cytochalasin-d and Neural-Tube-Defects

Valproic acid (VPA), cytochalasin D (CD) and 7-hydroxy-2-acetylaminofluorene (7-OH-AAF) each caused abnormal closure of the anterior neuropore in rat embryos cultured in vitro in the absence of an exogenous bioactivation system. Morphological comparisons showed that although all three compounds prevented normal neural tube closure, each did so in a distinctive manner. Modulation of GSH in cultured rat conceptuses was evaluated to determine whether common responses occurred relative to the ability of different chemicals to elicit neural tube defects. Malformation incidence in embryos (10-14 somites) varied widely following exposure to CD (44%), 7-OH-AAF (29%) or VPA (17%). The incidence of CD-elicited malformations was increased by 50% following GSH depletion by L-buthionine-S, R-sulfoximine (BSO) and was decreased by nearly 60% when the cysteine pro-drug 2-oxothiazolidine-4-carboxylate (OTC) was added to the culture medium. GSH modulation also exerted significant effects on the incidence of abnormal neurulation caused by VPA or 7-OH-AAF. A relatively low incidence of open neural tubes produced by VPA or 7-OH-AAF alone was shown to be a function of the state of maturation in the embryos. Conceptuses cultured in the presence of VPA or 7-OH-AAF from an earlier gestational age (6-10 somites) showed 2-3 fold increases in the number of embryos with open neural tubes. Differential alterations in protein and DNA content were observed among embryos and yolk sacs after various treatments indicating possible differences in the site of embryotoxicity. These results demonstrate the role of GSH status on the capacity of three chemically diverse compounds to elicit abnormal neurulation in cultured rat embryos and suggest some possible mechanisms by which normal neurulation may be compromised.

Topics: 2-Acetylaminofluorene; Animals; Cytochalasin D; Cytochalasins; Female; Glutathione; Hydroxyacetylaminofluorene; Neural Tube Defects; Organ Culture Techniques; Pregnancy; Rats; Rats, Inbred Strains; Reference Values; Teratogens; Valproic Acid

During the late stages of cranial neurulation in mammalian embryos, the neural epithelium becomes concave. A thick subapical band of microfilament bundles, attached to junctions which are both vertical and horizontal in orientation, can be seen by TEM. Prior to this the neural epithelium is first biconvex and then V-shaped in transverse section, microfilament bundles are absent, and the subapical junctions are only vertical in orientation. In order to determine the role of microfilaments in cranial neurulation, rat embryos were exposed to cytochalasin D (0.15 micrograms ml-1) for 1 h at three stages of development: convex neural fold stage, early concave (prior to midline apposition at the forebrain/midbrain junction: 'preapposition') and later concave ('postapposition'). They were subsequently washed and cultured in addition-free medium for 5, 12, 24 or 36 h, then examined alive and by LM, TEM, or SEM. The degree of neural fold collapse varied with the stage of development: at the convex stage there was only slight opening out of the neural groove; early concave (preapposition) neural folds collapsed laterally to a horizontal position; later concave (postapposition) neural folds showed widening of the midbrain/hindbrain neuropore and slight neuroepithelial eversion at the anterior neuropore. Neural epithelium which had been concave prior to cytochalasin D treatment changed in structure so that the cells were broader and shorter; most of the subapical junctions were vertical in orientation, and microfilament bundles were represented either as a mass of amorphous material adjacent to the junctions, or as separated and broken filaments. Re-elevation of neural folds in 'recovery' cultures was accompanied by regeneration of apical microfilament bundles and horizontal junctions. Embryos which had been exposed to cytochalasin D at the convex or later concave stage of cranial neural fold development were able to complete cranial neural tube closure; none of the early-concave-stage embryos achieved apposition at the forebrain/midbrain junction, and all had major cranial neural tube defects. The results suggest that contraction of apical microfilament bundles plays an essential role in elevation of the neural folds and in the generation of concave curvature during the later stages of cranial neurulation. During the convex neural fold stage, microfilaments are important in maintaining neuroepithelial apposition in the neural groove, but are not crucial to maintenan

Topics: Actin Cytoskeleton; Animals; Cranial Nerves; Culture Techniques; Cytochalasin D; Cytochalasins; Cytoskeleton; Embryo, Mammalian; Microscopy, Electron; Neural Tube Defects; Rats; Rats, Inbred Strains; Time Factors

Adult toxicity, embryo lethality, and teratogenicity of cytochalasins D and E (CD and CE) were determined for A/J, C57BL/6JH, aeae, and OELN mouse strains. The adult LD50 for CD (1.9-2.6 mg/kg) and CE (3.1-3.3) were similar for the strains. Although adult LD50s were similar, the embryo lethality and teratogenicity of CD and CE vary from strain to strain when all were treated with intraperitoneal injections on days 7-9 of gestation. CD was more teratogenic than CE and frequently produced exencephaly while CE did not. CD was more teratogenic in OELN and aeae (ED50 +/- 0.7 mg/kg) than in A/J or C57BL/6J (ED50 greater than 1.2 mg/kg). OELN fetuses were more resistant to lethal effects of CD than the other strains which had fetal LD50s +/- 1.2 mg/kg maternal weight. The F1 fetuses of a cross of sensitive aeae and resistant C57BL/6J showed intermediate sensitivity to exencephaly. The results suggest a genetic contribution to the response and differences in action of CD and CE as teratogens.

Topics: Abnormalities, Drug-Induced; Animals; Cytochalasin D; Cytochalasins; Dose-Response Relationship, Drug; Lethal Dose 50; Mice; Mice, Inbred A; Mice, Inbred C57BL; Neural Tube Defects; Skull