Placental transfer of maternal calcium (Ca² ) is carried out in vivo by the syncytiotrophoblast layer. Although this process is crucial for fetal development, it remains poorly understood. Cytotrophoblasts isolated from human term placenta undergo spontaneous syncytiotrophoblast-like morphological and biochemical differentiation in vitro and are thought to reflect in vivo syncytiotrophoblast. In the present study, we characterized the Ca² uptake potential and the expression of several Ca² channels by human trophoblasts during differentiation in vitro for up to 6 days. Secretion of hCG (specific differentiation marker) and uptake of Ca² by trophoblasts increased gradually as a function of days in culture. Both hCG secretion and Ca² uptake were maximal on Day 4 and declined on Days 5–6. Expression of the Ca² transporter proteins CaT1 and CaT2 was revealed by reverse transcription-polymerase chain reaction in cytotrophoblasts freshly isolated from human term placenta. In addition, messengers for two L-type Ca² channel isoforms (α_1C and α_1D) were also detected. Levels of CaT1, CaT2, and L-type Ca² channel mRNA increased gradually during culture, reaching a maximum between Days 2 and 3. In contrast to CaT1 and CaT2 expression that declined thereafter to levels observed on Day 1, L-type channel expression decreased by 50% but remained above the expression level of Day 1. Our results indicate that the pattern of CaT1 and CaT2 expression correlates with the Ca² uptake potential along the differentiation of cultured human trophoblasts isolated from term placenta. This correlation provides circumstantial evidence for a role of this family of channels in basal Ca² uptake by the syncytiotrophoblast.