Gender effects on the incidence of aneuploidy in mammalian germ cells.

Aneuploidy occurs in 0.3% of newborns, 4% of stillbirths, and more than 35% of all human spontaneous abortions. Human gametogenesis is uniquely and gender-specific susceptible to errors in chromosome segregation. Overall, between 1% and 4% of sperm and as many as 20% of human oocytes have been estimated by molecular cytogenetic analysis to be aneuploid. Maternal age remains the paramount aetiological factor associated with human aneuploidy. The majority of extra chromosomes in trisomic offspring appears to be of maternal origin resulting from nondisjunction of homologous chromosomes during the first meiotic division. Differences in the recombination patterns between male and female meiosis may partly account for the striking gender- and chromosome-specific differences in the genesis of human aneuploidy, especially in aged oocytes. Nondisjunction of entire chromosomes during meiosis I as well as premature separation of sister chromatids or homologues prior to meiotic anaphase can contribute to aneuploidy. During meiosis, checkpoints at meiotic prophase and the spindle checkpoint at M-phase can induce meiotic arrest and/or cell death in case of disturbances in pairing/recombination or spindle attachment of chromosomes. It has been suggested that gender differences in aneuploidy may result from more permissive checkpoints in females than males. Furthermore, age-related loss of chromosome cohesion in oocytes as a cause of aneuploidy may be female-specific. Comparative data about the susceptibility of human male and female germ cells to aneuploidy-causing chemicals is lacking. Increases of aneuploidy frequency in sperm have been shown after exposure to therapeutic drugs, occupational agents and lifestyle factors. Conversely, data on oocyte aneuploidy caused by exogenous agents is limited because of the small numbers of oocytes available for analysis combined with potential maternal age effects. The vast majority of animal studies on aneuploidy induction in germ cells represent cause and effect data. Specific studies designed to evaluate possible gender differences in induction of germ cell aneuploidy have not been found. However, the comparison of rodent data available from different laboratories suggests that oocytes are more sensitive than male germ cells when exposed to chemicals that effect the meiotic spindle. Only recently, in vitro experiments, analyses of transgenic animals and knockdown of expression of meiotic genes have started to address the molecular mechanisms underlying chromosome missegregation in mammalian germ cells whereby striking differences between genders could be shown. Such information is needed to clarify the extent and the mechanisms of gender effects, including possible differential susceptibility to environmental agents.

Mechanisms and chemical induction of aneuploidy in rodent germ cells.

The objective of this review is to suggest that the advances being made in our understanding of the molecular events surrounding chromosome segregation in non-mammalian and somatic cell models be considered when designing experiments for studying aneuploidy in mammalian germ cells. Accurate chromosome segregation requires the temporal control and unique interactions among a vast array of proteins and cellular organelles. Abnormal function and temporal disarray among these, and others to be identified, biochemical reactions and cellular organelles have the potential for predisposing cells to aneuploidy. Although numerous studies have demonstrated that certain chemicals (mainly those that alter microtubule function) can induce aneuploidy in mammalian germ cells, it seems relevant to point out that such data can be influenced by gender, meiotic stage, and time of cell-fixation post-treatment. Additionally, a consensus has not been reached regarding which of several germ cell aneuploidy assays most accurately reflects the human condition. More recent studies have shown that certain kinase, phosphatase, proteasome, and topoisomerase inhibitors can also induce aneuploidy in rodent germ cells. We suggest that molecular approaches be prudently incorporated into mammalian germ cell aneuploidy research in order to eventually understand the causes and mechanisms of human aneuploidy. Such an enormous undertaking would benefit from collaboration among scientists representing several disciplines.

Tamoxifen-induced alterations in meiotic maturation and cytogenetic abnormalities in mouse oocytes and 1-cell zygotes.

