Human spermatozoa are exposed to micromolar concentrations of oestrogen and progesterone within the female reproductive tract (Osman et.al., 1989). In vitro, 3 μM progesterone induces a biphasic Ca²⁺-influx, generating a [Ca²⁺]_i transient followed by a plateau (Kirkman-Brown et.al., 2002). 8-10% of cells generate slow [Ca²⁺]_i oscillations during the plateau phase. Harper et.al. (2004) reported that stimulation with a progesterone gradient induced a monotonic rise in [Ca²⁺]_i, 34 ± 2% of cells generating oscillations. Much less is known about the effect of oestrogen on human spermatozoa [Ca²⁺]_i. Fluorimetric population studies show a simple sustained [Ca²⁺]_i response that inhibits both transient and sustained components of the response to progesterone (Luconi et.al., 1999). However, populations of human spermatozoa are heterogenous, which is reflected in their [Ca²⁺]_i signalling (Lefièvre et.al., 2003). We have utilised single-cell imaging to analyse responses of individual spermatozoa [Ca²⁺]_i to oestrogen (5μM) and progesterone (3.2 μM). Values are means ± S.E.M. Spermatozoa were harvested by swim-up into sEBSS media with 0.3% BSA and incubated (5 h) at 6 million cells ml^-1. [Ca²⁺]_i was monitored in cells loaded with Oregon Green-1 BAPTA and adhered to the base of a perfusion chamber. >90% of spermatozoa responded to oestrogen with an increase in [Ca²⁺]_i,. 60±9% (n=7) of cells showed a plateau response and 9.2±1.0% generated transient and plateau phases, 23±3% generated a transient followed by oscillations and 7±1% generated a transient response only. Following pre-treatment with progesterone approximately 40% of spermatozoa responded with a transient response (sometimes followed by oscillations) as in control preparations. However, the plateau response to oestrogen was completely occluded. In contrast, pre-treatment of cells with oestrogen failed to alter the response to progesterone. Oestrogen, like progesterone, initiates a range of [Ca²⁺]_i signals in human spermatozoa, including oscillations. Although the response to oestrogen was sensitive to prior stimulation with progesterone, we did not observe effects of oestrogen pre-treatment. Further study is required to establish whether progesterone and oestrogen exert their effects through separate or shared transduction systems and how progesterone selectively inhibits the plateau component of the response to oestrogen. All procedures were in accord with local ethical guidelines and the Declaration of Helsinki.