Hypoxanthine Regulation of Oocyte Maturation in the Mouse: Insights Using Hypoxanthine Phosphoribosyltransferase-Deficient Animals1

Downs, Stephen M.

doi:10.1095/biolreprod57.1.54

Cited by 18 publications

(7 citation statements)

References 38 publications

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“…Therefore, FSH may trigger an alternative pathway that can decrease or bypass the high intracellular concentration of cGMP and consequently also cAMP in oocytes. It has been shown that FSH can trigger meiotic resumption in mammalian oocytes inhibited at the GV stage by drugs increasing the intracellular concentration of cAMP, like hypoxanthine [ 40 ], dibutyryl cAMP [ 41 ] or PDE3A inhibitors [ 42 ] via a mechanism involving the protein kinase C and EGFR/MAPK3/1 pathways [ 41 , 43 , 44 ]. Moreover, the cGMP analogues are probably hydrolyzed in the COCs by cGMP-specific phosphodiesterases, the activity of which may be affected by FSH.…”

Section: Discussionmentioning

confidence: 99%

Cyclic guanosine monophosphate does not inhibit gonadotropin-induced activation of mitogen-activated protein kinase 3/1 in pig cumulus-oocyte complexes

Blaha

Němcová

Procházka

2015

Reprod Biol Endocrinol

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BackgroundRecent results indicate a key role for cyclic guanosine monophosphate (cGMP) in the regulation of oocyte meiotic arrest in preovulatory mammalian follicles. The aim of our study was to determine whether the resumption of oocyte meiosis and expansion of cumulus cells in isolated pig cumulus-oocyte complexes (COCs) can be blocked by a high intracellular concentration of cGMP, and whether this effect is mediated by a cGMP-dependent inhibition of mitogen-activated protein kinase 3/1 (MAPK3/1).MethodsThe COCs were isolated from ovaries of slaughtered gilts and cultured in vitro in M199 supplemented with 5% fetal calf serum. The expression levels of the C-type natriuretic peptide (CNP) precursor (NPPC) and its receptor (NPR2) mRNAs during the culture of COCs were determined by real-time RT-PCR. To control the intracellular concentration of cGMP in the COCs, the culture medium was further supplemented with CNP or various concentrations of synthetic cGMP analogues; the concentration of cGMP in COCs was then assessed by ELISA. The effect of the drugs on oocyte maturation was assessed after 24 and 44 h of culture by determining nuclear maturation. The expansion of cumulus cells was assessed by light microscopy and the expression of cumulus expansion-related genes by real-time RT-PCR. A possible effect of cGMP on FSH-induced activation of MAPK3/1 was assessed by immunoblotting the COC proteins with phospho-specific and total anti-Erk1/2 antibodies.ResultsThe COCs expressed NPPC and NPR2, the key components of cGMP synthesis, and produced a large amount of cGMP upon stimulation with exogenous CNP, which lead to a significant (P < 0.05) delay in oocyte meiotic resumption. The COCs also responded to cGMP analogues by inhibiting the resumption of oocyte meiosis. The inhibitory effect of cGMP on meiotic resumption was reversed by stimulating the COCs with FSH. However, high concentration of intracellular cGMP was not able to suppress FSH-induced activation of MAPK3/1 in cumulus cells, cumulus expansion and expression of expansion-related genes (P > 0.05).ConclusionsThe findings of this study indicate that high cGMP concentrations inhibit the maturation of pig oocytes in vitro but the inhibitory mechanism does not involve the suppression of MAPK3/1 activation in cumulus cells.

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Section: Discussionmentioning

confidence: 99%

Cyclic guanosine monophosphate does not inhibit gonadotropin-induced activation of mitogen-activated protein kinase 3/1 in pig cumulus-oocyte complexes

Blaha

Němcová

Procházka

2015

Reprod Biol Endocrinol

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“…Additionally, many of the experiments showing successful meiotic induction during coculture have been carried out using hypoxanthine as the meiotic inhibitory agent. Studies demonstrating active uptake of this purine base by oocyte-CC complexes (Downs et al, 1986;Downs, 1994Downs, , 1997 raise the possibility that its sequestration by CCs reduces the inhibitory potency of the medium during coculture, leading to higher maturation percentages. In fact, one possible explanation for the lack of an inhibitory effect of conditioned hypoxanthinesupplemented medium on the maturation of CEO is that consumption of hypoxanthine by CEO during conditioning offset any negative influence that was generated.…”

