Cytoplasmic Determination of Meiotic Spindle Size Revealed by a Unique Inter-Species Germinal Vesicle Transfer Model

Abstract

Spindle sizes are different in diverse species and cell types. In frogs, the meiotic spindle size is positively correlated with the egg cell volume. Across species, relatively small mouse oocytes (70–80 μm) have a relatively large spindle while larger pig oocytes (about 120 μm) have a considerably smaller spindle. In this study we investigated whether species-specific oocyte spindle size was determined by cytoplasmic or nuclear factors. By exchanging the germinal vesicle between mouse and pig oocytes, we obtained two kinds of reconstructed oocytes: one with mouse ooplasm and pig GV (mCy-pGV oocyte), and the other with pig ooplasm and mouse GV (pCy-mGV oocyte). We show that the MII spindle size of the mCy-pGV oocyte is similar to that of the mouse meiotic spindle and significantly larger than that of the pig meiotic spindle. The timing of oocyte maturation also followed that of the species from which the oocyte cytoplasm arose, although some impact of the origin of the GV was observed. These data suggest that spindle size and the timing of meiotic progression are governed by cytoplasmic components rather than cytoplasmic volume and GV materials.

Reduction of nitric oxide level results in maturation promoting factor destabilization during spontaneous meiotic exit from diplotene arrest in rat cumulus oocytes complexes cultured in vitro

“…It starts during fetal life and gets arrested at diplotene stage of prophase‐I at the time of birth (Albertini ; Wang et al . ). The diplotene arrest in these follicular oocytes may last for several months or years in follicular microenvironment depending on the mammalian species (Sirard et al .…”

Reconstruction of mammalian oocytes by germinal vesicle transfer: A systematic review

“…3) IBMX medium containing 5 μg/ml CB and 0.2 μg/ml Dem for 0.5h (11) and then medium (M2) containing 0.2 mM IBMX, 5 μg/ml CB, and 0.2 μg/ml Dem for micromanipulation (11). 4) medium (HTF) with 10% FCS and 50 µg/mL IBMX for 6h incubation and modified medium (mHTF) with 10% FCS supplemented with 7.5 g/ml CB for 15 min at room temperature to interrupt microfilament and rise plasma membrane flexibility before manipulation (33), 5) medium (M2) complemented with 2.5 μ M milrinone for 2 hr to keep the oocytes at the GV stage during micro-manipulation and manipulation drops of medium (M2) with 2.5 μ M milrinone, 5 μ g/ml CB and 0.2 mg/ml Dem (37).…”

“…Micromanipulation can be done in two developmental stages of oocyte including prophase1 and metaphase1 (M1) of meiosis. To imagine the nuclear material (in prophase1oocytes) and meiotic spindle (in MI oocytes) localization, oocytes could be incubated in the enucleating medium with or without 2% sucrose for 1/2 hr (9,37,38). Indirect enucleation (9, 10, 18, 23), direct enucleation (4, 7, 9-12, 17-21, 32, 39, 40) and oocyte fraction (6,38) are three enucleation methods for GV oocytes.…”

“…In the second method, to speed removing and replacing the GV karyoplast and determine the size of it, the process was done "directly" by a tapered enucleation pipette with an inner diameter of about 20 μm (4, 7, 9-12, 17-21, 32, 33, 37, 39, 40). Also, GVs and karyoplasts can be located in the milrinone-medium (M16) for additional manipulation (37).…”

A novel role of Beclin-1, cytokinetic abscission

“…Thus, it does not appear that previous studies support the results of Beclin-1 knock down experiments, abscission failure and nonautophogy defect in meiotic oocytes. However, the improvement may be attributed to an increase in Beclin-1 expression because developmental competency of oocytes treated with rapamycin during 22-42 hours of in vitro maturation (IVM) is comparable to those during 0-42 hours of IVM in porcine oocytes that took 30-48 hours for first polar body extrusion, 6 implying that upregulated Beclin-1 in MI and MII staged porcine oocyte might lead to proper cytokinesis.…”