Proceedings of The Physiological Society

University of Liverpool (2002) J Physiol 543P, S053

Communications

Analysis of the non-genomic action of progesterone in human spermatozoa, at the single cell level

C.V. Harper*, J.C. Kirkman-Brown*† and S.J. Publicover*

*School of Biosciences, University of Birmingham, Birmingham B15 2TT and †Birmingham Women's Hospital NHS Trust and Department of Medicine, University of Birmingham, Birmingham, UK

Micromolar concentrations of the steroid hormone progesterone induce calcium influx and acrosome reaction (AR) in human sperm. Calcium influx occurs as an initial transient, lasting up to 4 min, followed by a sustained phase. The study aimed to analyse the effects of progesterone concentration on the calcium transient and subsequent AR.

Highly motile spermatozoa from human donors were harvested using direct swim-up with sEBSS media (0.3 % BSA), and were incubated for 6 h at 6 million cells ml^-1. For single cell analysis, sperm were labelled with Oregon Green Bapta AM, adhered to coverslips in a perfusion chamber and imaged at 25 °C. For fluorimetric studies sperm were labelled with fura-2 AM and imaged at 37 °C; a K_d = 224 nM was assumed.

Progesterone sensitivity was measured across a series of concentrations ranging from 0.3 nM to 30 mM. AR was induced at a concentration of 3 nM (P < 0.025, n = 9, paired t test) and all higher concentrations. The amplitude of the initial calcium transient was analysed using fluorimetry. A dose

Where applicable, experiments conform with Society ethical requirements