{"id":13235,"date":"2024-01-04T14:56:59","date_gmt":"2024-01-04T14:56:59","guid":{"rendered":"https:\/\/www.nanion.de\/?post_type=resource_library&#038;p=13235"},"modified":"2024-01-04T14:56:59","modified_gmt":"2024-01-04T14:56:59","slug":"2023-a-comparative-study-on-the-lysosomal-cation-channel-tmem175-using-automated-whole-cell-patch-clamp-lysosomal-patch-clamp-and-solid-supported-membrane-based-electrophysiology-functional-chara","status":"publish","type":"resource_library","link":"https:\/\/nanion.com\/ja\/resource_library\/2023-a-comparative-study-on-the-lysosomal-cation-channel-tmem175-using-automated-whole-cell-patch-clamp-lysosomal-patch-clamp-and-solid-supported-membrane-based-electrophysiology-functional-chara\/","title":{"rendered":"2023 &#8211; A Comparative Study on the Lysosomal Cation Channel TMEM175 Using Automated Whole-Cell Patch-Clamp, Lysosomal Patch-Clamp, and Solid Supported Membrane-Based Electrophysiology: Functional Characterization and High-Throughput Screening Assay Development"},"content":{"rendered":"<div class=\"html-p\">The lysosomal cation channel TMEM175 is a Parkinson\u2019s disease-related protein and a promising drug target. Unlike whole-cell automated patch-clamp (APC), lysosomal patch-clamp (LPC) facilitates physiological conditions, but is not yet suitable for high-throughput screening (HTS) applications. Here, we apply solid supported membrane-based electrophysiology (SSME), which enables both direct access to lysosomes and high-throughput electrophysiological recordings. In SSME, ion translocation mediated by TMEM175 is stimulated using a concentration gradient at a resting potential of 0 mV. The concentration-dependent K<sup>+<\/sup>\u00a0response exhibited an I\/c curve with two distinct slopes, indicating the existence of two conducting states. We measured H<sup>+<\/sup>\u00a0fluxes with a permeability ratio of P<sub>H<\/sub>\/P<sub>K<\/sub>\u00a0= 48,500, which matches literature findings from patch-clamp studies, validating the SSME approach. Additionally, TMEM175 displayed a high pH dependence. Decreasing cytosolic pH inhibited both K<sup>+<\/sup>\u00a0and H<sup>+<\/sup>\u00a0conductivity of TMEM175. Conversely, lysosomal pH and pH gradients did not have major effects on TMEM175. Finally, we developed HTS assays for drug screening and evaluated tool compounds (4-AP, Zn as inhibitors; DCPIB, arachidonic acid, SC-79 as enhancers) using SSME and APC. Additionally, we recorded EC<sub>50<\/sub>\u00a0data for eight blinded TMEM175 enhancers and compared the results across all three assay technologies, including LPC, discussing their advantages and disadvantages.<\/div>\n","protected":false},"featured_media":0,"template":"","products_tags":[42,36,132],"resource_type":[75],"application_areas":[50,76,108],"therapeutic_areas":[55,143,403],"target_categories":[335,60],"target_families":[],"targets":[399],"resource_keyword":[],"class_list":["post-13235","resource_library","type-resource_library","status-publish","hentry","products_tags-syncropatch-384","products_tags-surfe2r-n1","products_tags-surfe2r-96se","resource_type-publication","application_areas-ion-channels","application_areas-drug-discovery","application_areas-transporters","therapeutic_areas-cns","therapeutic_areas-assay-development-and-validation","therapeutic_areas-lysosomal-diseases","target_categories-intracellular-regulated-ion-channels","target_categories-transporters","targets-tmem175"],"acf":[],"_links":{"self":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_library\/13235","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_library"}],"about":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/types\/resource_library"}],"wp:attachment":[{"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/media?parent=13235"}],"wp:term":[{"taxonomy":"products_tags","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/products_tags?post=13235"},{"taxonomy":"resource_type","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_type?post=13235"},{"taxonomy":"application_areas","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/application_areas?post=13235"},{"taxonomy":"therapeutic_areas","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/therapeutic_areas?post=13235"},{"taxonomy":"target_categories","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/target_categories?post=13235"},{"taxonomy":"target_families","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/target_families?post=13235"},{"taxonomy":"targets","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/targets?post=13235"},{"taxonomy":"resource_keyword","embeddable":true,"href":"https:\/\/nanion.com\/ja\/wp-json\/wp\/v2\/resource_keyword?post=13235"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}