Section: Discussionmentioning

confidence: 99%

Differential response of cumulus cell-enclosed and denuded mouse oocytes in a meiotic induction model system

et al. 2006

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In this study we have examined the effects of denuded oocyte coculture with dissociated cumulus cells (CC) or intact oocyte-CC complexes on meiotic resumption. When denuded oocytes (DO) or cumulus cell-enclosed oocytes (CEO) were cultured in 40-microl drops of medium under oil, and held in meiotic arrest with 4 mM hypoxanthine plus 25 microM dbcAMP, they underwent germinal vesicle breakdown (GVB) at similar frequencies (34%-35%). Coculture of DO with complexes or dissociated CCs stimulated maturation (50% and 61% GVB, respectively), with no effect of DO on maturation of cocultured CEO (32% GVB). This coculture effect was increased with the number of CCs added to the culture drop. When either glucose or glutamine was eliminated from the medium, no meiotic induction resulted from cocultured CCs. When CEO were cultured alone in microdrops, increasing their number from 10 to 50 significantly lowered the percentage resuming maturation, an effect also reduced by removing glucose and/or glutamine from the medium. This effect was not observed with DO. When inhibitory medium was conditioned overnight with complexes, subsequent culture with DO led to higher maturation percentages than culture in unconditioned medium; however, when CEO were cultured in conditioned medium, there was either no effect or increased inhibition of maturation. Assay of glucose and pyruvate in spent medium showed that DO cultured alone consumed glucose and pyruvate, but under CC coculture conditions more glucose was consumed and significant amounts of pyruvate accumulated in the medium, changes that led to an increase in the maturation of DO. Further experiments showed that DO were more sensitive than CEO to the meiosis-inducing effect of pyruvate. These results demonstrate different responsiveness of DO and CEO to coculture conditions and question the physiological relevance of denuded oocyte/CC coculture to study meiotic induction.

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“…Moreover, Acacb −/− mice show normal fertility, suggesting that there is some redundancy of function wherein ACACA produces enough malonyl CoA to dampen FAO and premature meiotic resumption. The line of Acacb −/− mice used in this study overexpress hypoxanthine‐guanine phosphoribosyltransferase (Abu‐Elheiga et al, ), but this cannot account for the effects seen on meiosis and FAO because increased levels of this enzyme would be expected to produce effects opposite to those observed (Downs, ).…”

Section: Discussionmentioning

confidence: 99%

Acetyl CoA carboxylase inactivation and meiotic maturation in mouse oocytes

Valsangkar

Downs

2015

Mol. Reprod. Dev.

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In mouse oocytes, meiotic induction by pharmacological activation of PRKA (adenosine monophosphate-activated protein kinase; formerly known as AMPK) or by hormones depends on stimulation of fatty acid oxidation (FAO). PRKA stimulates FAO by phosphorylating and inactivating acetyl CoA carboxylase (ACAC; formerly ACC), leading to decreased malonyl CoA levels and augmenting fatty-acid transport into mitochondria. We investigated a role for ACAC inactivation in meiotic resumption by testing the effect of two ACAC inhibitors, CP-640186 and Soraphen A, on mouse oocytes maintained in meiotic arrest in vitro. These inhibitors significantly stimulated the resumption of meiosis in arrested cumulus cell-enclosed oocytes, denuded oocytes, and follicle-enclosed oocytes. This stimulation was accompanied by an increase in FAO. Etomoxir, a malonyl CoA analogue, prevented meiotic resumption as well as the increase in FAO induced by ACAC inhibition. Citrate, an ACAC activator, and CBM-301106, an inhibitor of malonyl CoA decarboxylase, which converts malonyl CoA to acetyl CoA, suppressed both meiotic induction and FAO induced by follicle-stimulating hormone, presumably by maintaining elevated malonyl CoA levels. Mouse oocyte-cumulus cell complexes contain both isoforms of ACAC (ACACA and ACACB); when wild-type and Acacb(-/-) oocytes characteristics were compared, we found that these single-knockout oocytes showed a significantly higher FAO level and a reduced ability to maintain meiotic arrest, resulting in higher rates of germinal vesicle breakdown. Collectively, these data support the model that ACAC inactivation contributes to the maturation-promoting activity of PRKA through stimulation of FAO.

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Hypoxanthine Regulation of Oocyte Maturation in the Mouse: Insights Using Hypoxanthine Phosphoribosyltransferase-Deficient Animals1

Cited by 18 publications

References 38 publications

Cyclic guanosine monophosphate does not inhibit gonadotropin-induced activation of mitogen-activated protein kinase 3/1 in pig cumulus-oocyte complexes

Cyclic guanosine monophosphate does not inhibit gonadotropin-induced activation of mitogen-activated protein kinase 3/1 in pig cumulus-oocyte complexes

Differential response of cumulus cell-enclosed and denuded mouse oocytes in a meiotic induction model system

Acetyl CoA carboxylase inactivation and meiotic maturation in mouse oocytes

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