TY  - JOUR
AU  - Pajić, Tanja
AU  - Stevanović, Katarina
AU  - Todorović, Nataša
AU  - Krmpot, Aleksandar J
AU  - Živić, Miroslav
AU  - Savić-Šević, Svetlana
AU  - Lević, Steva M
AU  - Stanić, Marina
AU  - Pantelić, Dejan
AU  - Jelenković, Brana
AU  - Rabasović, Mihailo D
PY  - 2024
UR  - http://radar.ibiss.bg.ac.rs/handle/123456789/6666
AB  - 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 pathogens. However, their plasma membrane has been inaccessible for a micron-sized patch-clamp pipette for pA current recordings due to the rigid chitinous cell wall. Here, 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 developed. Protoplast release from the nanosurgery-generated incisions in the cell wall was achieved from different regions of the hyphae. The plasma membrane of the obtained protoplasts formed tight and high-resistance (GΩ) contacts with the recording pipette. The entire nanosurgical procedure followed by the patch-clamp technique could be completed in less than 1 hour. Compared 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.
PB  - Springer Nature
T2  - Microsystems & Nanoengineering
T1  - In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi
VL  - 10
DO  - 10.1038/s41378-024-00664-x
SP  - 47
ER  - 

@article{
author = "Pajić, Tanja and Stevanović, Katarina and Todorović, Nataša and Krmpot, Aleksandar J and Živić, Miroslav and Savić-Šević, Svetlana and Lević, Steva M and Stanić, Marina and Pantelić, Dejan and Jelenković, Brana and Rabasović, Mihailo D",
year = "2024",
abstract = "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 pathogens. However, their plasma membrane has been inaccessible for a micron-sized patch-clamp pipette for pA current recordings due to the rigid chitinous cell wall. Here, 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 developed. Protoplast release from the nanosurgery-generated incisions in the cell wall was achieved from different regions of the hyphae. The plasma membrane of the obtained protoplasts formed tight and high-resistance (GΩ) contacts with the recording pipette. The entire nanosurgical procedure followed by the patch-clamp technique could be completed in less than 1 hour. Compared 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.",
publisher = "Springer Nature",
journal = "Microsystems & Nanoengineering",
title = "In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi",
volume = "10",
doi = "10.1038/s41378-024-00664-x",
pages = "47"
}

Pajić, T., Stevanović, K., Todorović, N., Krmpot, A. J., Živić, M., Savić-Šević, S., Lević, S. M., Stanić, M., Pantelić, D., Jelenković, B.,& Rabasović, M. D.. (2024). In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi. in Microsystems & Nanoengineering
Springer Nature., 10, 47.
https://doi.org/10.1038/s41378-024-00664-x

Pajić T, Stevanović K, Todorović N, Krmpot AJ, Živić M, Savić-Šević S, Lević SM, Stanić M, Pantelić D, Jelenković B, Rabasović MD. In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi. in Microsystems & Nanoengineering. 2024;10:47.
doi:10.1038/s41378-024-00664-x .

Pajić, Tanja, Stevanović, Katarina, Todorović, Nataša, Krmpot, Aleksandar J, Živić, Miroslav, Savić-Šević, Svetlana, Lević, Steva M, Stanić, Marina, Pantelić, Dejan, Jelenković, Brana, Rabasović, Mihailo D, "In vivo femtosecond laser nanosurgery of the cell wall enabling patch-clamp measurements on filamentous fungi" in Microsystems & Nanoengineering, 10 (2024):47,
https://doi.org/10.1038/s41378-024-00664-x . .