The aim of the present study was to investigate the interaction of estradiol-17β-bovine serum albumin (E2-BSA) and calcitropic hormones, such as parathyroid hormone, calcitonin, and vitamin D, in regulation of Ca2⁢ uptake in primary cultured renal proximal tubule cells. Statistically significant increase in Ca2⁢ uptake was found from 2 hours after E2-BSA (10⁣9 M) treatment, while estradiol-17β (10⁣9 M) did not affect. Treatment of the cells with E2-BSA (10⁣9 M) together with parathyroid hormone (PTH) (10⁣8 M), vitamin D (10⁣8 M), or calcitonin (10⁣8 M) significantly stimulated Ca2⁢ uptake by 32.50%, 29.30%, or 27.75%, respectively, compared with the control. However, calcitropic hormones did not exhibit any synergistic effect on the E2-BSA-induced stimulation. E2-BSA significantly increased cAMP generation and PKC activity. The stimulatory effect of cotreatment of E2-BSA and PTH or vitamin D was blocked by SQ22536 (an adenylate cyclase inhibitor) and staurosporine (a PKC inhibitor), but the effect of cotreatment of E2-BSA and calcitonin was not blocked. Furthermore, 8-Br-cAMP and TPA (an artificial PKC promoter) increased Ca2⁢ uptake by 25.51% and 16.47%, respectively, compared with the control. In conclusion, E2-BSA combined with calcitropic hormones regulated Ca2⁢ uptake partially via cAMP and PKC-dependent mechanisms in renal proximal tubule cells.