A mesencephalic flexure, particularly marked in the more advanced embryos e.g. 1878, demarcates the mesencephalon. Rostral to the midbrain lies the prosencephalon and caudally the rhombencephalon, the latter can be divided into 4 segments or pro-rhombomeres; A, B,C and D. B underlies the otic placode, and D lies medial to the occipital somites and comprises the hypoglossal region of the hindbrain. O’ Rahilly R., Müller F., (1994) pp35-42. The neural crest migrating from the neurosomatic junction of rhombomere A (future hindbrain) is forming lateral condensations which are thought to be the primordia of the fifth cranial nerve ganglia. Ingalls N.W. (1920) p.67
The pro-rhombomeric divisions of the hindbrain can be further subdivided in this stage. Pro-rhombomere A contains rhombomeres 1-3, pro-rhombomere B comprises of rhombomere 4, pro-rhombomere C is divided into rhombomeres 5,6 and 7 while pro-rhombomere D, the region of the occipital somites, constitutes the 8th rhombomere. The rhombomeres are transverse swellings in the brain and some are characterised by their associated developing cranial ganglia. e.g. the trigeminal in rhombomere 2, the vestibulocochlear in rhombomere 4 and the glossopharyngeal and vagal in rhombomere 6 and rhombomere 7, respectively. O’Rahilly R., Müller F., (1987) p. 117. Neural crest cells are being formed at the neurosomatic junction as before. A cell free layer is visible in the lateral walls of RhD Müller F., O’Rahilly R., (1986)
An eighth rhombomere is distinguishable at this stage. The rhombomeres are transverse swellings in the hindbrain and some are characterised by associated developing cranial nerve ganglia. Expansions in the floor of rhombomeres 2,4 and 6 are seen while the roof is becoming thin. The first nerve fibres have been observed in silver-impregnated specimens. e.g. Carnegie Embryo No.6097. These were seen in the future region of the sulcus limitans in rhombomeres 2,4,6 and 7 and will contribute to the lateral longitudinal tract. Fibres of the future ventral longitudinal tract may also be observed in rhombomere 7. Müller F., O’Rahilly R., (1987) Fig. 5b. Intramural roots of the hypoglossal nerve (XII) can be seen at the levels of somites 2-4. Müller F., O’Rahilly R., (1987) Fig. 5b. A marginal layer is present in the rhombencephalic wall extending from the floor towards the roof as far as the dorsal limit of the developing cranial ganglia. Müller F., O’Rahilly R., (1987) Fig. 3C & E
Between rhombomere 1 and the midbrain, a constriction develops in the wall of the neural tube. It is labelled by some as the isthmic segment or rhombencephalic isthmus. See Müller F., O’Rahilly R., (1988b) Fig. 9.
According to Müller F., O’Rahilly R., (1988b), the primordial cerebellum is detectable in the alar lamina of rhombomere 1 by the appearance of loosely arranged cells that will form its intermediate layer. The marginal layer can be seen around both dorsal and ventral portions of the neural tube.
The lateral longitudinal fasciculus extends, to varying degrees between embryos, from the midbrain to the caudal end of the hindbrain. 4 out of 4 embryos studied by O’Rahilly R., et al., (1984) possess this fibre tract. In Carnegie embryo no. 8066, loosely arranged unipolar neurons, with fibres running along the level of the sulcus limitans, extend from the isthmic area to the last rhombomere. Situated more ventrally, unipolar cells give rise to fibres that join to form the ventral longitudinal fasciculus. The fibres run ventrally in the marginal layer and some cross to the other side forming a rudimentary ventral commissure. Only 7of 12 stage 13 embryos possessed fibres of the ventral longitudinal tract O’Rahilly R., et al., (1984). The medial longitudinal fasciculus only extends as far as the isthmic segment in embryos No 6473 and 8006 Müller F., O’Rahilly R., (1988b).