Alterations in the rate of oocyte meiotic maturation (OM) and the timing of the metaphase-anaphase transition may predispose oocytes to premature centromere separation (PCS) and aneuploidy. Tamoxifen has the potential for perturbing the rate of OM since it can function as a calcium antagonist by binding to calmodulin and inhibiting the formation of a calcium-calmodulin complex which is needed for activating calmodulin-dependent cAMP phosphodiesterase and initiating OM. The objective of this study was to test the hypothesis that tamoxifen alters the rate of OM and predisposes oocytes to PCS and aneuploidy. Different does of tamoxifen were administered by oral gavage to female mice preovulation. Metaphase II oocyte and 1-cell zygote chromosomes were C-banded and cytogenetically analysed. Tamoxifen treatment resulted in a modest, but significant (p < 0.05), increase in oocytes with PCS. Similar frequencies of hyperploidy and oocytes with unpaired, single chromatids (SC) were found. Metaphase I, diploid and premature anaphase (PA) oocytes were not detected. Hyperploidy, polyploidy, PCS, PA and SC were not detected in zygotes. These data indicate that the levels of tamoxifen-induced PCS found in mouse oocytes did not predispose zygotes to aneuploidy. Tamoxifen did, however, reduce the proportion of females exhibiting oestrus.

Centrosome-specific perturbations during in vitro maturation of mouse oocytes exposed to cocaine.

Previous studies indicating that cocaine may perturb meiotic chromosome segregation in mammalian oocytes prompted an analysis of the effects of cocaine on mouse oocytes matured in vitro under defined exposure conditions. Cumulus-enclosed mouse oocytes were matured in vitro in the continuous presence of cocaine and assessed for meiotic cell cycle progression and centrosome-microtubule organization using a combination of cytogenetic and fluorescence microscopic techniques. Both of these approaches demonstrated that cocaine had little effect on meiotic cell cycle progression to metaphase of meiosis-2 except at the highest dose tested (1000 microg/ml) where progression from metaphase-1 to metaphase-2 was inhibited. Cytogenetic analyses further showed that bivalent segregation was moderately affected and the incidence of premature centromere separation was significantly decreased following cocaine treatment. Under conditions of cocaine exposure, striking changes in meiotic spindle structure and cytoplasmic centrosome organization were observed. A 36% reduction in spindle length was associated with a loss of nonacetylated microtubules and fragmentation of spindle pole centrosomes. Moreover, in oocytes exposed to cocaine during maturation, a doubling in cytoplasmic centrosome number was observed. These results are discussed with respect to the relative roles of chromosomes and centrosomes in establishing and maintaining functional microtubule organization during meiosis in oocytes.

Clomiphene citrate-induced perturbations during meiotic maturation and cytogenetic abnormalities in mouse oocytes in vivo and in vitro.

OBJECTIVE: To determine if clomiphene citrate induces temporal perturbations during meiotic maturation and aneuploidy in mouse oocytes. DESIGN: A controlled dose study involving mouse oocytes in vivo and in vitro. SETTING: Clinical and academic research setting in a university medical center. INTERVENTION(S): Oocytes were obtained after superovulation and from mature follicles. MAIN OUTCOME MEASURE(S): Cytogenetic analysis of oocytes for aneuploidy, premature centromere separation, premature anaphase, and single chromatids, and the frequencies of metaphase I and diploid oocytes. RESULT(S): Clomiphene citrate resulted in a decrease in the number of ovulated oocytes and a significant (P<.05) increase in hyperploidy at 100 mg/kg in vivo. In vitro, 5.0 microg/mL of clomiphene citrate significantly (P<.05) increased hyperploidy and reduced the proportion of metaphase I oocytes. CONCLUSION(S): These findings suggest that clomiphene citrate has the potential for inducing aneuploidy in mouse oocytes both in vivo and in vitro and that the rate of oocyte maturation is altered after clomiphene exposure in vitro. Additional data are needed to support the results of this study.

Sensitivity of mouse oocytes to nicotine-induced perturbations during oocyte meiotic maturation and aneuploidy in vivo and in vitro.

