Follicle-Stimulating Hormone Induces a Gap Junction-Dependent Dynamic Change in [cAMP] and Protein Kinase A in Mammalian Oocytes

Abstract

The second messenger cyclic adenosine 5'monophosphate (cAMP) has been implicated in controlling meiotic maturation. To date, there have been no direct measurements of cAMP in living mammalian oocytes. Here, we have used the fluorescently labelled cAMP-dependent protein kinase A (PKA), FlCRhR, to monitor cAMP in mouse oocytes. In cumulus-enclosed oocytes, follicle-stimulating hormone (FSH) stimulated an increase in the oocyte [cAMP] that was prevented by using the gap junction inhibitor, carbenoxolone. The FSH-induced increase in oocyte [cAMP] was suppressed in a time-dependent manner by prior exposure to ATP, while epidermal growth factor had no effect on basal or stimulated levels of cAMP. Finally, using confocal microscopy, we show that the regulatory and catalytic subunits of the microinjected PKA are distributed in a punctate manner with a stronger accumulation in the perinuclear region. On an increase in [cAMP], in response to phosphodiesterase inhibition or FSH, the catalytic subunit diffused throughout the cytoplasm and germinal vesicle, while the regulatory subunit remained anchored. These experiments show that increases in cAMP in ovarian somatic cells are communicated via gap junctions to the oocyte, where it can lead to a redistribution of the catalytic subunit of PKA.

Stops and starts in mammalian oocytes: recent advances in understanding the regulation of meiotic arrest and oocyte maturation

“…One long-standing hypothesis is that cAMP is produced by follicle cells and diffuses through gap junctions to the oocyte (Anderson & Albertini 1976, Bornslaeger & Schultz 1985, Piontkewitz & Dekel 1993, Webb et al 2002b. Gap junctions are present between the cumulus cells and the oocyte (Albertini & Anderson 1974, Anderson & Albertini 1976.…”

“…However, the lack of specific inhibitors against gap junctions in the oocyte has complicated efforts to clarify their possible role in the maintenance of meiotic arrest. For further discussion, see Piontkewitz & Dekel (1993), Webb et al (2002b) and Eppig et al (2004).…”

“…(3) Microinjection of the non-hydrolyzable GTP analog, GTPgS, which activates Gproteins including G s , transiently and dose-dependently maintains meiotic arrest in isolated mouse oocytes (Downs et al 1992). (4) cAMP levels increase in isolated oocytes maintained in meiotic arrest with the phosphodiesterase inhibitors, IBMX or hypoxanthine (Bornslaeger & Schultz 1985, Webb et al 2002b). (5) Cholera toxin, which irreversibly activates G s (De Haan & Hirst 2004), has been shown to delay oocyte maturation in isolated oocytes (Dekel & Beers 1978, Schultz et al 1983b, Urner et al 1983, Vivarelli et al 1983, Downs et al 1992, Grøndahl et al 2000a.…”

Mathematical model of morphogen electrophoresis through gap junctions

“…In contrast, invertebrate gap junctions are made of innexins, a family formerly called OPUS (Barnes, 1994). A variety of small molecules and metabolites are thought to permeate gap junctional paths, including cAMP (Burnside and Collas, 2002;Webb et al, 2002;Bedner et al, 2003), ATP (Bao et al, 2004;Pearson et al, 2005), and Ca ϩϩ (Toyofuku et al, 1998;Blomstrand et al, 1999;Paemeleire et al, 2000).…”

Effects of puberty and gonadotropins on the molecular events controlling meiotic resumption of mouse oocytes

“…The cAMP could be produced either by the oocyte or by cumulus cells. One long-standing hypothesis is that cAMP is produced by follicle cells, and diffuses through gap junctions to the oocyte (Bornslaeger & Schultz 1985, Dekel 1988, Webb et al 2002. However, several lines of evidence support the hypothesis that the oocyte produces its own cAMP through a G-protein-linked receptor in the oocyte plasma membrane that stimulates G s and, subsequently, ADCY (Olsiewski & Beers 1983, Urner et al 1983, Downs et al 1992, Mehlmann 2005.…”