Developmental basis of severe neural tube defects in the loop-tail (Lp) mutant mouse: use of microsatellite DNA markers to identify embryonic genotype.

Mouse embryos homozygous for the mutation loop-tail (Lp) develop lethal defects in which the neural tube remains open from the hindbrain to the caudal extremity, a condition that closely resembles the human malformation craniorachischisis. Heterozygotes develop tail defects and occasional spina bifida, but are generally viable. In order to study the early development of these defects, it is necessary to determine the genotype of embryos at stages prior to the first appearance of the morphological abnormalities. We used a microsatellite DNA sequence, Crp, that is closely linked to the Lp locus and which segregates polymorphic variants in matings between Lp/+ mice, thus permitting identification of embryos of Lp/Lp, Lp/+ and +/+ genotypes. We found that the severe phenotype craniorachischisis is present at 9.5 and 10.5 days of gestation only in Lp/Lp embryos in utero, whereas Lp/+ and +/+ littermates show neural tube closure throughout most of the body axis. The open neural tube phenotype also develops in Lp/Lp embryos growing in whole embryo culture. A small proportion of Lp/+ embryos were found to develop this phenotype in vitro, but only when culture conditions were suboptimal. Analysis of 8.5-day embryos revealed that the initial defect in Lp/Lp embryos is failure to initiate neural tube closure at the cervical/hindbrain boundary when the embryo has 6-7 somites. Thereafter, the neural tube remains open throughout the body axis, with the exception of the midbrain and forebrain where neural tube closure is initiated independently. Closure at the midbrain/forebrain boundary does not appear to be defective in Lp/Lp embryos. Heterozygous Lp/+ embryos initiate neural tube closure at the cervical/hindbrain boundary with a slight delay compared with +/+ littermates. Moreover, at 10.5 days of gestation, Lp/+ embryos undergo delayed closure of the posterior neuropore. Thus, Lp/+ embryos are defective in several aspects of the neurulation process. The pattern of delayed neuropore closure in Lp/+ embryos resembles that caused by the ct and Sp mutations and is likely to be responsible for the development of tail defects (i.e., looped tails) and spina bifida in Lp/+ mice. The use of microsatellite markers to determine the genotype of mutant embryos has general application: microsatellites are widespread throughout the mouse genome, so that informative sequences are likely to be available with close linkage to the majority of mutant genes. Moreover, polymorphisms can be detected using the polymerase chain reaction, making it possible to determine the genotype of very early embryos when only small amounts of material are available.

Copp AJ ，Checiu I ，Henson JN 《DEVELOPMENTAL BIOLOGY》

Identification of a new chemically induced allele (Lp(m1Jus)) at the loop-tail locus: morphology, histology, and genetic mapping.

Loop-tail (Lp) is a semidominant mutation that affects neurulation in mice. Heterozygous animals are characterized by a looped-tail appearance (pig tail) and wobbly head movements while homozygous embryos exhibit a neural tube closure defect that extends from the caudal midbrain to the tip of the tail. The Lp gene has been finely mapped to the distal part of chromosome 1, and a positional cloning strategy has been initiated to isolate the defective gene. This study represents the characterization of a new Lp allele (Lp(m1Jus)) induced by N-ethyl-N-nitrosurea mutagenesis. Lp(m1Jus)/+ mice have a looped-tail appearance, and both Lp(m1Jus)/Lp(m1Jus) homozygotes and Lp/Lp(m1Jus) compound heterozygotes fail to initiate neural tube closure along most of the embryonic axis. These data indicate that the Lp(m1Jus) allele causes a neural tube defect and overall phenotype similar to that of the original Lp allele. Segregation analysis of 90 (Lp(m1Jus)/+ x C57BL/6J)F(1) x C57BL/6J looped-tail mice with seven markers that define the Lp genetic map (D1Mit455/D1Mit146/D1Mit148/D1Mit270-1 cM-D1Mit113-0.4 cM-Lp-0.2 cM-D1Mit149-0.8 cM-D1Mit115) showed significant linkage between Lp(m1Jus) and all loci analyzed (P < 0.0001). Eight crossovers were detected with the proximal cluster of D1Mit455, D1Mit146, D1Mit148, and D1Mit270, indicating a recombination rate higher than expected in this region, and a single recombinant was encountered with the distal markers D1Mit149 and D1Mit115. Based on these phenotypic and genetic data, Lp(m1Jus) is most likely allelic to Lp, thereby representing a valuable additional tool for the positional cloning of the Lp gene and its subsequent molecular characterization.

