Neurulation in the cranial region--normal and abnormal

Abstract

Cranial neurulation is the embryonic process responsible for formation of the brain primordium. In the mouse embryo, cranial neurulation is a piecemeal process with several initiation sites and two neuropores. Variation in the pattern of cranial neurulation occurs in different mouse strains, and a simpler version of this morphogenetic scheme has been described in human embryos. Exencephaly is more common in females than in males, an unexplained phenomenon seen in both mice and humans. As the cranial neural tube closes, a critical morphogenetic event is the formation of dorsolateral bending points near the neural fold tips, which enables subsequent midline fusion of the neural folds. Many mutant and gene-targeted mouse strains develop cranial neural tube defects, and analysis of the underlying molecular defects identifies several requirements for normal dorsolateral bending. These include a functional actin cytoskeleton, emigration of the cranial neural crest, spatio-temporally regulated apoptosis, and a balance between cell proliferation and the onset of neuronal differentiation. A small number of mouse mutants exhibit craniorachischisis, a combined brain and spine neurulation defect. Recent studies show that disturbance of a single molecular signalling cascade, the planar cell polarity pathway, is implicated in mutants with this defect.

Fig. 1

Newborn (A) and E15.5 fetuses (B,C) showing the main cranial neural tube defects in mice. (A) Anencephaly in a curly tail mutant. Note the lack of skull vault (between small arrows). The skull base is overlaid by the ‘area cerebrovasculosa’, the remnant of the degenerate brain tissue. Open spina bifida (strictly, myelocele) is present in the lumbosacral region (arrowhead). (B) Exencephaly of the midbrain in a curly tail mutant, showing the everted cranial neural folds (between small arrows). Open spina bifida is also present, affecting the lumbosacral region (arrowhead). (C) Craniorachischisis in a Celsr1 mutant, in which the neural tube is open from midbrain to low spine (between small arrows). Note the presence of a curled tail in all cases (large arrows in A–C). B and C are modified from Copp et al. (2003b).

Fig. 2

Stages of mouse cranial neurulation, as seen diagrammatically in whole embryos at (A) E8.5 and (B) E9 (with extra-embryonic membranes removed). At E8.5 (five-somite stage) the neural folds are approaching one other at the site of Closure 1 (arrow in A), whereas the cranial neural folds are still wide apart. Ten hours later, at E9 (ten-somite stage), Closure 1 has occurred and neural fold fusion has also been initiated at Closures 2 and 3 (arrows in B). These closure initiation events define two neuropores where cranial closure will later be completed: the anterior or rostral neuropore (ANP) and the hindbrain neuropore (HNP). A third neuropore (the posterior neuropore, PNP) is present in the spinal region. Somites are shown in yellow.

Fig. 3

Variation in the position of Closure 2 (arrowhead) among different inbred mouse strains, as shown on diagrams of the cranial region (viewed from the left side). Open neural folds in the midbrain and hindbrain are shown in red; forebrain neural folds are shown in green. Arrows show the direction of closure between initiation sites.(A) In the majority of strains, Closure 2 occurs at the forebrain/midbrain boundary.(B) One variation is for Closure 2 to occur caudally, within the midbrain, as in the DBA/2 strain. This mode of closure supports midbrain neural fold apposition and counteracts any tendency for these neural folds to remain open, thereby diminishing risk of exencephaly.(C) Another variation is for Closure 2 to occur rostral to the forebrain/midbrain boundary, as in the NZW strain. This type of Closure 2 destabilizes elevation and apposition of the midbrain neural folds, and increases the chance of exencephaly.(D) The most extreme situation, as seen in the SELH/Bc strain, is where Closure 2 is absent altogether. Susceptibility to exencephaly is very high in the absence of Closure 2, with 17% of SELH/Bc mice exhibiting exencephaly. Note that human embryos are suggested to complete cranial neurulation, as shown in D (see also Fig. 4). Abbreviations: fb, forebrain; hb, hindbrain; mb, midbrain. Figure modified from Fleming & Copp (2000).

Fig. 4

Mouse (A) and human (B) embryos soon after the completion of cranial neurulation. Note the relatively smaller midbrain region (mb) in the human embryo compared with the mouse. (A) The mouse embryo (E10) has closed its brain by piecemeal neurulation between three initiation points (1, 2 and 3) with completion of closure at two neuropores (anterior neuropore, ANP; hindbrain neuropore, HNP). (B) The human embryo (35days) seems likely to have completed brain closure by caudally directed spread of neurulation from a rostral initiation site (Closure 3) and rostrally directed spread from Closure 1. The relatively smaller brain may make this simpler pattern of cranial neurulation feasible mechanically in human development. Scale bar: 0.5mm in A; also applies to B. A is modified from Van Straaten & Copp (2001).

Fig. 5

Appearance of the midbrain neural folds before (A) and after (B) the stage of Closure 2 during mouse cranial neurulation.(A) Initially, the midbrain neural folds are convex, with divergence of the fold apices (small arrows).(B) A few hours later, Closure 2 has been initiated at the forebrain/midbrain boundary (large arrow). Dorsolateral bending points have developed, enabling the midbrain neural folds to reverse their curvature and adopt a bi-concave morphology. This allows the folds apices to approach one other in the dorsal midline (small arrows). Note the open anterior neuropore (arrowhead in B). Scale bar: 0.15mm. Figure modified from Fleming & Copp (2000).

Fig. 6

Summary of the principal developmental mechanisms that appear essential for successful closure of the cranial neural tube, with particular reference to the formation of dorsolateral bending points. Examples of mouse mutants in which each mechanism appears to be disrupted are shown in parentheses. Mechanisms are depicted unilaterally for the sake of clarity, but operate bilaterally. See text for explanation. Figure modified from Copp et al. (2003b).