is available and the EU BIOTECH Programme has as a goal the development of such assays for the in-vitro detection of teratogens. Previously, members of this consortium have been contracted under the BIOTECH programme to identify in-vitro endpoints for the development of such tests (BIO2CT930471). As the most significant initial steps in the emergence of developmental toxicity is inhibition of cell division coupled with a premature differentiation, coincident evaluation of these endpoints was found to reliably discriminate known teratogens from non-teratogens in structurally related agents using single cells in-vitro. To determine change in differentiation state change in cell morphology was determined using an automated, computer assisted image analysis system which was developed specifically to allow unbiased estimations in large numbers of single cells. The aim of the present proposal is to modify this system in a manner which allows high volume, coincident analysis of proliferation rate and differentiation state; to develop a prototypic system with standard operating procedures; and to demonstrate the validity of the system for routine use in the public and industrial sectors. The proposal fits well with area 7.1.1 of the BIOTECH Work Programme and is within the remit of a Demonstration Project. Prime Contractor: National University of Ireland Dublin, Department of Pharmacology; Dublin; Eire/Ireland.