Effect of maternal food restriction during gestation on early development of F1 and F2 offspring in the rat-like hamster (Cricetulus triton).

Maternal food restriction (FR) may have strong and long-term effects on body weight, brain and behavior development of offspring. However, it is still not well understood whether such an effect is carried over to the next generation. Our objective was to examine the differences of maternal behavior, body growth, cranial growth and early development of F1 and F2 offspring of rat-like hamsters between a FR group and a control group. Results show that FR has a significant influence on maternal gathering behavior. The body weight of F1 offspring was significantly lower in the food-restricted group compared with that of the control animals, while the body weight of food-restricted F2 offspring was not significantly different from that of the control group. The physical development and neurodevelopment of food-restricted F1 and F2 offspring were significantly delayed compared to the controls. These results suggest that FR in female rat-like hamsters affected negatively the body growth of F1 offspring, and the physical and neurodevelopment of both F1 and F2 offspring. The effect of maternal FR on F2 offspring was smaller than that on F1 offspring. These factors may, in turn, play an important role in the population regulation of this species.

Contribution of chromosomal imbalance to sperm selection and pre-implantation loss in translocation-heterozygous Chinese hamsters.

Chinese hamster stocks with various structurally abnormal chromosomes have been produced by X irradiation. Among these stocks, 18 with various reciprocal translocations were used to investigate the participation of unbalanced gametes in fertilization and the development of unbalanced embryos. Among males as well as females heterozygous for the same translocation, there is no difference in the frequency of each disjunctional class. The participation of chromosomally unbalanced gametes in fertilization was investigated by chromosomal analysis of meiotic cells in heterozygotes for the 18 reciprocal translocations and pronuclei of fertilized ova obtained from crossing these heterozygotes. Compared with the expected frequencies from MII scoring, the frequencies of male pronuclei having a common deficiency of chromosome 1 (1q17-->1q42) or chromosome 3 (3p23-->3q31) decreased significantly in one-cell embryos. However, the frequencies of male pronuclei with other abnormalities were all consistent with those expected from MII scoring. In contrast, the frequencies of female pronuclei with any karyotype including the same abnormalities as those decreased in male pronuclei from the translocation heterozygotes were all consistent with those estimated from MII scoring. These results revealed clearly that most gametes with nullisomies as well as disomies for any chromosomal segments may participate in fertilization, whereas only male gametes nullisomic for certain segments of chromosomes 1 and 3 failed to participate in fertilization. The zygotic selection of chromosomal imbalance was also investigated by direct chromosomal and morphological analyses of preimplantation embryos from crosses between karyotypically normal females and male heterozygotes from the 18 stocks with various reciprocal translocations. These analyses revealed that some embryos were arrested in development at the two-cell stage. The karyotype of these two-cell embryos had a common deficiency in a segment of chromosome 1 or chromosome 2. Embryos with partial monosomy including chromosomes 1, 3, 4 and 5 showed arrested development at four- to eight-cell stages. Among day 4 embryos, some chromosomally unbalanced embryos, mainly with a deficiency of segments of chromosomes 1p, 1q, 2q, 5q, 7q and 8, had fewer blastomeres than karyotypically normal and balanced embryos. The homology between Chinese hamster and mouse chromosomes relating to abnormal embryogenesis at early stages has been partially confirmed from reported maps of chromosomes. The Chinese hamster is useful for further cytogenetic studies during the stages of meiosis and early embryogenesis.

Transmission of chromosomal abnormalities: participation of chromosomally unbalanced gametes in fertilization and early development of unbalanced embryos in the Chinese hamster.

