In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi

作     者：Tanja Pajić Katarina Stevanović Nataša V.Todorović Aleksandar J.Krmpot Miroslav Živić Svetlana Savić-Šević Steva M.Lević Marina Stanić Dejan Pantelić Brana Jelenković Mihailo D.Rabasović 

作者机构：Institute of Physiology and Biochemistry“Ivan Djaja”Faculty of BiologyUniversity of BelgradeStudentski trg 1611158BelgradeSerbia Institute for Biological Research“Siniša Stanković”University of BelgradeNational Institute of the Republic of SerbiaBulevar Despota Stefana 14211000BelgradeSerbia Institute of Physics BelgradeUniversity of BelgradeNational Institute of the Republic of SerbiaPregrevica 11811080BelgradeSerbia University of BelgradeFaculty of AgricultureNemanjina Street 611080BelgradeSerbia Institute for Multidisciplinary ResearchUniversity of BelgradeKneza Višeslava 111030BelgradeSerbia 

出 版 物：《Microsystems & Nanoengineering》 (微系统与纳米工程（英文）)

年 卷 期：2024年第10卷第2期

页      面：251-267页

核心收录：

学科分类：0831[工学-生物医学工程（可授工学、理学、医学学位）] 0710[理学-生物学] 1002[医学-临床医学] 080901[工学-物理电子学] 0809[工学-电子科学与技术（可授工学、理学学位）] 08[工学] 080401[工学-精密仪器及机械] 0804[工学-仪器科学与技术] 0805[工学-材料科学与工程（可授工学、理学学位）] 0803[工学-光学工程] 

基　　金：supported by the Ministry of Science,Technological Development and Innovations,Republic of Serbia[contract numbers:451-03-47/2023-01/200178 and 451-03-47/2023-01/200007] the Project Advanced Biophysical Methods for Soil Targeted Fungi-Based Biocontrol Agents-BioPhysFUN[Grant number 4545]from the Program DEVELOPMENT-Green program of cooperation between science and industry,Science Fund of the Republic of Serbia the Project HEMMAGINERO[Grant number 6066079]from the Program PROMIS,Science Fund of the Republic of Serbia and the Institute of Physics Belgrade through grants from the Ministry of Science,Technological Development and Innovations of the Republic of Serbia 

主　　题：patch recording laser 

摘      要：Studying the membrane physiology of filamentous fungi is key to understanding their interactions with the environment and crucial for developing new therapeutic strategies for disease-causing ***,their plasma membrane has been inaccessible for a micron-sized patch-clamp pipette for pA current recordings due to the rigid chitinous cell ***,we report the first femtosecond IR laser nanosurgery of the cell wall of the filamentous fungi,which enabled patch-clamp measurements on protoplasts released from hyphae.A reproducible and highly precise(diffraction-limited,submicron resolution)method for obtaining viable released protoplasts was *** release from the nanosurgery-generated incisions in the cell wall was achieved from different regions of the *** plasma membrane of the obtained protoplasts formed tight and high-resistance(GΩ)contacts with the recording *** entire nanosurgical procedure followed by the patch-clamp technique could be completed in less than 1 *** to previous studies using heterologously expressed channels,this technique provides the opportunity to identify new ionic currents and to study the properties of the ion channels in the protoplasts of filamentous fungi in their native environment.