The effect of ethanol upon early development in mice and rats. XV. The late effect of chronic preconceptional and preimplantational consumption of beer and wine in rats.

The late effect upon fetal development of chronic preconceptional and preimplantational consumption of beer and wine was investigated in rats. Both beverages applied showed a noxious fetal effect, manifested by an increased number of resorptions and the partial retardation of skeletal development. The correlation of experimental blastopathy with late fetal effects is discussed.

The effect of ethanol upon early development in mice and rats. XIX. The late effect of acute preimplantation intoxication with beer and cognac, on the background of chronic consumption, in mice.

The effect upon late foetal development of acute preimplantation intoxication with beer and cognac, on the background of chronic consumption was investigated in mice (controlled on day 19 of pregnancy), by using the following criteria: mean number of embryos/animal, number of resorptions, mean foetal and placental weight, changes of internal organs (Wilson's sections), skeletal development. The results showed that both beverages applied had a noxious foetal effect, manifested by: a decrease of the mean number of embryos/animal (cognac), an increased late mortality (beer), a decrease of the mean foetal and placental weight (both beverages) and the presence of some structural anomalies (especially after beer consumption). The more marked effect of beer and the correlation of late effects with preimplantation changes are discussed.

The effect of ethanol upon early development in mice and rats. XVI. In vivo effect of acute preimplantation intoxication with beer and cognac on the background of chronic consumption.

The effect of acute preimplantation intoxication with beer and cognac on the background of chronic consumption was investigated in mice (controlled on day 4 of pregnancy) by using the following criteria: mean number of embryos/animal; oviductal-uterine migration of embryos; developmental rate; number of pathologically modified embryos. It resulted that both beverages used have a noxious effect upon preimplantation development: retardation of development, slowing down of oviductal-uterine migration and presence of pathologically modified embryos. This effect was more marked than the effect of chronic administration of these beverages in rats (1) and than the effect of ethanol administered similarly in mice (2). This difference may be due to a species difference of susceptibility, to the additional acute intoxication (for 1) or to the various congeners present in the beverages used (for 2).

The effect of ethanol upon early development in mice and rats. XVIII. In vivo effect of acute preimplantation intoxication with beer and cognac on the background of chronic biparental intake (in mice).

The effect of acute preimplantation intoxication with beer and cognac, on the background of chronic biparental consumption, was investigated in mice (controlled on day 4 of pregnancy), by using the following criteria: mean number of embryos/animal; oviductal-uterine migration of embryos; developmental rate; number of pathologically modified embryos. It resulted that both beverages used have a noxious effect upon preimplantation development, producing: retardation of oviductal-uterine migration and the increase of the number of pathologically modified embryos. Although the experimental model applied represents a maximal schedule of beverage administration (chronic biparental + acute), the effects were not enhanced as compared with previous results obtained by a less "forced" administration (chronic maternal + acute).

The effect of ethanol upon early development in mice and rats. XXII. The effect of chronic consumption of nonalcoholic beer upon preimplantation development in rats.

The effect upon preimplantation development of chronic consumption of nonalcoholic beer was investigated in rats (controlled on day 5 of pregnancy) by using the following criteria: mean number of embryos/animal, oviductal-uterine migration of embryos, developmental rate and number of pathologically modified embryos. It resulted that this beverage had a noxious effect (less marked than normal beer) upon preimplantation development, manifested by the decrease of the mean number of embryos/animal and the presence of abnormal embryos. These results suggest a possible action of various congeners present in this beverage, which potentiate the noxious effect of ethanol in the case of normal beer.

The effect of ethanol upon early development in mice and rats. XX. The effect of chronic biparental beer intake in mice upon early implantation events.

The effect of chronic biparental beer intake upon early implantation in mice was investigated. In order to study the changes taking place in the blastocyst and locally in the uterus at the site of blastocyst attachment, the uteri were tested for the presence of Evans blue areas (Psychoyos test), histochemically for alkaline phosphatase in the stroma and histologically examined. It resulted that the beverage used has a noxious effect upon the implantation process, manifested by: delayed attachment of blastocysts, decreased intensity or absence of decidual reaction, the presence of dead cells in the decidua, the desynchronisation of the implantation process and the diminution or absence of alkaline phosphatase reaction. These results may explain some of our previous observations regarding the late (fetal) effect of preconceptional intake of beer.

The effect of ethanol upon early development in mice and rats. XXIII. The effect of indomethacin on beer-induced disturbances of preimplantation development in rats.