Kibar Z ，Underhill DA ，Canonne-Hergaux F ，Gauthier S ，Justice MJ ，Gros P ... -  《GENOMICS》

Analysis of the embryonic phenotype of Bent tail, a mouse model for X-linked neural tube defects.

Neural tube defects, mostly believed to result from closure defects of the neural tube during embryonic development, are frequently observed congenital malformations in humans. Since the etiology of these defects is not well understood yet, many animal models for neural tube defects, either arising from spontaneous mutations or generated by gene targeting, are being studied. The Bent tail mouse is a model for X-linked neural tube defects. This mutant has a characteristic short and kinked tail. Exencephaly occurs in Bent tail embryos with a frequency of 11-16%. Laterality defects also belong to the phenotypic spectrum. In this study, we analyzed the embryonic phenotype in further detail using scanning electron microscopy during the stages of neurulation. We observed a number of defects in both wild type and Bent tail embryos, including a kinked neural tube, tight amnion, delay in axial rotation and even malrotation. The severity or frequency of most defects, the delay in axial rotation excluded, was significantly higher in Bent tail embryos compared to wild type embryos. Other abnormalities were seen in Bent tail embryos only. These defects were related to anterior and posterior neural tube closure and resulted in exencephaly and a closure delay of the posterior neuropore, respectively. The exencephalic phenotype was further analyzed by light microscopy in ED14 embryos, showing disorganization and overgrowth in the mesencephalon and rhombencephalon. In conclusion, the anterior and posterior neural tube closure defects in the Bent tail are strictly linked to the genetic defect in this mouse. Other phenotypic features described in this study also occur in the wild type genetic background of the Bent tail strain. Apparently, the genetic background contains elements conducive to these developmental abnormalities.

Franke B ，Klootwijk R ，Hekking JW ，de Boer RT ，ten Donkelaar HJ ，Mariman EC ，van Straaten HW ... -  《anatomy and embryology》

Circletail, a new mouse mutant with severe neural tube defects: chromosomal localization and interaction with the loop-tail mutation.

Circletail (Crc) is a new mouse mutant that exhibits a severe form of neural tube defect, craniorachischisis, in which almost the entire neural tube fails to close. This phenotype is seen in very few other mutants, the best characterized of which is loop-tail (Ltap(Lp), referred to hereafter as Lp). We tested the possibility of allelism between Lp and Crc by intercrossing Lp/+ and Crc/+mice. A proportion of double heterozygotes (Lp/+,Crc/+) exhibit craniorachischisis, revealing failure of complementation. However, genetic analysis shows that Crc is not linked to the markers that flank the Lp locus and cannot, therefore, be an allele of Lp. A genome-wide scan has localized the Crc gene to a region of 8.8 cM on central chromosome 15. Partial penetrance of the craniorachischisis phenotype in Crc/+,Lp/+double heterozygotes suggests the existence of a third, unlinked genetic locus that influences the interaction between Crc and Lp.

Murdoch JN ，Rachel RA ，Shah S ，Beermann F ，Stanier P ，Mason CA ，Copp AJ ... -  《GENOMICS》

Neural tube development in mutant (curly tail) and normal mouse embryos: the timing of posterior neuropore closure in vivo and in vitro.

A dye-injection technique has been used to determine the developmental stage at which posterior neuropore (PNP) closure occurs in normal and mutant curly tail mouse embryos. In vivo, the majority of non-mutant embryos undergo PNP closure between 30 and 34 somites whereas approximately 50% of all mutant embryos show delayed closure, and around 20% maintain an open PNP even at advanced stages of development. A similar result has been found for embryos developing in vitro from the headfold stage. Later in development, 50--60% of mutant embryos in vivo develop tail flexion defects, and 15--20% lumbosacral myeloschisis. This supports the view that delayed PNP closure is the main developmental lesion leading to the appearance of caudal neural tube defects in curly tail mice. The neural tube is closed in the region of tail flexion defects, but it is locally over-expanded and abnormal in position. The significance of these observations is discussed in relation to possible mechanisms of development of lumbosacral and caudal neural tube defects. This paper constitutes the first demonstration of the development of a genetically induced malformation in vitro.

Copp AJ ，Seller MJ ，Polani PE 《-》