NCKX4 regulates hippocampal Ca²⁺ homeostasis and contributes to contextual memory and anxiety-related behaviours

The K⁺-dependent Na⁺/Ca²⁺-exchanger (NCKX) family comprises five members (NCKX1-5) that serve as key regulators of Ca²⁺ homeostasis. The fourth member, NCKX4, has been implicated in several neuronal processes, including vision, olfaction, and hypothalamic satiety signaling. We recently showed high NCKX4 expression in the mossy fibers of the hippocampus, a brain region essential for learning and memory. In this study, we investigated the impact of NCKX4 loss on Ca²⁺ dynamics in primary hippocampal neurons and assessed hippocampal-dependent behaviour in Nckx4^-/- mice. Using Ca²⁺ imaging, we found that NCKX4 actively contributes to Ca²⁺ homeostasis in hippocampal cells and its loss is not compensated for by other Ca²⁺ transporters. Neurons lacking NCKX4 had significantly reduced rates of total Na⁺/Ca²⁺-exchange activity and showed a decrease in the rate of Ca²⁺ clearance following a depolarization event. Complementary behavioural testing showed deficits in contextual memory formation and recall, as well as reduced anxiety when placed in an open field, in Nckx4^-/- mice. These findings reveal a previously unrecognized role for NCKX4 in regulating hippocampal Ca²⁺ dynamics, and support the hypothesis that NCKX4 expression is mechanistically linked to learning and memory.