LH, FSH, and chorionic gonadotropin (CG) are comprised of a common α subunit and a hormone-specific β subunit. Using Madin-Darby canine kidney (MDCK) epithelial cells to examine the polarized secretion of human CG/LH, we previously reported that CG and LH were detected in the apical and basolateral compartments, respectively, and the carboxyl terminal end of the CGβ subunit contains a strong apical signal. Here we show that the carboxyl seven amino acids in the LHβ subunit contribute to the basolateral secretion of LH, and an LH chimera bearing the CGβ apical signal is redirected from the basolateral to the apical compartments. Because LH and FSH are synthesized in the same cell, we also compared the secretion polarity of LH with FSH. MDCK cells expressing the FSH dimer displayed an almost equal distribution of protein into the apical and basolateral compartments. Given that the LHβ and CGβ carboxy terminal sequences, which differ from that in the FSHβ subunit, occupy a pivotal role in their polarized behavior, the results support the hypothesis that pituitary exit of LH and FSH occur via different secretion pathways, and are released spatially from the pituitary via different circulatory routes.