Oocyte meiosis is sensitive to endogenous and exogenous perturbations that upset the temporal sequence of biochemical reactions during oocyte maturation (OM) and predispose oocytes to aneuploidy. Nicotine is an alkaloid that has been reported to disrupt the rate of OM, reduce ovulation and fertilization rates, and increase diploidy. The objective of this study was to test the hypothesis that nicotine perturbs the rate of OM and induces aneuploidy in mouse oocytes in vivo and in vitro. Female mice were given 7.5 IU pregnant mare's serum and either 0, 5.0, 7.5, or 10 mg/kg nicotine in vivo at -3, 0, and +3 h relative to a 5 IU injection of HCG. Oocytes were also cultured in vitro in the presence of 0, 1.0, 5.0, or 10.0 mmol/l nicotine. In vivo, significant (P < 0.05) differences in the proportions of oocytes with premature centromere separation and premature anaphase were found at 10.0 mg/kg nicotine suggesting that the rate of OM was advanced. Also, at this dose the proportion of ovulated oocytes was reduced by approximately 50% relative to controls. In vitro, only non-significant differences were found among the parameters measured. Although nicotine reduced the ovulation rate and perturbed the rate of OM in vivo, these data show that the rate of aneuploidy was not significantly elevated.

Taxol-induced meiotic maturation delay, spindle defects, and aneuploidy in mouse oocytes and zygotes.

To increase our understanding about the potential risks of chemically-induced aneuploidy, more information about the various mechanisms of aneuploidy induction is needed, particularly in germ cells. Most chemicals that induce aneuploidy inhibit microtubule polymerization. However, taxol alters microtubule dynamics by enhancing polymerization and stabilizing the polymer fraction. We tested the hypothesis that taxol induces meiotic delay, spindle defects, and aneuploidy in mouse oocytes and zygotes. Super-ovulated ICR mice received 0 (control), 2.5, 5.0, and 7.5 mg/kg taxol intraperitoneally immediately after HCG. Females were paired (1:1) with males for 17 h after taxol treatment. Mated females were given colchicine 25 h after taxol and their one-cell zygotes were collected 16 h later. Ovulated oocytes from non-mated females were collected 17 h after taxol. Chromosomes were C-banded for cytogenetic analyses. Oocytes were also collected from another group of similarly treated females for in situ chromatin and microtubule analyses. Taxol significantly (p<0.01) enhanced the proportion of oocytes exhibiting parthenogenetic activation, chromosomes displaced from the meiotic spindle, and sister-chromatid separation. Moreover, 7.5 mg/kg taxol significantly (p<0.01) increased the proportions of metaphase I and diploid oocytes and polyploid zygotes. A significant (p<0.01) dose response for taxol-induced hyperploidy in oocytes and zygotes was found. These results support the hypothesis that taxol-induced meiotic delay and spindle defects contribute to aneuploid mouse oocytes and zygotes.

Postovulatory ageing of mouse oocytes in vivo and premature centromere separation and aneuploidy.

Two paramount observations exist regarding aneuploidy in human oocytes: its association with maternal age and its more frequent occurrence during meiosis I. Numerous experimental studies have shown that fertilization of postovulatory aged oocytes is coupled with reproductive failure and cytogenetic aberrations in embryos. However, the basic cytogenetic defect(s) of aged oocytes that causes these abnormalities has not been adequately described. The objective of this study was to test the hypothesis that postovulatory oocyte ageing results in increased frequencies of premature centromere separation (PCS) in metaphase II (MII) oocytes and aneuploidy in zygotes. MII oocytes and one-cell zygotes were collected from superovulated mice at different times after ovulation and fertilization. Chromosomes were C-banded and analyzed for structural and numerical aberrations. The frequencies of PCS in oocytes significantly (p < 0.01) increased with time postovulation: 15 h (15 of 529, 2.8%), 20 h (82 of 627, 13.1%), and 25 h (118 of 502, 23.5%). In zygotes, the frequencies of hyperploidy significantly (p < 0.01) increased with time post-fertilization: 0-4 h (0 of 260), 4-8 h (5 of 212, 2.4%), and 8-12 h (8 of 262, 3.1%). These data support the hypothesis that postovulatory ageing results in elevated levels of PCS in oocytes and of aneuploidy in zygotes. The link between PCS and aneuploidy may be random segregation of sister chromatids during anaphase II.

