Cellular and subcellular distribution of the K⁺-dependent Na⁺/Ca²⁺-exchanger subtype 4, NCKX4, in mouse brain

SLC24A4, the gene encoding the K⁺-dependent Na⁺/Ca²⁺-exchanger, NCKX4, is widely expressed in the brain and involved in many neuronal functions. Here we use immunofluorescent staining and electron microscopy to map the expression of the NCKX4 protein across the mouse brain. Our data show that NCKX4 expression is higher in forebrain with particularly intense staining in specific brain regions, including olfactory bulb, ventral pallidum, globus pallidus, hippocampal mossy fibers, substantia nigra, inferior olive and choroid plexus. These areas are closely associated with crucial functions such as learning and memory, reward processing, motor activity, and cerebrospinal fluid production. Co-immunostaining of NCKX4 with marker proteins and immuno-electron microscopy demonstrate that neuronal NCKX4 is not expressed in cell bodies but is confined to distal neuronal processes with preferential expression on the cell surface. In contrast, NCKX4 expression in epithelial cells of the choroid plexus is found exclusively at intracellular sites. We also compared NCKX4 to two other Na⁺/Ca²⁺-exchangers, NCKX2 and NCX1. NCKX4 is the only exchanger expressed in choroid plexus and hippocampal mossy fibers. In the substantia nigra, NCKX4 and NCKX2 show striking overlap while NCX1 is absent. In many other brain regions all three exchangers are present. These data suggest both distinct and redundant roles for these exchangers in different brain regions. This novel information provides valuable insight into the mechanisms underlying the contribution of NCKX4 to various physiological processes associated with different brain regions and suggests the existence of undiscovered roles for NCKX4 in the brain.