Using 14 Chinese hamster stocks with various reciprocal translocations, chromosomally unbalanced gametes were produced and used to investigate the participation of the unbalanced gametes in fertilization and the development of unbalanced embryos. The selection of chromosomally abnormal gametes during fertilization was investigated by the chromosomal analysis of meiotic cells in heterozygotes for the 14 reciprocal translocations and pronuclei of fertilized ova obtained from crossing these heterozygotes. Compared with the expected frequencies from meiotic metaphase II (MII) scoring, the frequencies of male pronuclei having commonly a deficiency of chromosome 1 (q14-->q42) or chromosome 3 (p23-->q31) in one-cell embryos decreased significantly. However, the frequencies of male pronuclei with other abnormalities were all consistent with those expected from MII scoring. In contrast, the frequencies of female pronuclei with any karyotype including the same ones, as those decreased in male pronuclei from the translocation heterozygotes were all consistent with those estimated from MII scoring. These results suggest that gametes with nullisomies as well as disomies for any chromosomal segments may mostly participate in fertilization, whereas some sperm nullisomic for the specific segments of chromosomes 1 and 3 may fail to fertilize. On the other hand, the zygotic selection of chromosomal imbalance was investigated by direct analyses of pre-implantation embryos from crosses between chromosomally normal females and male heterozygotes from the 14 stocks with various reciprocal translocations. The chromosomal and morphological analysis revealed that some embryos were arrested in development at the two-cell stage and their common abnormality was partial monosomy for chromosome 1 or 2. Embryos with partial monosomy including chromosomes 1, 3 and 4 showed arrested development at four-eight-cell stages. Among day 4 embryos, some chromosomally unbalanced embryos, mainly with a deficiency of other segments, such as chromosomes 1p, 2q, 5q and 8, had fewer blastomeres than karyotypically normal and balanced embryos. The homology between the mouse and the Chinese hamster chromosomes relating to the developmental abnormalities at early stages was partially confirmed.

The timing and sequence of appearance of neuromeres and their derivatives in staged human embryos.

Serial sections of 215 human embryos from Carnegie stages 6-17 were investigated, and 85 graphic reconstructions were prepared. It is proposed that neuromeres be defined as morphologically identifiable transverse subdivisions perpendicular to the longitudinal axis of the embryonic brain and extending onto both sides of the body. It is proposed further that primary neuromeres be redefined as the early-appearing larger divisions of the open neural folds, and secondary neuromeres as the smaller subdivisions that are found both before and after closure of the neural tube. In the light of these definitions, 6 primary neuromeres can be detected in the human brain at stage 9, and a maximum of 16 secondary neuromeres at stage 14. The relationships of the 8 rhombomeres to the associated neural crest, as well as to the pharyngeal arches and the exits of the cranial nerves, are tabulated. Rhombomere 8 (Rh. 8) is intermediate between the more rostral neuromeres and the spinal cord, and its neural relationships indicate that the four occipital somitic pairs do not impress a strictly repetitive pattern as in the spinal cord. Hence, it is suggested that Rh. 8 depends on both intrinsic and extrinsic factors. The synencephalon, parencephalon, and isthmic neuromere can be distinguished in stage 13. In stage 14, rostral and caudal portions of the parencephalon are recognizable, and the full complement of 16 neuromeres is now present. The medial ventricular eminence appears in the diencephalon (D1). A longitudinal organisation begins to be superimposed on the neuromeres, as now indicated by the appearance of the hypothalamic cell cord. This continues in stage 15, when the hypothalamic sulcus develops. That groove, however, is not continuous with the sulcus limitans. In the diencephalon, five longitudinal zones can be discerned. In stage 16, fibre tracts, such as the habenulo-interpeduncular (fasciculus retroflexus) and the tract of the posterior commissure, outline the boundaries of the synencephalon. In stage 17, the tract of the zona limitans intrathalamica (along the marginal ridge in the parencephalon) is an important landmark. This is the last stage in which all the neuromeres can be distinguished. The supramamillary recess becomes defined and is the termination of the sulcus limitans: the alar/basal distinction is inappropriate in the human forebrain. The number and identity of the neuromeres in the human brain, their precise sequence of appearance, and the stages at which they appear are here clarified for the first time. The results of various studies of domains of gene expression indicate that, although in some instances such territories follow the morphological neuromeres, in others they may cross interneuromeric boundaries. It is concluded that the precise morphological study of neuromeres in any given species is necessary for correlative investigations of gene expression.

Down-regulation of topoisomerase I in mammalian cells following ionizing radiation.

