Modulation of calcium signalling in human sperm by nitric oxide

Nitric oxide (NO) generation by nitric oxide synthase (NOS) is implicated in gamete interaction and fertilization. In vitro studies were undertaken to assess the ability of human sperm and cumulus cells (surrounding the oocyte) to generate NO, investigate the mechanism of action of NO, the NO-mediated [Ca²⁺]_i signalling pathways, the possible interaction of NO with progesterone (a product of cumulus) and its impact in the regulation of human sperm functions. Immunofluorescent staining revealed constitutive NOS in human cumulus. DAF-FM diacetate staining demonstrated NO production by cumulus cells. Human sperm exposure to NO donors caused mobilization of stored Ca²⁺ by a mechanism not requiring guanylate cyclase activation but mimicked by S-nitrosoglutathione (GSNO; an S-nitrosylating agent). Dithiothreitol application, to reduce protein –SNO groups, rapidly reversed the actions of NO and GSNO on [Ca²⁺]_i. The effects of NO, GSNO and dithiothreitol on protein S-nitrosylation, assessed using the biotin-switch assay, closely paralleled their actions on [Ca²⁺]_i. Progesterone mobilizes stored Ca²⁺ in human sperm, by a mechanism involving ryanodine receptor (RyR) activation. Pre-treatment with NO reduced the amplitude of the Ca²⁺ response to ryanodine (a RyR agonist), suggesting convergence of the actions of NO and ryanodine. Sperm pre-treatment with NO greatly enhanced the progesterone effect on [Ca²⁺]_i, causing a prolonged increase in flagellar excursion. We conclude that NO regulates mobilization of stored Ca²⁺ in human sperm by protein (possibly RyRs) S-nitrosylation, that this action is synergistic with that of progesterone and that this synergism is potentially highly significant in gamete interactions leading to fertilization.

Name dc:type.qualificationname

d_ph

Level dc:type.qualificationlevel

d_ph

Grantor dc:publisher.institution

University of Birmingham

Year dc:date.issued

2008

Author dc:creator

• Machado de Oliveira, Gisela Sofia Mendes

• Q Science (General)
• QH301 Biology

Repository record dc:identifier.uri

http://etheses.bham.ac.uk//id/eprint/197/2/Decl_IS_MachadoOliveira08PhD.gif

OAI identifier oai:identifier

oai:etheses.bham.ac.uk:197