Electromagnetic fields enhance chemically-induced hyperploidy in mammalian oocytes.

Epidemiological studies suggest that exposure to electromagnetic fields (EMFs) in the environment may be associated with mutagenic changes, but the relation between EMF exposure and aneuploidy has not previously been studied. Environmental EMFs apparently lack the energy necessary to function as aneugens, but the possibility exists that EMFs could influence the incidence of aneuploidy synergistically because EMFs can activate the neuroendocrine system, and ovulation and oocyte meiotic maturation are under neurohormonal control. This hypothesis was tested by examining the effect of EMF exposure on the occurrence of hyperploidy in mouse oocytes induced by vinblastine sulphate (VBS), which was employed as a surrogate for aneugens in the environment. The incidence of hyperploidy in metaphase II oocytes of individual mice following superovulation was determined, and statistical methods were developed to assess whether EMF exposure during oogenesis in the presence of VBS altered the rate of hyperploidy. A significant effect of EMF exposure on VBS-induced hyperploidy was found (P < 0.05). The data suggested that the EMF primarily affected the mice that exhibited a high incidence of VBS-induced hyperploidy. Exposure had no effect on the number of oocytes ovulated nor on the occurrence of hypoploidy. The results support the hypothesis that EMF exposure can promote the occurrence of aneuploidy caused by an aneugen via a mechanism involving the neuroendocrine system.

1,2-propanediol-induced premature centromere separation in mouse oocytes and aneuploidy in one-cell zygotes.

Aneuploidy in germ cells results in reproductive failure and mental and physical disorders in humans. Unfortunately, little is known about the causes and mechanisms of aneuploidy induction. The objective of this study was to test the hypothesis that propylene glycol (1,2-propanediol; PG) induces cytogenetic aberrations in mouse metaphase II (MII) oocytes that predispose zygotes to aneuploidy. Female ICR mice received 7.5 IU eCG and 5.0 IU hCG 48 h later. PG doses of 1300, 2600, and 5200 mg/kg body weight were given 3 h post-hCG; controls received the solvent deionized water. Ovulated oocytes were collected 16 h after administration of PG and processed for cytogenetic analysis. For the one-cell zygote cytogenetic study, females were given PG and paired (1:1) with ICR males for 16 h. Females that mated were given 2 x 10(-3) M colchicine 22 h post-PG, and zygotes were collected 18 h later. PG significantly (p < 0.05) increased both the proportion of MII oocytes with premature centromere separation (PCS) and the proportion of aneuploid one-cell zygotes. These results support the hypothesis that PG-induced PCS in MII oocytes predisposes zygotes to aneuploidy.

Thiabendazole-induced cytogenetic abnormalities in mouse oocytes.

Of the various classes of human genetic disorders, aneuploidy is the most prevalent. Besides its association with maternal age and its predominant origin during maternal meiosis I, little is known about the etiology of aneuploidy. Although various classes of chemicals have been shown to induce aneuploidy in experimental systems, there is no definitive evidence for the role of chemically induced aneuploidy and adverse human health effects, particularly germ cell effects. Thus, it is important to understand the potential of chemicals for inducing aneuploidy in germ cells. There are conflicting data in the literature about the ability of thiabendazole (TBZ) to induce aneuploidy; therefore, we investigated the potential of TBZ for inducing aneuploidy in oocytes. Superovulated ICR female mice were administered 0, 50, 100, or 150 mg/kg TBZ by intraperitoneal injection. The frequencies and percentages of hyperploid oocytes were 0/472 (0), 2/410 (0.5), 6/ 478 (1.3), and 3/427 (0.7) for control, 50, 100, and 150 mg/kg TBZ, respectively. The difference between controls and the 100 mg/kg dose was statistically significant. Also, the proportions of ovulatory mice and the number of oocytes collected per ovulatory female were reduced in the TBZ groups relative to controls. Based on these results, we conclude that TBZ induces a small, but significant increase in the frequency of aneuploid oocytes at toxic doses that also impair ovulation.

