Failure of neural tube closure in the loop-tail ( Lp) mutant mouse: analysis of the embryonic mechanism

Abstract

Loop-tail ( Lp) is unique among mouse mutants in failing to initiate neural tube closure at the cervical/hindbrain boundary (so-called `Closure 1'), at the 5–7 somite stage. Lp/Lp embryos go on to develop a malformation that closely resembles cranio-rachischisis, the most severe neural tube defect found in humans. We investigated several possible embryological mechanisms that may underlie this failure of neural tube closure in Lp. The genotypes of Lp/Lp, Lp/+ and +/+ embryos from mixed litters were identified using the polymerase chain reaction to amplify a polymorphic microsatellite sequence that is very closely linked to Lp. At post-neurulation stages of development, Lp/Lp embryos have a shortened body axis, which could suggest a defect of axial elongation as the primary anomaly in Lp. However, we found that axial elongation is normal in Lp homozygotes prior to the stage of defective Closure 1, indicating that the shortened body axis of later embryos is a secondary effect of the neurulation anomaly, or an independent effect of the Lp mutation. Some workers have reported cell proliferation rates to be abnormal in later stage Lp/Lp embryos. We observed variations in [ 3H]thymidine labelling index, and mitotic index, between embryonic tissues, and between embryos at different somite stages. However, Lp/Lp, Lp/+ and +/+ embryos had closely similar cell proliferation parameters, arguing against a mechanism based on faulty embryonic growth. Thirdly, we tested the hypothesis that the defect in loop-tail results from an inability of the neural folds to become apposed, specifically at the site of Closure 1. By tying a silk suture around the embryonic axis, at the future site of Closure 1, we were able to effect convergence of the neural folds at this site. Neural fold closure failed to progress along the body axis in sutured Lp/Lp embryos, however, in contrast to operated Lp/+ and +/+ embryos which exhibited normal progression of neural tube closure. The embryonic defect in loop-tail appears, therefore, to involve either a general inability of the spinal neural folds to become apposed along the spinal region, or a defect in the process of neural fold fusion.