[Ca²⁺]_i rise at in vitro maturation in bovine cumulus–oocyte complexes

Abstract

An intracellular calcium ([Ca(2+)]i) rise has been described in cumulus-oocyte complexes (COCs) following luteinizing hormone (LH) exposure. Together with cAMP, Ca(2+) is a candidate signal for resumption of meiosis. Here, we analyzed if the most common hormones involved in oocyte maturation can induce the same Ca(2+) signal. In addition, we characterized the source of this signal. Immature, in vitro-matured, and roscovitine-meiotically arrested COCs were loaded with Fluo-4 AM, stimulated with hormones/growth factors, and tested for [Ca(2+)](i) variations in cumulus cells. Reagents known to inhibit or stimulate [Ca(2+)](i) rises were used to characterize these [Ca(2+)](i) dynamics. Finally, expression of LH receptors (LHRs) in COCs was analyzed by immunofluorescence. In immature COCs, follicle-stimulating hormone (FSH) elicited a single [Ca(2+)](i) rise that was higher than those induced by LH and growth hormone (GH), whereas epithelial growth factor failed to induce any changes in [Ca(2+)](i). The [Ca(2+)](i) rise induced by FSH was higher in immature COCs; was reduced in roscovitine-arrested, immature COCs; and was negligible in gonadotropin-induced, in vitro-matured COCs. In the case of spontaneous- and GH-matured COCs, however, FSH stimulation caused a lower [Ca(2+)](i) rise. The hormone-induced [Ca(2+)](i) rise was due to: (i) external Ca(2+) entry; (ii) intercellular communication; and (iii) intracellular Ca(2+) stores. Immunofluorescence revealed that LHRs were expressed throughout the cumulus cells. The above results show that: (i) gonadotropins and GH cause a [Ca(2+)](i) rise in cumulus cells; (ii) this [Ca(2+)](i) rise results from extra-, inter-, and intra-cellular cumulative Ca(2+) fluxes; and (iii) LHRs are distributed on either outer or inner cumulus cells.

Electrophysiology and Fluorescence Spectroscopy Approaches for Evaluating Gamete and Embryo Functionality in Animals and Humans

“…[Ca 2+ ] i rises have been described in cumulus–oocyte complexes (COCs) following luteinizing hormone (LH) exposure and candidate as a signal for resumption of meiosis in the mammalian oocyte [ 113 , 114 , 115 ]. However, the role of Ca 2+ entry through ion channels on the plasma membrane has been described only in a few mammalian species.…”

Short term culture with cAMP modulators before vitrification significantly improve actin integrity in bovine oocytes

“…In many cell types, the activation of hormones and growth factors receptors causes an increase in intracellular free calcium (reviewed by (Williamson and Monck, 1990)). A calcium rise was described in bovine COCs after their exposure to IVM hormones (Silvestre et al, 2012). In vivo, the LH surge causes a rise in calcium free that starts in cumulus cells (Gudermann et al, 1992;Mattioli et al, 1998).…”

The Calcium-Sensing Receptor and the Reproductive System

“…Activation of MAPKs triggers degradation of cAMP, allowing oocyte maturation to proceed. Ca 2+ plays a role in controlling LH-induced as well as spontaneous oocyte maturation, possibly by modulating intracytoplasmic cAMP concentrations via Ca 2+ -sensitive AC (Silvestre et al, 2011, 2012; Chen et al, 2013). By termination of meiosis I, the diploid primary oocyte generates two haploid daughter cells.…”