Numerical and structural chromosome aberrations induced by etoposide (VP16) during oocyte maturation of mice: transmission to one-cell zygotes and damage to dictyate oocytes.

The antineoplastic drug etoposide (ET) inhibits topoisomerase II (topo II) activity by forming a ternary complex (DNA-ET-topo II). This complex prevents the DNA-strand-rejoining activity of topo II and may result in structural chromosome aberrations. Inhibition of topo II activity may also predispose cells to aneuploidy because this enzyme is needed for removing regions of DNA catenation prior to chromosome segregation. Our objectives were to study the dose response for ET-induced numerical and structural chromosomal aberrations in mouse one-cell zygotes, to compare these data with those obtained from a contemporary metaphase II (MII) oocyte study and to evaluate the sensitivity of dictyate oocytes to ET-induced aneuploidy. ICR female mice were superovulated and injected i.p. with either 6% dimethylsulphoxide (controls) or 20, 40 or 60 mg/kg ET 2 h after human chorionic gonadotrophin (HCG). ICR males were paired (1:1) with females immediately after treatment. After 17 h the males were removed, and after 24 h the females with a vaginal plug were given colchicine. One-cell zygotes were harvested for cytogenetic analysis 17 h after colchicine. The percentages of hyperploid zygotes were 1.1, 5.7, 13.8 and 20.7 and of zygotes with structural aberrations were 2.5, 16.3, 37.7 and 64.7, for control, 20, 40 and 60 mg/kg ET respectively. The differences between each succeeding dose for both structural and numerical aberrations were statistically significant (P < 0.01). When the ET dose response aneuploidy data from zygotes were compared with similar data from a contemporary study involving metaphase II oocytes, the frequencies of hyperploidy were greater in zygotes than in oocytes. We conclude that when ET is administered during the preovulatory phase of meiosis, it is both an aneugen and a clastogen in mouse one-cell zygotes.

Cytogenetic effects of caffeine during in vivo mouse oocyte maturation.

Numerous investigators have studied the reproductive and genetic toxicity of caffeine. Caffeine has also been reported to retard meiotic progression and induce aneuploidy in hamster oocytes in vitro. However, the ability of caffeine to induce aneuploidy in mammalian oocytes in vivo has not been reported. The objective of this study was to test the hypothesis that chemical-induced perturbations during in vivo oocyte meiotic maturation (OM) predispose oocytes to chromosome missegregation. Caffeine inhibits cAMP phosphodiesterase, which is needed for dephosphorylating p34(cdc2) kinase and initiating OM. Following superovulation, a dose of 150 mg/kg caffeine was administered to Institute of Cancer Research (ICR) female mice at various times prior to metaphase I (MI). Ovulated oocytes were collected from the oviducts and processed for cytogenetic analysis. Statistical analyses of the frequencies of hyperploid, MI, diploid, premature centromere separation and single chromatids revealed nonsignificant (P > 0.05) differences between the controls and each of the caffeine groups. Structural chromosome aberrations were not found. Under our experimental conditions, we rejected the hypothesis and concluded that caffeine neither retarded the rate of OM nor increased the incidence of aneuploidy in mouse oocytes. The factors responsible for the different in vivo and in vitro responses require investigation.

Dose-response study and threshold estimation of griseofulvin-induced aneuploidy during female mouse meiosis I and II.