Cranial nerve nuclei A common sheet of motor neurons is visible which will soon become separated into distinct columns. Fibres from neurons located ventromedially in the basal plate ascend in the marginal layer and exit at the level of the sulcus medians. These fibres are thought to be the first visceral efferent fibres, later forming the parasympathetic output for the trigeminal (V), facial (VII), glossopharyngeal (IX), vagal (X) and accessory (XI) nerves. Müller F., O’Rahilly R., (1988b)Fig. 10 C,D. The trigeminal, glossopharyngeal, vagal and accessory motor nuclei give off intramural fibres and motor roots which leave at the level of the sulcus limitans. The extramural fibres of the accessory nerve are the longest and are accompanied by neural crest cells. These fibres extend to the lower cervical levels of the spinal cord. Fibres from the facial motor nucleus are not yet extramural. Müller F., O’Rahilly R., (1988b)Fig 8, 10 Neurons located laterally to those of the future visceral efferent neurons give rise to fibres that exit the neural tube ventrally. This ‘ventrolateral’ group of fibres will later form the somatic efferent motor output. The hypoglossal (XII)fibres belong to this group and the extramural fibres are obvious, whereas those of the abducens (VI) do not yet exit the neural tube. The hypoglossal nucleus is distinguishable containing unipolar neurons and the abducens nucleus is mentioned by Müller F., O’Rahilly R., (1988b) although no pictorial evidence of its presence is given. The common afferent tract containing fibres from ganglia of nerves V, VII, IX and X is developing. Although no pictorial evidence of this is given Müller F., O’Rahilly R., (1988b), The trigeminal spinal tract is present in 4 out of 14 embryos studied and the spinal tract of nerves VII and VIII are present in 3 out of 13 embryos studied O’Rahilly R., et al., (1984).
Eight rhombomeres are still distinguishable in this stage as well as the isthmic segment. Expansions can be seen in the floor of rhombomeres 2,4 and 6 and adjacent to rhombomere 4 is the facial-acoustic (VII-VIII) ganglion, rhombomere 6 is associated with ganglion 9 and rhombomere 7 with the vagal ganglion. The otic vesicle characterises rhombomere 5 but is beginning to expand over rhombomere 6. The marginal layer can be seen around both dorsal and ventral portions of the tube, and an intermediate layer extends over more than two-thirds of the surface. According to Müller F., O’Rahilly R., (1988a), the cerebellum is detectable in the alar lamina of Rh1. Most of the cerebellar plate has developed an intermediate layer which possesses short fibres.. Surrounding it dorsally is a dense area which is the future site of the rhombic lip.
The lateral longitudinal fasciculus extends, to various degrees between embryos, from the midbrain to the caudal end of the hindbrain. The nucleus of this fasciculus is situated level with the sulcus limitans. The ventral longitudinal fasciculus is continuous with the medial longitudinal fasciculus in the midbrain.
Cranial nerve nuclei The common sheet of motor neurons has become separated into distinct columns, the visceral motor nuclei of cranial nerves V, VII/VIII, IX, X/XI are situated near the medial plane forming the ventromedial column with efferent fibres exiting the neural tube at the level of the sulcus limitans whereas those of nerves VI and XII are more lateral (somatic efferent). This is not the definitive state and during this stage some of the cells of the ventromedial column (primarily those of the trigeminal and facio-acoustic nuclei) migrate to the level of the sulcus limitans forming a dorsolateral column. Müller F., O’Rahilly R., (1988a)Fig 8A-D The first signs of the relocation of the trigeminal nucleus can be seen as according to Müller F., O’Rahilly R., (1988a), ‘a small group of neurons accumulates in the angle between the efferent and afferent trigeminal fibres and forms a dorsolateral nucleus’ This has occurred in 9/29 embryos studied by O’Rahilly R., et al., (1984). The dorsolateral nucleus of the glossopharyngeal nerve is also migrating away from the ventromedial position during this and the next stage. It is found in a dorsolateral position in 3/23 embryos studied by O’Rahilly R., et al., (1984) and will form the inferior salivatory nucleus. According to O’Rahilly R., et al., (1984) nuclei of cranial nerves III and IV have developed in nearly all embryos of this stage. A ‘nucleus’ is defined as an area of ‘lower cellular density and slightly larger cellular size’. The trochlear (IV) nucleus is located in the isthmic segment between fibres of the medial longitudinal fasciculus and the ventricular layer and its neurons give rise to fibres which ascend towards the roof in about half the embryos of this stage. In almost all embryos of this stage the spinal tract of the trigeminal (V), facial (VII) and vestibular (VIII) nerves have formed, and O’Rahilly R., et al., (1984) claim that in 6/34 embryos of this stage the common afferent tract (defined as identifiable when ‘the sensory axons of nerves V-XI have extended as a continuous tract’) has developed.
An intermediate layer extends over the dorsal half of the rhombencephalon and presages the development of the sensory nuclei in the alar plate. Fibres in the ventral commissure increase and the septum medullae (future median raphe) develops which extends as far as the isthmus.