The effect of Indomethacin on beer-induced modifications of preimplantation development was investigated in rats (control on day 5 of pregnancy), using the following criteria: the mean number of embryos/animal, topographical distribution of embryos, the developmental stage attained, the appearance of pathological forms, the mean cell number/embryo. It resulted that previous administration of Indomethacin antagonized the main deleterious effect of repeated acute administration of beer in the preimplantation period, supporting the hypothesis of prostaglandins (PG) being involved in the pathogenetic action of this alcoholic beverage.

On the prenatal noxious effects of trypan blue and of a related azo dye.

Since 1948 trypan blue has been a well-known and extensively used experimental teratogen, belonging to the group of azo dyes. Chemically, trypan blue consists of a biphenyl molecule (0-tolidine or benzidine) combined by means of azo linkages with two molecules of a substituted naphthalene. Between 1987-89 the effect of the replacement of the biphenyl molecule by a molecule of p,p'-diaminobenzanilide upon the prenatal noxious action of trypan blue has been controlled. Investigations were carried out on three species: chick embryos, albino rats and albino mice. In the species used, the replacement annihilates the teratogenic properties of the dye, with the persistence of some embryotoxic effects. On the other hand, the control of o-tolidine and of p,p'-diaminobenzanilide revealed that no one had teratogenic properties (only some embryotoxic effect, more marked in the case of o-tolidine). It results that the teratogenic action of trypan blue cannot be attributed to the o-tolidine molecule proper but to an effect which results (in a for the moment unknown manner) from its combination with the other parts of the dye molecule.

The effect of ethanol upon early development in mice and rats. XIV. The effect of acute intoxication with beer and wine upon preimplantation development in rats.

The preimplantation effect of acute intoxication during the preimplantation period of pregnancy (i.p. or i.v. injection on day 4) with beer and wine has been investigated on female albino rats (Wistar strain). The following criteria were applied for checking preimplantation development: mean number of embryos/animal; topographical distribution of the embryos; developmental stage attained; occurrence of pathological embryonic forms. The control on day 5 of pregnancy revealed no significant effect upon the developmental criteria used. The data obtained are compared with our own previous results.

Researches on the formation of axial organs in the chick embryo. XI. Experimental investigations into the role of Hensen's node in somitogenesis.

In order to obtain new data with respect to the role of the node area in somitogenesis and to its "individuality" and real regression, three experimental models were applied to 1-7 somite chick embryo 1) UV irradiation of the node area (in vitro); 2) subnodal transsection (in vitro and in ovo); 3) combination of the two interventions. The main results obtained were as follows: The UV irradiation of the node area in chick embryos of early somite stage (1-7 somites) prevents, by necrotizing the cell population of the irradiated zone, the further regression of the node. This result attests the existence of a real, distinct, node cell population and the real character of regression movement. The subnodal transsection of similar embryos of about 0.1-0.2 mm caudal of the node leads (as observed also by several other authors) to the development of a "tail", projecting into the hole formed after the intervention. The "tail" contains axial organs and results from an "autonomous" regression of the node area. The previous irradiation of the node area prevents the shaping of the "tail". In both experimental models, segmentation and somite differentiation is possible caudal of the stopped node area (with the development of median somite blocks) and on the edges of the hole, respectively. Thus the node seems not to be an absolute contributor--by its regression--to the determination (to the second morphogenetic "wave") of somitogenesis (Cooke and Zeeman, 1976; Bellairs, 1980). The arrest of the node area regression does not influence (during the developmental stages studied) the rate of somitogenesis in the anterior part of the segmental plate.(ABSTRACT TRUNCATED AT 250 WORDS)

Researches on the formation of axial organs in the chick embryo. X. Further investigations on the role of ecto- and endoderm in somitogenesis.

In continuation of previous experiments, recent results concerning the determinism of somitogenesis are reported. By means of two experimental devices on explanted chick embryos the influence upon mesoderm segmentation of the early removal of ectoderm (neural plate)--before and during the formation of the first somite pairs--and removal of endo- and ectoderm after 36-40 hours of incubation was investigated. Up to date results attest that during the shaping of the first somites no essential epigenetic interrelations between overlying ectoderm (neural plate) and paraxial mesoderm are necessary for segmentation. Just before the onset of segmentation a labile determination seems to be present in the presumptive somitogenic mesoderm. As to the later role of endo- and ectoderm in segmentation, present results reveal a relative independence of the segmentation process. The absence of the above-mentioned layers does not prevent further segmentation but induces a progressive slowing down of the process.

The effect of ethanol upon early development in mice and rats. VIII. The effect of chronic consumption of some beverages upon preimplantation development in rats.

