{"id":{"repo_id":"birmingham","oai_identifier":"oai:etheses.bham.ac.uk:197"},"canonical_url":"https://search.dev.ndltd.org/etd/birmingham/oai:etheses.bham.ac.uk:197","repository":{"repo_id":"birmingham","name":"University of Birmingham","base_url":"https://etheses.bham.ac.uk/cgi/oai2"},"display":{"title":"Modulation of calcium signalling in human sperm by nitric oxide","abstract":"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 [Ca\\(^{2+}\\)]\\(_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 Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\). The effects of NO, GSNO and dithiothreitol on protein S-nitrosylation, assessed using the biotin-switch assay, closely paralleled their actions on [Ca\\(^{2+}\\)]\\(_i\\). Progesterone mobilizes stored Ca\\(^{2+}\\) in human sperm, by a mechanism involving ryanodine receptor (RyR) activation. Pre-treatment with NO reduced the amplitude of the Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\), causing a prolonged increase in flagellar excursion. We conclude that NO regulates mobilization of stored Ca\\(^{2+}\\) 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.","abstract_html":"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 [Ca<span class=\"etd-inline-math\"><sup>2+</sup></span>]<span class=\"etd-inline-math\"><sub>i</sub></span> 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 Ca<span class=\"etd-inline-math\"><sup>2+</sup></span> 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 [Ca<span class=\"etd-inline-math\"><sup>2+</sup></span>]<span class=\"etd-inline-math\"><sub>i</sub></span>. The effects of NO, GSNO and dithiothreitol on protein S-nitrosylation, assessed using the biotin-switch assay, closely paralleled their actions on [Ca<span class=\"etd-inline-math\"><sup>2+</sup></span>]<span class=\"etd-inline-math\"><sub>i</sub></span>. Progesterone mobilizes stored Ca<span class=\"etd-inline-math\"><sup>2+</sup></span> in human sperm, by a mechanism involving ryanodine receptor (RyR) activation. Pre-treatment with NO reduced the amplitude of the Ca<span class=\"etd-inline-math\"><sup>2+</sup></span> 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 [Ca<span class=\"etd-inline-math\"><sup>2+</sup></span>]<span class=\"etd-inline-math\"><sub>i</sub></span>, causing a prolonged increase in flagellar excursion. We conclude that NO regulates mobilization of stored Ca<span class=\"etd-inline-math\"><sup>2+</sup></span> 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.","abstract_has_math":true,"creators":["Machado de Oliveira, Gisela Sofia Mendes"],"institution":"University of Birmingham","degree_name":"d_ph","degree_level":"d_ph","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2008,"date_issued":"2008-12","date_published":"2008-12","updated_at":"2026-07-24T01:10:46Z","subjects":["Q Science (General)","QH301 Biology"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://etheses.bham.ac.uk//id/eprint/197/2/Decl_IS_MachadoOliveira08PhD.gif","outbound_label":"Repository record","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.sponsor","label":"Sponsor","values":["other"]},{"key":"dc:creator","label":"Author","values":["Machado de Oliveira, Gisela Sofia Mendes"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2008-12"]},{"key":"dc:date.issued","label":"Date","values":["2008-12"]},{"key":"dc:publisher.department","label":"Dc Publisher Department","values":["College of Life & Environmental Sciences","School of Biosciences"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["University of Birmingham"]},{"key":"dc:relation.isreferencedby","label":"Dc Relation Isreferencedby","values":["http://etheses.bham.ac.uk//id/eprint/197/"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["d_ph"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["d_ph"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Q Science (General)","QH301 Biology"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights.embargodate","label":"Dc Rights Embargodate","values":["3000-01-01"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://etheses.bham.ac.uk//id/eprint/197/1/MachadoOliveira08PhD.pdf","http://etheses.bham.ac.uk//id/eprint/197/2/Decl_IS_MachadoOliveira08PhD.gif"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["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 [Ca\\(^{2+}\\)]\\(_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 Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\). The effects of NO, GSNO and dithiothreitol on protein S-nitrosylation, assessed using the biotin-switch assay, closely paralleled their actions on [Ca\\(^{2+}\\)]\\(_i\\). Progesterone mobilizes stored Ca\\(^{2+}\\) in human sperm, by a mechanism involving ryanodine receptor (RyR) activation. Pre-treatment with NO reduced the amplitude of the Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\), causing a prolonged increase in flagellar excursion. We conclude that NO regulates mobilization of stored Ca\\(^{2+}\\) 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."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf","image/gif"]},{"key":"dc:title","label":"Title","values":["Modulation of calcium signalling in human sperm by nitric oxide"]}]}],"canonical_facts":{"dc:contributor.sponsor":["other"],"dc:creator":["Machado de Oliveira, Gisela Sofia Mendes"],"dc:date":["2008-12"],"dc:date.issued":["2008-12"],"dc:description.abstract":["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 [Ca\\(^{2+}\\)]\\(_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 Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\). The effects of NO, GSNO and dithiothreitol on protein S-nitrosylation, assessed using the biotin-switch assay, closely paralleled their actions on [Ca\\(^{2+}\\)]\\(_i\\). Progesterone mobilizes stored Ca\\(^{2+}\\) in human sperm, by a mechanism involving ryanodine receptor (RyR) activation. Pre-treatment with NO reduced the amplitude of the Ca\\(^{2+}\\) 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 [Ca\\(^{2+}\\)]\\(_i\\), causing a prolonged increase in flagellar excursion. We conclude that NO regulates mobilization of stored Ca\\(^{2+}\\) 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."],"dc:format":["application/pdf","image/gif"],"dc:identifier.uri":["http://etheses.bham.ac.uk//id/eprint/197/1/MachadoOliveira08PhD.pdf","http://etheses.bham.ac.uk//id/eprint/197/2/Decl_IS_MachadoOliveira08PhD.gif"],"dc:publisher.department":["College of Life & Environmental Sciences","School of Biosciences"],"dc:publisher.institution":["University of Birmingham"],"dc:relation.isreferencedby":["http://etheses.bham.ac.uk//id/eprint/197/"],"dc:rights.embargodate":["3000-01-01"],"dc:subject":["Q Science (General)","QH301 Biology"],"dc:title":["Modulation of calcium signalling in human sperm by nitric oxide"],"dc:type":["Thesis"],"dc:type.qualificationlevel":["d_ph"],"dc:type.qualificationname":["d_ph"]},"updated_at":"2026-07-24T01:10:46Z"}