This application is for renewal of a project concerned with signal transduction during oocyte maturation and fertilization. It addresses the long-standing unanswered question of how luteinizing hormone (LH) causes meiotic resumption in mammalian follicle- enclosed oocytes. Recent findings, which establish that gap junctions between the somatic cells of the follicle close rapidly in response to LH, form the basis for the 3 aims of this project: 1) to investigate if LH-induced gap junction closure in the follicle is caused by TACE protease-mediated activation of the EGF receptor, 2) to investigate whether phosphorylation-mediated closure of Cx43 gap junctions in the follicle cells is required for resumption of meiosis in response to LH, and 3) to investigate the role of gap junction closure in regulating cAMP and cGMP in the oocyte in response to LH. Gap junction permeability will be investigated by microinjection of live follicle-enclosed oocytes with fluorescent tracers, and analysis by 2-photon microscopy and fluorescence recovery after photobleaching. These studies will be complemented by investigations of the phosphorylation of connexin 43 on specific regulatory sites. Concentrations of cAMP and cGMP will be monitored in follicle-enclosed oocytes using newly developed optical probes and confocal microscopy. These methods for live tissue microscopy are a major advance, because they avoid the necessity of disrupting the regulatory environment of the follicle in order to investigate its function. Genetically modified mice will be used, in combination with these microscopic approaches, in order to test the hypothesis that proteolytic processing of EGF receptor ligands and phosphorylation of connexin 43 on identified serines are essential for transduction of signals from the somatic cells of the follicle to the oocyte. The proposed research will advance knowledge of a crucial biological process, and establish a basis for future clinical developments, especially in the treatment of infertility. In vitro oocyte maturation is an emerging component of methods for human in vitro fertilization, and understanding of signaling mechanisms that control oocyte meiosis will facilitate such advances.

Public Health Relevance

The proposed research concerns the mechanisms by which hormonal signals cause mammalian oocytes, stored in the ovary, to develop to the fertilizable stage. This research will advance knowledge of a crucial biological process, and establish a basis for future clinical developments, especially in the treatment of infertility.

National Institute of Health (NIH)
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Research Project (R01)
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Study Section
Cellular, Molecular and Integrative Reproduction Study Section (CMIR)
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Ravindranath, Neelakanta
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University of Connecticut
Anatomy/Cell Biology
Schools of Medicine
United States
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Lee, Kyung-Bon; Zhang, Meijia; Sugiura, Koji et al. (2013) Hormonal coordination of natriuretic peptide type C and natriuretic peptide receptor 3 expression in mouse granulosa cells. Biol Reprod 88:42
Robinson, Jerid W; Zhang, Meijia; Shuhaibar, Leia C et al. (2012) Luteinizing hormone reduces the activity of the NPR2 guanylyl cyclase in mouse ovarian follicles, contributing to the cyclic GMP decrease that promotes resumption of meiosis in oocytes. Dev Biol 366:308-16
Norris, Rachael P; Freudzon, Marina; Nikolaev, Viacheslav O et al. (2010) Epidermal growth factor receptor kinase activity is required for gap junction closure and for part of the decrease in ovarian follicle cGMP in response to LH. Reproduction 140:655-62
Jaffe, Laurinda A; Norris, Rachael P; Freudzon, Marina et al. (2009) Microinjection of follicle-enclosed mouse oocytes. Methods Mol Biol 518:157-73
Norris, Rachael P; Ratzan, William J; Freudzon, Marina et al. (2009) Cyclic GMP from the surrounding somatic cells regulates cyclic AMP and meiosis in the mouse oocyte. Development 136:1869-78
Norris, Rachael P; Freudzon, Marina; Mehlmann, Lisa M et al. (2008) Luteinizing hormone causes MAP kinase-dependent phosphorylation and closure of connexin 43 gap junctions in mouse ovarian follicles: one of two paths to meiotic resumption. Development 135:3229-38
Norris, Rachael P; Freudzon, Leon; Freudzon, Marina et al. (2007) A G(s)-linked receptor maintains meiotic arrest in mouse oocytes, but luteinizing hormone does not cause meiotic resumption by terminating receptor-G(s) signaling. Dev Biol 310:240-9
Mehlmann, Lisa M; Kalinowski, Rebecca R; Ross, Lavinia F et al. (2006) Meiotic resumption in response to luteinizing hormone is independent of a Gi family G protein or calcium in the mouse oocyte. Dev Biol 299:345-55
Mehlmann, Lisa M; Jaffe, Laurinda A (2005) SH2 domain-mediated activation of an SRC family kinase is not required to initiate Ca2+ release at fertilization in mouse eggs. Reproduction 129:557-64
Mehlmann, Lisa M (2005) Oocyte-specific expression of Gpr3 is required for the maintenance of meiotic arrest in mouse oocytes. Dev Biol 288:397-404

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