The effects of chronic consumption of some beverages (plum-brandy 24% and cognac 20%) upon preimplantation development in rats were studied. The control of possible effects was performed on day 5 by usual flushing, examination and photographying of oviductal and uterine embryos. In order to evaluate the effect of the beverage applied, the following criteria were used: mean litter size, migration of the embryos from the oviduct to the uterus, the developmental stage attained by the pre-implantation embryos and the appearance of pathological embryos. The main results were the following: both beverages applied influenced the preimplantation development; with respect to the developmental rate and to the induction of pathological changes, the effect of both beverages was similar (retardation and an increased, number of pathological morulae and blastocysts); a different action could be detected as to the mean litter size and to the migration of preimplantation embryos: plum-brandy reduced more substantially the mean litter size, whereas cognac had a more marked retarding effect upon the migration of embryos from the oviduct to the uterus: all the changes detected show a more or less marked "litter-effect". The present data were compared with the corresponding effects of chronic ethanol administration observed previously in our laboratory. No obvious potentiating effect of beverage congeners could be established. The findings are discussed in connection with other experimental models of alcohol embryo and fetopathy.

Regeneration of retinal pigment epithelium damaged in rat foetuses by 6-aminonicotinamide.

The pathological changes induced by 6-AN (administered i. p. on day 15 of pregnancy) in the pigment epithelium of the retina were studied on days 17, 18, 19 and 20 of pregnancy by light and electronmicroscopy. The marked vacuolar degeneration (enlargement of the perinuclear cisterns) observed until day 18 of pregnancy is followed on days 19 and 20 by a practically total regeneration, by the restitution of the normal microscopic and submicroscopic feature. Within the eye, the regenerative phenomena mentioned are limited to the pigment epithelium (although initially similar pathological changes are observed also in its other components.

A microcinematographic study of the in vitro development of postimplantation rat embryos.

A method for time-lapse cinematography of in vitro cultured rat embryos is presented. Technical details (especially with respect to the possibility of manipulation during filming) and the first results obtained are presented and discussed.

The effect of ethanol upon early development in mice and rats. I. In vivo effect upon preimplantation and early postimplantation stages.

The preimplantation and early postimplantation effect of chronic alcohol consumption (at least a month before mating and during pregnancy until killing) and of acute ethanol intoxication during the preimplantation period (i.v. infection of ethanol) was studied on albino rats (Wistar) and albino mice (RAP). The main results were as follows: Chronic alcoholization. Rats: significant retardation of preimplantation development and in early postimplantation stages; a tendency of lowering of the mean litter size. Mice: significant increase of the number of preimplantation pathological forms; a tendency of lowering of the mean litter size. Pathological changes show, both in rats and mice, an obvious "litter effect". Acute ethanol intoxication. Rats: significant retardation in some litters, normal or even advanced development in others. This effect differs from the previously reported effect of acute ethanol intoxication during early postimplantation stages. The results obtained attest the prenatal noxious effect of chronic ethanol consumption in both species used and of acute ethanol intoxication during preimplantation development upon early postimplantation development in rats. Within the limits of extrapolation possibilities, they represent a risk signal for other species (including human).

The effect of ethanol upon early development in mice and rats. II. In vitro effect upon early postimplantation rat embryos.

The direct effect of ethanol upon in vitro cultured 9.5 day rat embryos was investigated (2, 4, 8 and 10% ethanol added to the culture medium). The main effects recorded were as follows: 1. Significant increase of the number of "dying" embryos (beating heart without yolk sac circulation); 2. No significant increase of mortality; 3. Significant increase of the number of living embryos with deficient blood circulation; 4. Significant retardation of coiling in living embryos with a significant dose-effect relation, when the effects of 20/00 and 80/00 ethanol were compared; 5. Lowering of the mean somite number in living embryos; 6. Various macro- and macroscopical pathological changes (mainly necrotic areas in the central nervous system).

Researches on the formation of axial organs in the chick embryo. IX. On the development of somites in axial-paraaxial segments explanted to the zona pellucida.

Axial-paraaxial segments (neural tube, chorda, unilateral meso- and endoderm) excized from explanted 36--40-hour-incubated chick embryos at the level of unsegmented mesoderm, after removal of the ectoderm, were grafted onto subectodermal pockets of the zona pellucida. Under these conditions somites develop and differentiate normally. Paraaxial segments (unilateral meso- and endoderm) grafted under the same conditions show (retarded) somitogenesis only in 15% of the cases. Pure paraaxial unsegmented mesoderm grafted under the same conditions develops somites in 14% of the cases. Since in situ, the removal of the axial organs and of the endo- and ectoderm does not inhibit somitogenesis, the above-mentioned results prove that under conditions of grafting, some additionary "factors of realization" necessary for normal somitogenesis are lacking.