Modulators of the DNA unwinding enzyme, topoisomerase I (Topo I), inhibit DNA repair and augment the lethal effects of X-rays and other agents that create breaks in DNA. To investigate the role of Topo I in DNA repair, we examined Topo I activity before and after X-irradiation using confluence-arrested hamster and human cells. Topo I activities were higher in unirradiated neoplastic compared to normal hamster or human cells. Following ionizing radiation, however, enzyme activities were dramatically down-regulated to a greater extent in tumor than in normal cells. The extent of Topo I down-regulation correlated to some extent with survival enhancement in irradiated cells. Enzyme activities were down-regulated within 5 min in hamster and human cells. Recovery of Topo I activities in X-irradiated hamster cells required 12 h, whereas irradiated human cells recovered in only 70 min. Decreased Topo I activity was also noted after UV irradiation. In contrast, Topo I protein and mRNA levels remained unchanged following radiation. Administration of 5 mM 3-aminobenzamide or 0.5 mM PD 128763, inhibitors of poly(ADP-ribose) transferase, prevented Topo I down-regulation in X-irradiated or UV-irradiated human or hamster cells. Thus, decreases in Topo I activity following DNA damage are likely caused by ADP-ribosylation of the enzyme. Down-regulation of Topo I may prevent its binding to single-stranded DNA nicks created by X-irradiation, allowing the DNA repair complex (which is concomitantly activated by ADP-ribosylation) access to these lesions.

Nuclear localization of 68 kDa calmodulin-binding protein is associated with the onset of DNA replication.

In Chinese hamster embryo fibroblast cells, an increase in intracellular calmodulin levels coincided with the nuclear localization of a calmodulin-binding protein of about 68 kDa as the cells progressed from G1 to S phase. When cells were limited from entering into S phase, by omitting insulin a defined medium, intracellular CaM levels did not increase and the 68 kDa calmodulin-binding protein was completely absent from the nuclei. Corresponding to the nuclear localization of calmodulin and the 68 kDa calmodulin-binding protein in S phase cells, there was a dramatic increase in DNA polymerase and thymidine kinase activities in the nuclei of S phase cells as compared to G1 phase cells. In addition, the 68 kDa calmodulin-binding protein, along with calmodulin, is observed to be an integral component of replitase complex responsible for nuclear DNA replication in S phase cells. These observations point to the association of calmodulin and calmodulin-binding protein(s) with the replication machinery responsible for nuclear DNA replication during S phase. A possible regulatory role of these proteins in the onset of DNA replication and cell proliferation is discussed.

Neurogenesis in the basal forebrain in the Chinese hamster (cricetulus griseus). II. Site of neuron origin: morphogenesis of the ventricular ridges.

In the present study the morphogenesis of the ventricular ridges, i.e. the site of origin for neurons in the basal ganglia and various related basal forebrain structures, has been studied in the Chinese hamster with the aid of three-dimensional and graphical reconstructions. The first ridge appears at developmental stage 14 (E 12 1/2). It originates at the level of the torus hemisphaericus, thereby obscuring the basal part of the telodiencephalic boundary. Later on this ridge passes into the medial ventricular ridge. Subsequently, the lateral ventricular ridge arises at stage 16 (E 13 1/2). Initially, both ridges are completely separated by the sulcus subpallii intermedius. During further development, however, this limiting groove fades away, a process starting caudally and gradually proceeding in the rostral direction. Eventually, this process results in the formation of one single ventricular eminence at the second fetal stage (E 18). In the adult stage the ventricular eminence curves around the cerebral stem area. Thickening of the telencephalic walls and local coarctations have considerably reduced the lumen of the lateral ventricle. The preoptic region in the adult must be considered as a derivative of the diencephalic part of the medial ventricular ridge.

Neurogenesis in the basal forebrain of the Chinese hamster (Cricetulus griseus). I. Time of neuron origin.

The time of neuron origin has been determined in the basal ganglia and related basal forebrain structures of the Chinese hamster with the aid of 3H-thymidine autoradiography. Large-celled structures like the globus pallidus, nucleus of the horizontal limb of the diagonal band of Broca as well as large cells in the rostral part of the substantia innominata, in the caudate-putamen-complex and in the olfactory tubercle arise early (E12--E16), whereas medium-sized and small cells in the basal forebrain have a persistent origin over a much longer period. Neuron formation in the basal forebrain persists decrementally until P4. A clear caudorostral spatiotemporal gradient as well as a distinct 'outside-in' gradient have been observed in the caudate-putamen-complex. Medium-sized neurons in the neostriatum and in the nucleus accumbens, generated simultaneously, are usually arranged in scattered clusters. The present data on time of neuron origin strongly support other evidence which points to the conclusion that the nucleus accumbens can be considered as a ventromedial extension of the caudate-putamen-complex.