Mechanistic ion channel interactions in red cells of patients with Gărdos channelopathy

In patients with Gárdos channelopathy (p.R352H), an increased concentration of intracellular Ca21 was previously reported. This is a surprising finding because the Gárdos channel (KCa3.1) is a K1 channel. Here, we confirm the increased intracellular Ca21 for patients with the KCa3.1 mutation p.S314P. Furthermore, we provide the concept of KCa3.1 activity resulting in a flickering of red blood cell (RBC) membranepotential, which activates the CaV2.1 channel allowing Ca21 to enter the RBC. Activity of the nonselective cation channel Piezo1 modulates the aforementioned interplay in away that a closed Piezo1 is in favor of the KCa3.1-CaV2.1 interaction. In contrast, Piezo1 openings compromise the membrane potential flickering, thus limiting the activity of CaV2.1. With the compound NS309, we mimic a gain-of-function mutation of KCa3.1. Assessing the RBC Ca21 response by Fluo-4–based flow cytometry and by measuring the membrane potential using the Macey-Bennekou-Egée method, we provide data that support the concept of the KCa3.1/CaV2.1/Piezo1 interplay as a partial explanation for an increased number of high Ca21 RBCs. With the pharmacological inhibition of KCa3.1 (TRAM34 and Senicapoc), CaV2.1 (v-agatoxin TK), and Piezo1 (GsMTx-4), we could project the NS309 behavior of healthy RBCs to the RBCs of Gárdos channelopathy patients.