Characterisation of the rheotaxis and thigmotaxis responses of stallion sperm

To navigate their way towards the site of fertilisation in the oviducts, sperm orientate and swim against a flow of mucus (rheotaxis) and while doing so, are guided by the sensation of touch along the epithelial lining of the reproductive tract (thigmotaxis). The aim of this study was to characterise the rheotactic and thigmotactic response of stallion sperm within a microfluidic channel (800 μm wide, 20 μm deep and 58.5 mm in length). Stallion sperm rheotaxis was assessed within the microfluidic channel in response to (i) A range of flow velocities (ii) Varying media viscosity (iii) Sperm distribution across the microfluidic channel and (iv) Sperm hyperactivation and compared to human and ram sperm. Stallion sperm progressed furthest at a velocity range of 10-30 μm/s, with an optimum velocity of 20 μm/s, and a viscosity of 2.5 cP or greater reduced sperm rheotaxis (P<0.05). Stallion sperm that were hyperactivated were unable to exhibit rheotaxis within the microfluidic channel, whereas, both hyperactivated human and ram sperm did exhibit positive rheotaxis. Both hyperactivated and non-hyperactivated human and ram sperm reorietated and swam against the flow, and while non-hyperactivated stallion sperm reoriented and swam against the flow, hyperactivated stallion sperm did not and were swept downstream with the flow. The number of sperm swimming near the microfluidic channel walls was higher than in the centre (P<0.05). This is the first study to illustrate that stallion sperm are rheotactically responsive and increasing viscosity reduces this response. We also demonstrated that sperm are inclined to swim along a surface and uniquely, hyperactivated stallion sperm are non-progressive and do not exhibit a rheotactic response unlike other species. The swimming pattern of hyperactivated stallion sperm against a fluid flow is uncoordinated and non-progressive similar to the erratic pattern traditionally observed in a static environment. This study provides a novel insight into species differences in sperm rheotatic response.