{"id":4759,"date":"2023-02-17T07:09:54","date_gmt":"2023-02-17T07:09:54","guid":{"rendered":"http:\/\/ionchanneltargets.com\/resource_library\/neurons-peri-4u-and-dopa-4u-stem-cell-derived-neurons-recorded-on-nanions-patchliner\/"},"modified":"2023-02-17T07:09:54","modified_gmt":"2023-02-17T07:09:54","slug":"2009-an-automatic-electrophysiological-assay-for-the-neuronal-glutamate-transporter-meaac1","status":"publish","type":"resource_library","link":"https:\/\/nanion.com\/zh-hans\/resource_library\/2009-an-automatic-electrophysiological-assay-for-the-neuronal-glutamate-transporter-meaac1\/","title":{"rendered":"2009 &#8211; An automatic electrophysiological assay for the neuronal glutamate transporter mEAAC1"},"content":{"rendered":"<p>A rapid and robust electrophysiological assay based on solid supported membranes (SSM) for the murine neuronal glutamate transporter mEAAC1 is presented. Measurements at different concentrations revealed the EAAC1 specific affinities for l-glutamate (Km\u00a0= 24 \u03bcM), l-aspartate (Km\u00a0= 5 \u03bcM) and Na<sup>+<\/sup>\u00a0(Km\u00a0= 33 mM) and an inhibition constant Ki for dl-threo-\u03b2-benzyloxyaspartic acid (TBOA) of 1 \u03bcM. Inhibition by 3-hydroxy-4,5,6,6a-tetrahydro-3aH-pyrrolo[3,4-d]isoxazole-6-carboxylic acid (HIP-B) was not purely competitive with an IC<sub>50<\/sub>\u00a0of 13 \u03bcM. Experiments using SCN\u2212\u00a0concentration jumps yielded large transient currents in the presence of l-glutamate showing the characteristics of the glutamate-gated anion conductance of EAAC1. Thus, SSM-based electrophysiology allows the analysis of all relevant transport modes of the glutamate transporter on the same sample.K<sup>+<\/sup>\u00a0and Na<sup>+<\/sup>\u00a0gradients could be applied to the transporter. Experiments in the presence and absence of Na<sup>+<\/sup>\u00a0and K<sup>+<\/sup>\u00a0gradients demonstrated that the protein is still able to produce a charge translocation when no internal K<sup>+<\/sup>\u00a0is present. In this case, the signal amplitude is smaller and a lower apparent affinity for l-glutamate of 144 \u03bcM is found.Finally the assay was adapted to a commercial fully automatic system for SSM-based electrophysiology and was validated by determining the substrate affinities and inhibition constants as for the laboratory setup. The combination of automatic function and its ability to monitor all transport modes of EAAC1 make this system an universal tool for industrial drug discovery.<\/p>\n","protected":false},"featured_media":0,"template":"","products_tags":[36],"resource_type":[75],"application_areas":[108],"therapeutic_areas":[],"target_categories":[60],"target_families":[375],"targets":[133],"resource_keyword":[],"class_list":["post-4759","resource_library","type-resource_library","status-publish","hentry","products_tags-surfe2r-n1","resource_type-publication","application_areas-transporters","target_categories-transporters","target_families-symporter","targets-eaat3-slc1a1"],"acf":[],"_links":{"self":[{"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/resource_library\/4759","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/resource_library"}],"about":[{"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/types\/resource_library"}],"wp:attachment":[{"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/media?parent=4759"}],"wp:term":[{"taxonomy":"products_tags","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/products_tags?post=4759"},{"taxonomy":"resource_type","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/resource_type?post=4759"},{"taxonomy":"application_areas","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/application_areas?post=4759"},{"taxonomy":"therapeutic_areas","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/therapeutic_areas?post=4759"},{"taxonomy":"target_categories","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/target_categories?post=4759"},{"taxonomy":"target_families","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/target_families?post=4759"},{"taxonomy":"targets","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/targets?post=4759"},{"taxonomy":"resource_keyword","embeddable":true,"href":"https:\/\/nanion.com\/zh-hans\/wp-json\/wp\/v2\/resource_keyword?post=4759"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}