When: 19.04. – 23.04.2024

Venue: Fletcher Hotel-Restaurant Noordsee-Ameland. Strandweg 42, 9163 GN Nes, The Netherlands.

The Nanion team will be present with a talk

Join us for an insightful talk at session 2 “Ion channels of the red blood cell – function and disorders” on the characterization of red blood cells using automated patch clamp. Our representatives Dr. Nicoletta Murciano (Application Scientist) and Reetta Penttinen (PhD candidate) would be glad to discuss further with you!

Title of talk: “Electrophysiological characterization of red blood cells using the automated patch clamp”

Presenter: Dr. Nicoletta Murciano (Application Scientist)

When: April 20th, 2024 from 11:45-12:00 h CEST

Abstract of the talk:

Introduction: The ion channels Piezo1, K_Ca3.1 (Gardos), and TRPV2 regulate Ca²⁺ and control important functions of red blood cells (RBCs), such as volume adaptation and deformability. These functions are relevant during differentiation and in the circulation, while transporting gases throughout the body. In addition, dysfunctions of these channels can disrupt normal erythropoiesis and may be associated with diseases such as hereditary xerocytosis, Gardos channelopathy, sickle cell anaemia and thalassaemia^1,2,3,4, suggesting their potential as promising therapeutic targets.

Methods:  In this study, we examined the role of Ca²⁺-permeable and Ca²⁺-regulated channels in cultured erythroblasts (EBLs), cultured reticulocytes (retics), healthy and diseased RBCs using manual and high throughput automated patch clamp (APC, SyncroPatch 384). The latter allows for 384 parallel recordings of individual cells, thereby enabling the analysis of heterogeneous cell populations such as native RBC samples based on single cell responses^2,3.

Results: Our results have shown that a significantly higher percentage of EBLs and retics responded to Yoda1 compared to RBCs, indicating a higher activity of Piezo1 in the RBC progenitors, likely due to the different number of receptor copies during differentiation⁵. Only in EBLs, Yoda1-induced current was significantly decreased in the presence of the specific Gardos blocker TRAM-34, suggesting that Gardos is mainly involved at early stages of differentiation. At later stages, calcium influx seemed insufficient to activate Gardos. Piezo1 was mechanically activated in EBLs, and not in retics or RBCs, possibly due to a different membrane tension and/or increased expression of Piezo1 channel in EBLs⁵. Manual patch clamp was used to investigate TRPV2 in RBCs of Sickle cell Anemia (SCA) patients by ∆9THC. SCA patient RBCs were more responsive to acute ∆9THC stimulation compared to control RBCs. These findings may help to interpret recent and very encouraging results on the use of cannabinoid derivatives (CBD) for the treatment of pain associated with the disease⁶.

Conclusion/Discussion:  This work provides valuable insights into the role of Piezo1, Gardos and TRPV2 channels in RBCs differentiation and diseases, and demonstrates that automated patch clamping provides robust assays to investigate Ca²⁺-regulating ion channels in primary cells. The high throughput technology overcomes challenges such as heterogeneous and/or low expression of ion channels across a cell population. This makes the approach suited to detect channelopathies and to screen against diseases related to ion channel dysfunctions in general.

1. Kaestner L et al. (2020). Calcium Signaling, 625-648.
2. Fermo E et al. (2017). Scientific reports, 7(1), 1-13.
3. Rotordam MG et al. (2019). Haematologica, 104(5), e179.
4. Aglialoro F. et al. (2018). Blood (2018) 132 (Supplement 1): 2321.
5. Zarychanski et al. (2012). Blood, 120 (9): 1908–1915.
6. Cherukury et al. (2023). Blood, 141(2):203-208.

About the European Red Cell Society Meeting

This biannual International meeting attracts leading experts in the fields of erythropoiesis, red blood cell physiology and function, and red blood cell related disorders. It is an event that brings together 100+ people from Europe and beyond, and connects the fields of research, diagnostics and clinical implications.