The events during migration of human sperm through the female reproductive tract remain almost completely uncharacterised. Not only are sperm known to be sensitive to products of the female tract but animal data suggests the female tract alters gene expression after exposure to sperm (1), implying two-way communication. We have used single-cell fluorescence imaging to observe calcium signalling in both human sperm and human reproductive tract cells upon initial contact and during sperm adhesion and release. Explants and primary cell lines were prepared from donated human reproductive tract tissue removed during surgery (2). An immortalised oviductal cell line (OE E6/E7) has been used as an internal standard (3). Human sperm were harvested via a modified swim-up technique. Briefly, sperm were selected by their ability to migrate through a viscous medium (~ 140 centipoise) into sEBSS media, then washed and resuspended in sEBSS media containing 0.3% FBS (charcoal stripped) at a concentration of 1 x106 cells per ml. Sperm were incubated for at least 3 hours at 37oC 6% CO2 before use. To investigate whether there was rapid cell signalling occurring in human female tract cells upon exposure to sperm, tract cells were labelled with 7.6 µM Calcium Green-1, AM for 1h at 37oC 6% CO2 to monitor intracellular calcium levels. Sperm were labelled with 5 µM Syto64, a red fluorescent nuclear dye, to allow tracking of sperm movement and contact with cells. Raw intensity values were imported into Microsoft Excel and normalised. Values are percentage change in fluorescence ±S.E.M. Initial data with tract cells show that explants demonstrated transient responses of 9±5%(n=3) and an average peak duration of ~ 20s. Primary tract culture responses were 15±10% (isthmus, n=3), 7±4% (ampulla, n=3), both with similar peak durations of ~ 30s. OE E6/E7 cells had larger responses of 22±10%(n=6) and generated transients with an average peak duration of ~ 35s. Human sperm bound to and interacted with cells from all tract zones. Sperm swimming patterns appear to be modified when in the presence of reproductive tract cells. We then investigated whether calcium signals occurred within sperm whilst in contact with tract cells. Sperm were loaded with 7.6 µM Calcium Green-1, AM for 1h at 37oC 6% CO2. Detailed observations of signals in attached cells and responses to progesterone exposure are currently being examined. Preliminary evidence suggests that certain calcium signals may relate to observed motility changes. Research into these processes may lead to a better understanding of the basic reproductive biology, which will allow the development of rational treatments and diagnostics. All procedures were in accordance with Local Ethics Committee guidelines and patient and donors provided informed consent.