Plasma membrane sodium–calcium exchangers are an important component of intracellular calcium homeostasis and electrical conduction. The potassium-dependent or sodium–calcium exchangers NCKX3 (gene SLC24A3) and NCX1 (gene SLC8A1) play a critical role in the transport of intracellular calcium across the cell membrane in exchange for extracellular sodium ions. The transcripts SLC24A3 and SLC8A1 are most abundant in the brain and smooth muscle, but many other tissues, particularly the uterus, aorta, and intestine, also express this gene at lower levels. However, the expression and physiological roles of NCKX3 and NCX1 are largely unknown in the human endometrium during the menstrual cycle. Thus, we examined the endometrial expression of NCKX3 and NCX1 at the transcriptional and translational levels at different phases of the menstrual cycle. Our findings revealed that NCKX1 and NCKX3 were differentially expressed during the menstrual cycle. The endometrial expression of NCKX3 mRNA and protein was enhanced up to 1.5- to 2.5-fold at the early proliferative phase, midproliferative phase, and early secretory phase compared with other phases, whereas a significant alteration in NCX1 expression during the human menstrual cycle was not observed. Subsequent immunohistochemical analysis revealed a large number of uterine NCKX3 and NCX1 proteins in the cytoplasm of luminal and glandular epithelial cells throughout the menstrual cycle. Taken together, these results suggest that human endometrial NCKX3 is abundantly expressed in the endometrium and that NCKX3 may be involved in reproductive function during the menstrual cycle in the human endometrium.