The relative sensitivity of the two meiotic divisions of mouse oogenesis to griseofulvin (GF)-induced aneuploidy was investigated. The first meiotic division was studied by administering GF 4 h after human chorionic gonadotrophin (HCG) injection and analyzing metaphase II (MII) oocytes, whereas study of the second meiotic division involved treating the females 10 h after HCG and analyzing one-cell (1-Cl) zygotes. Data from previous studies have shown that these treatment times represented the most sensitive times for aneuploidy induction during meioses I and II. The statistical analyses of the data showed that the dose-response curves for aneuploidy induction did not differ quantitatively or qualitatively between the two meiotic divisions. The percentages of hyperploid MII oocytes and 1-Cl zygotes were significantly higher (P < 0.001) than in the controls for all doses except 125 mg/kg GF. The highest percentages of hyperploid cells were found after administering 1500 mg/kg GF. However, these percentages were not different (P > 0.05) from those observed after 500 or 1000 mg/kg GF, suggesting saturation of the GF aneuploid target(s). These results suggest that the relative sensitivity to GF-induced aneuploidy between the two meiotic divisions of oogenesis are similar. They also suggest the presence of a lower (125 mg/kg) and an upper 500 mg/kg) threshold for GF-induced aneuploidy.

Important biological variables that can influence the degree of chemical-induced aneuploidy in mammalian oocyte and zygotes.

The ability of certain chemicals to increase the frequency of aneuploidy in mammalian oocytes elicits concern about human health and well-being. This concernment exists because aneuploidy is the most prevalent class of human genetic disorders, and very little information exists about the etiology of aneuploidy. Although there are experimental models for studying aneuploidy in female germ cells and zygotes, these models are still being validated because insufficient information exists about the biological variables that can influence the degree of chemical-induced aneuploidy. In this regard, variables such as dose, solvent, use of gonadotrophins, mode and preovulatory time of chemical administration, time of cell harvest relative to the possibility of chemical-induced meiotic delay, criteria for cytogenetic analysis and data reporting, and an introduction to differences between cell types and sexes are presented. Besides these variables, additional information is needed about the various molecular mechanisms associated with oocyte meiotic maturation and the genesis of aneuploidy. Also, differences between the results from selected chromosome analysis and DNA-hybridization studies are presented. Based upon the various biologic endpoints measured and the differences in cellular physiology and biochemical pathways, agreement among the results from different aneuploidy assays cannot necessarily be expected. To gain further insight into the etiology of aneuploidy in female germ cells, information is needed about the chemical interactions between endogenous and exogenous compounds and those involved with oocyte meiotic maturation.

Synergism between gonadotrophins and vinblastine relative to the frequencies of metaphase I, diploid and aneuploid mouse oocytes.

Ovulated oocytes are used to obtain estimates of germ cell aneuploidy following in vivo treatment with various chemicals. Such studies commonly use exogenous gonadotrophins to increase the number of ovulated oocytes and to induce ovulation during specific time periods. However, the gonadotrophin dosages have not been evaluated relative to the possibility that synergistic and antagonistic interactions may exist between the hormones and the chemical being studied. We, therefore, studied the interactions of different hormone dosages (5 IU pregnant mare's serum (PMS) and 2.5 IU human chorionic gonadotrophin (HCG), 7.5 IU PMS and 5 IU HCG, and 10 IU PMS and 7.5 IU HCG) alone (controls) and in combination with 0.2 mg/kg of the aneugen vinblastine sulfate (VBS) relative to the number of oocytes ovulated, the frequencies of metaphase I (MI), diploid and aneuploid oocytes. The results indicated that the proportions of oocytes ovulated were not significantly (P > 0.05) different among the various control and VBS groups. VBS, however, resulted in significantly (P < 0.01) higher levels of MI oocytes than in controls. In the VBS-treated females, higher frequencies (P < 0.05) of MI oocytes were found in the 7.5 IU PMS and 5 IU HCG and 10 IU PMS and 7.5 IU HCG groups as compared to the 5 IU PMS and 2.5 IU HCG group. The frequencies of diploid oocytes did not differ significantly (P > 0.05) within the control of VBS groups; whereas, when comparisons were made between each control and VBS group, only the difference between the 5 IU PMS and 2.5 IU HCG groups was significant (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Variation of mouse oocyte sensitivity to griseofulvin-induced aneuploidy during the second meiotic division.

