Functional characterization of ion channels expressed in kidney organoids derived from human induced pluripotent stem...

Kidney organoids derived from human or rodent pluripotent stem cells have glomerular structures and differentiated/polarized nephron segments. Although there is an increasing understanding of the patterns of expression of transcripts and proteins within kidney organoids, there is a paucity of data regarding functional protein expression, in particular on transporters that mediate the vectorial transport of solutes. Using cells derived from kidney organoids, we examined the functional expression of key ion channels that are expressed in distal nephron segments: the large-conductance Ca²⁺-activated K⁺ (BK_Ca) channel, the renal outer medullary K⁺ (ROMK, Kir1.1) channel, and the epithelial Na⁺ channel (ENaC). RNA-sequencing analyses showed that genes encoding the pore-forming subunits of these transporters, and for BK_Ca channels, key accessory subunits, are expressed in kidney organoids. Expression and localization of selected ion channels was confirmed by immunofluorescence microscopy and immunoblot analysis. Electrophysiological analysis showed that BK_Ca and ROMK channels are expressed in different cell populations. These two cell populations also expressed other unidentified Ba²⁺-sensitive K⁺ channels. BK_Ca expression was confirmed at a single channel level, based on its high conductance and voltage dependence of activation. We also found a population of cells expressing amiloride-sensitive ENaC currents. In summary, our results show that human kidney organoids functionally produce key distal nephron K⁺ and Na⁺ channels.NEW & NOTEWORTHY Our results show that human kidney organoids express key K⁺ and Na⁺ channels that are expressed on the apical membranes of cells in the aldosterone-sensitive distal nephron, including the large-conductance Ca²⁺-activated K⁺ channel, renal outer medullary K⁺ channel, and epithelial Na⁺ channel.