According to Müller F., O’Rahilly R., (1988c), the cerebellum extends from the trochlear decussation to the rostral boundary of the 2nd rhombomere. It is formed rostrally by the bilateral alar plates of the isthmic segment (the junction of the two being the future site of the superior medullary velum). The thickened wall of the 1st rhombomere will later form the corpus cerebelli. Dorsal to this thickening there is a conspicuous absence of marginal layer- an area that will form the rhombic lip. The various cerebellar areas may be distinguished by the arrangement of cells and fibres in the marginal and intermediate layers. Deep running fibres between the intermediate and ventricular layers are thought to be ascending trigeminal fibres while fibres running superficially in the marginal areas are probably vestibular. Müller F., O’Rahilly R., (1988c)Fig 9C,D
Cranial nerve nuclei The ventromedial cell column containing nuclei of the visceral efferent type and the venterolateral column of somatic efferent nuclei are still in the same location as before, but as nuclei from the ventromedial cell column migrate towards the sulcus limitans, dorsolateral nuclei begin to form in the angle between emerging motor fibres and entering sensory fibres Müller F., O’Rahilly R., (1988c). Thus by this stage somatic efferent nuclei (III, IV, VI, XII) are present in a ventrolateral position, general visceral nuclei (superior salivatory VII, inferior salivatory IX, and dorsal vagal X) have begun to migrate to a dorsolateral position and special visceral (branchial) efferent nuclei (trigeminal V,) have also migrated to a dorsolateral position. Later the dorsolateral nuclei form a distinct ventral and dorsal group which constitute the more definitive general and special visceral divisions. According to Lemire R.J., et al., (1975) chapter 8, the abducens nucleus is not defined until this stage but Müller F., O’Rahilly R., (1988c) state ‘The somatomotor nuclei of cranial nerves 3,4,6 and 12 were present before stage 15’ Of the dorsolateral visceral efferent nuclei the superior salivatory nucleus is only just beginning to form- seen in 4 out of 26 embryos studied by O’Rahilly R., Müller F.,(1994). The inferior salivatory nucleus is visible in 9 out of 26 embryos studied by O’Rahilly R., Müller F.,(1994)but the dorsolateral nucleus of the vagus has not yet formed. O’Rahilly R., Müller F.,(1994). Sensory nuclei have not yet developed but the common afferent tract contains most afferent fibres and by the end of this stage the future tractus solitarius has begun to separate from the common tract, beginning in the area of nerve VII and occuring in 4/24 embryos studied by O’Rahilly R., et al., (1984) According to Yokoh Y., (1968) fibres of the tractus solitarius are distinguishable in the marginal zone. The special afferent fibres come from the vestibular and cochlear nerves. According to Müller F., O’Rahilly R., (1988c) the vestibular nerve now possesses sensory nerve fibres that ‘join the common afferent tract’
The rhombomeres are now no longer distinguishable from the outer surface of the hindbrain. Müller F., O’Rahilly R., (1989a).
Cranial nerve nuclei The hypoglossal nucleus is depicted in a graphic reconstruction of an embryo of this stage Streeter G.L., (1908a) Plate II. It is continuous with the motor column in the ventral horn of the spinal cord. The visceral efferent nuclei continue their relocation to a dorsolateral position and the first signs of a division into a special visceral (branchial) and general visceral division is apparent as the nuclei separate into a dorsolateral and ventrolateral position. The superior salivatory nucleus (VII), inferior salivatory nucleus (IX) and dorsal vagal nucleus (X) have formed in their dorsolateral position. The inferior salivatory nucleus is visible in 29 out of 35 embryos studied by O’Rahilly R.,Muller F., (1994). The site of the relocated vagal nucleus (X) has caused some debate as to whether or not it may be the nucleus ambiguus. O’Rahilly R., et al., (1987) claim that a ‘ventrolateral and/or dorsolateral nucleus of X is present’ in 30/32 embryos of this stage. However in O’Rahilly R.,Muller F., (1994)p. 104 it is stated that ‘An examination of first class material shows that it is not possible to distinguish the nucleus ambiguus at stage 16 or earlier’.
In 65% of embryos studied by Müller F., O’Rahilly R., (1989a), the tractus solitarius is beginning to separate off from the common afferent tract. The is most obvious in the area where the fibres of the nervus intermedius (VII) join the tract. The common afferent tract reaches the trigeminal area rostrally, and the mesencephalic tract of the trigeminal (V) extends caudally to the trigeminal ganglion. According to Lemire R.J., et al., (1975) the mesencephalic and chief sensory nuclei of the trigeminal appeared at stage 15 however O’Rahilly R.,Muller F., (1994)claim that while the mesencephalic nucleus is present in 27 of 28 embryos studied of this stage, the chief sensory nucleus is not present until stage 18. The vestibular fibres contribute a large proportion of the common afferent tract fibres. Afferent fibres from the cochlear ganglion also now enter the brain wall. Running in the marginal layer, vestibulocerebellar fibres are visible in sagittal sections and trigeminocerebellar fibres run between the intermediate and ventricular layers of the tegmentum. By this stage the fibres have reached the base of the cerebellum. The first internal arcuate fibres are visible in the caudal rhombencephalon, crossing in the ventral commissure.