The effects of griseofulvin (GF) treatment during the second meiotic division of oogenesis were investigated by cytogenetic analysis of mouse one-cell (1-Cl) zygotes. After determining the duration of fertilization and the second meiotic division, 1500 mg/kg GF were administered to superovulated mice at 10, 12 or 14 h after human chorionic gonadotrophin (HCG) injection. The results showed that GF reduced the frequencies of fertilized oocytes (P < 0.01) and of 1-Cl zygotes that reached the first mitotic metaphase stage (P < 0.001). Aneuploidy was induced regardless of the moment of treatment, but the various treatment times were associated with statistically different (P < 0.05) levels of hyperdiploidy. The maximum frequency of hyperdiploidy (39.6%) occurred when GF was given 10 h after HCG. Polyploid 1-Cl zygotes were significantly induced only at the 10 and 12 h treatment times and their level never exceeded 1.8% of fertilized oocytes. GF also induced abnormalities of chromosome condensation along with centromeric chromosome associations. These results support the conclusion that GF treatment during the second meiotic division induced aneuploidy, polyploidy, and reduced rates of fertilization and zygotic development. Also, the time of chemical treatment influenced the frequencies of these effects.

Susceptibility to vinblastine-induced aneuploidy and preferential chromosome segregation during meiosis I in Robertsonian heterozygous mice.

Chromosome segregation at meiosis I was studied in oocytes and spermatocytes of four different Robertsonian (Rb) heterozygous mouse stocks by cytogenetic analysis of meiotic products. Two Rb heterozygotes spontaneously yielded high frequencies of unbalanced oocytes. In one case, Rb(2.18)Rma, the excess hyperploidy was mainly accounted for by nondisjunction of normal bivalents, suggesting a generalized impairment of meiotic segregation. In each stock, frequencies of hyperploid spermatocytes were either not significantly different or significantly lower than the corresponding frequencies in the oocytes. This confirmed the greater risk of segregational errors in female than in male carriers of the same Rb metacentric. The hypothesis that an error prone system of meiotic segregation, such as the trivalent configuration of single Rb heterozygous oocytes, could be hypersensitive to chemically induced malsegregation was tested by injecting Rb heterozygous females with low doses of vinblastine (VBL). An intraperitoneal injection of 0.06 or 0.09 mg/kg VBL before the first meiotic division significantly increased the spontaneous frequency of hyperploid oocytes, inducing segregational errors of both the trivalent and normal bivalents. The comparison of these data with VBL effects in B6C3F1 mice showed that single Rb heterozygous oocytes are more sensitive to VBL-induced meiotic aneuploidy than oocytes with a standard karyotype. Although segregation distortion has been repeatedly shown in the progeny of Rb heterozygous mice with a significant excess of all telocentric balanced offspring, it has never been demonstrated whether this is a primary event occurring during meiotic segregation or a consequence of selective postconceptional death. In this study, we showed that preferential segregation occurred during female meiosis in all the Rb stocks tested. When segregation distortion was analyzed separately in balanced and unbalanced oocytes, the latter did not show preferential segregation, suggesting that, when the two telocentrics segregated from each other, then the metacentric was randomly directed to the ovum or the polar body.

The influence of postovulatory ageing on the retardation of mouse oocyte maturation and chromosome segregation induced by vinblastine.

Certain compounds can induce ovulated metaphase I (MI) oocytes. To study if these MI oocytes can overcome this blockage, ICR mice were given human chorionic gonadotrophin and 0.6 mg/kg vinblastine sulfate (VBS). Their ovulated oocytes were collected at 17, 19, 21, 23 and 25 h later. The results showed that the frequencies of MI oocytes decreased, the proportions of diploid metaphase II (MII) oocytes increased and the frequencies of hyperploid MII oocytes did not significantly differ (P > 0.05) among the five harvest or postovulatory times. We also found that the proportions of MII oocytes exhibiting premature anaphase II and premature centromere separation increased with postovulatory ageing, and that these frequencies were consistently higher in controls than in the VBS groups. These findings indicate that some of the oocytes blocked in MI can overcome this inhibition and produce primarily diploid MII oocytes. The ultimate fate of ovulated MI and diploid oocytes on aneuploid production hinges upon the formation of a functioning meiotic spindle, which is affected by both dosage and the specific chemical.