Slight grooves on the ventricular surface of the rhombencephalon may help distinguish the former site of some of the rhombomeres. Medial to fibres of the mesencephalic tract of V, in the basal region of the isthmic segment and Rh1, elongated cells lie at boundary between the intermediate and ventricular layers. These cells constitute the locus coeruleus. According to Yokoh Y., (1968) a nerve bundle in the ventral/medial part of the cerebellar plate which extends to the dorsal and lateral side of the spinal tract of the trigeminal, represents the beginnings of the inferior cerebellar peduncle.
Cranial nerve nuclei The relocating cells of the visceral efferent nuclei are clearly visible Müller F., O’Rahilly R., (1989b)Fig. 14D-F. By now the general visceral efferent nuclei (inferior/superior salivatory, dorsal vagal) are present in a dorsolateral position in all embryos and the special visceral efferent nuclei are also forming in a slightly more ventrolateral position- the definitive trigeminal nucleus is present in about half the embryos studied but the nucleus ambiguus is still difficult to discern according to Müller F., O’Rahilly R., (1989b)The common afferent tract reaches as far rostrally as the cerebellum. It contains mainly trigeminospinal fibres as gustatory fibres are beginning to separate off and form the tractus solitarius. Müller F., O’Rahilly R., (1989b)Fig. 14A The first signs of the sensory nuclei are appearing in the vestibular nerve area.
The pontine flexure deepens and enables the division of the rhombencephalon into future pontine and medullary regions. The isthmic segment has developed a ventral outpocket sometimes named Isthmushöcker or Tuberculum interpedunculare. Later dopaminergic cells develop rostral to this and it serves to locate the interpeduncular nucleus. Otherwise known as the dorsal nucleus of the trapezoid body, , the superior olivary nucleus is depicted in a reconstruction of embryo no. D-7924, Müller F., O’Rahilly R., (1990). It is lateral to the reticular formation and fibres from it form the lateral lemniscus The developing dentate nucleus in the cerebellum apparently receives afferent fibres via the vestibulo and trigeminocerebellar tracts, which constitute a primary inferior cerebellar peduncle. Efferent fibres from it run to the isthmic nucleus and finally to the red nucleus (the dentatorubral tract) O’Rahilly R., et al., (1988)
Cranial nerve nuclei In all embryos of stage 18 studied by O’Rahilly R., et al., (1988), the primordium of the chief sensory nucleus of trigeminal V was found. It is described as consisting of ‘ small, poorly differentiated, fusiform cells, which are located between the incoming fascicles of the sensory root’ depicted in O’Rahilly R., Müller F., (1994)Fig 18-14 C . The tractus solitarius has completely separated from the common afferent tract. Dorsal and ventral cochlear (special somatic afferent) nuclei begin to appear. (Depicted in a reconstruction of Embryo D-7924) O’Rahilly R., Müller F., (1994)Fig 18-14 F). But according to O’Rahilly R., et al., (1988)the nuclei are not present but appear rapidly in stage 19. Fibres leave the vestibular area in the alar plate and join the medial longitudinal fasciculus. O’Rahilly R., Müller F., (1994) Fig 18-14 F. The general visceral afferent component has not yet developed into distinct glossopharyngeal and vagal sensory nuclei. The dorsal funiculus extends further rostrally to reach the cuneate and gracile tubercles.
A groove has formed in the isthmic area between the cerebellum and midbrain and the floor of the rhombencephalon is becoming thicker with the amount of fibres passing through it. The lemniscal decussation is visible just caudal to the junction of the medulla to the spinal cord. Fibres passing from the dentate nucleus to the red nucleus (dentatorubral tract) indicate the first signs of the superior cerebellar peduncle. O’Rahilly R., Müller F., (1994)p.158. According to O’Rahilly R., et al.,(1988) , the medial accessory olivary nucleus appears as a condensation of undifferentiated cells medial to the intramural fibres of the hypoglossal nerve roots.
Cranial nerve nuclei The ventral cochlear nucleus is described as a dense mass of cells lateral to the primitive inferior cerebellar peduncle in embryo D-9588 studied by O’Rahilly R., Müller F., (1994)Fig. 9.13
According to O’Rahilly R., Müller F., (1994)Fig 20.3, decussating fibres of the fasciculus cuneatus and gracilis can be found at the transitional area from the rhombencephalon to spinal cord. This decussation is known as the decussation of medial leminisci and fibres continue in the medial lemniscus to the ventral posterior thalamus. Cranial nerve nuclei An accessory abducens nucleus has been identified in embryos of this stage by Muller F., O’Rahilly R., (1990) Fig 14D An accessory hypoglossal nucleus has been identified in embryos of this stage by Muller F., O’Rahilly R., (1990)Fig 14B. The main nucleus contains bigger cells than the more ventral accessory nucleus