the early development of the hypoglossal nerve and occipital somites in staged human embryos

THE AMERICAN JOURNAL OF ANATOMY 169:237-257 (1984)

The Early Development of the Hypoglossal Nerve and Occipital Somites in Staged Human Embryos

RONAN O’RAHILLY ANU FABIOLA MULLER Carnegie Laboratories of Embryology, California Primate Research Center, and Departments of Human Anatomy and Neurology, University of California, Davis, California 95616

ABSTRACT Serial sections of 105 human embryos (including 20 silver preparations) from stage 11 (24 days) to stage 22 (54 days) were studied, and 23 graphic reconstructions were prepared. The hypoglossal nucleus is evident a t stage 12 and becomes isolated from other efferent nuclei at stage 14. The first hypoglossal nerve fibers appear a t stage 12. The roots unite a t stage 14 and the main trunk arrives in the tongue at stage 15. Four occipital somites can be identified during stage 13, and the sclerotomic material forms two bilateral masses. The fourth sclerotome separates in stage 14 and develops like a vertebra. This and the remaining sclerotomic material form the basioccipital and exoccipital parts of the chondrocranium, which are the first to appear. Four occipital myotomes develop and grow towards the tongue as the “hypoglossal cord”, which arrives prior to the hypoglossal nerve. The developmental similarity in the hypoglossal region between birds and mammals, combined with experimental studies in birds, renders it extremely likely that the hypoglossal musculature in mammals also is derived from occipital somites. The present study is the first in which this conclusion is adequately supported in the human. This investigation aids in the interpretation and timing of origin of variations (e.g., bipartite hypoglossal canal) and anomalies (e.g., persistent hypoglossal artery).

Developmentally, the hypoglossal nerve is a segmental structure the roots of which are intersomitic, as are those of the spinal nerves. A dorsal root and ganglion are lacking in the hypoglossal nerve, although claims have been made concerning their presence in rare instances, or even “occasionally” (Streeter, 1912). Illustrations offered in support, however, generally show a ganglion on the accessory nerve (Fig. 94 of Streeter, 1912) and hence not a (Froriep’s) hypoglossal ganglion. Indeed, Froriep, who described the ganglion in hoofed animals, found no trace of hypoglossal dorsal roots or ganglia in the adult human (Froriep and Beck, 1895). The development of the hypoglossal nerve has been examined in human embryos chiefly by Streeter (1904), Pearson (19391, and Windle (1970). No systematic study in staged embryos, however, has appeared.

The foregut arches dorsally in association with the arching pattern of the brain, and an elevation on the basal wall of the foregut

represents the tongue (Blechschmidt and Gasser, 1978). Vaulting of the tongue pro- gresses tridimensionally, and consequently its musculature develops in all three dimen- sions (Blechschmidt and Gasser, 1978).

The lingual musculature of mammals is believed to arise either (1) in situ from local mesenchyme, or (2) from the myotomes of rostra1 somites. The latter view was first pro- posed in the 1880s by Froriep, who related the origin to the course of the hypoglossal nerve (Hunter, 1935b; Bates, 1948). In most accounts, “it is usually assumed that the tongue musculature is derived from the occipital myotomes which appear to be serially related to the N. hypoglossus. There is, however, no direct evidence whatever for this statement” (Lewis, 1910) in the human. That such assumptions are by no means always

Address reprint requests to Dr. O’Rahilly, Carnegie Laborato- ries of Embryology, California Primate Research Center, Uni- versity of California, Davis, CA 95616.

Received May 2, 1983. Accepted October 17,1983

@ 1984 ALAN R. LISS. INC.

238 R. O’RAHILLY AND F. MULLER

justified can be seen from the equally com- mon description of a pronephros, although it has been shown that “the concept of the pronephros does not apply to the human embryo” (Torrey, 1954).

The accumulated conclusions of morphologic studies have been reaffirmed by several experimental investigations in the chick embryo. Occipital somites were marked with carbon particles, which were later found in the tongue musculature and geniohyoid (Deuchar, 1958). Unilateral extirpation of rostral somites or their myotomes was followed by absence or reduction of the lingual muscles (Hammond, 1965). In a further study (in which the last-mentioned work was not cited), occipital somites and overlying ectoderm marked with tritiated thymidine were transplanted to untreated host embryos (Ha- zelton, 1970). It was shown that “the hypoglossal as well as other hypopharyngeal (e.g., laryngeal) musculature originates from the occipital somites.” In addition, “there is an occipital somitic contribution to the primitive meninx, to the endothelial walls of developing blood vessels, possibly to microglial cells and to the cartilage surrounding the notochord.”

It is still true that “the origin of the (lingual) musculature has not yet been thor- oughly studied in the human embryo” (McMurrich, 1912). This is in large measure caused by (1) the much greater difficulty encountered in tracing the course of the occipital, compared to the cervical and thoracic, somites, and (2) the lack of precise reconstructions. A major objective of the present study is to elucidate the occipital somites in a closely graded series of staged human embryos.

MATERIALS AND METHODS

Serial sections of 105 human embryos (2.5- 28 mm) from stage 11 (24 postovulatory days) to stage 22 (54 days) were examined, and graphic reconstructions were prepared (Ta- ble 1). Only one embryo of stage ll was studied because this stage is well documented by other authors. Myotomes and their derivatives, which lose their original character early, were closely followed up to stage 16. In addition, the developing muscle fibers of the tongue were examined in two embryos of stage 19 and stage 23. The skeletal parts keep their segmented, original character longer, especially the vertebral column, but also the occipital part of the skull. Their development, therefore, was followed up to

TABLE I . Number of embryos examined and reconstructed at each stage

No. silver No. preparations reconstructions

Stage No. embryos included prepared

11 1 1 -

12 13 14 15 16 17 18 19 20 21 22 Total I

22 2 22 5 23 3

7 4 8 1 7 2 2 3 2 3 6 1 1

-05 20

-

-

-

1 3 2 4 4 3

2 2 1 1

24

-

stage 22. Stage 23 has already been described (Miiller and O’Rahilly, 1980b; O’Ra- hilly et al., 1983).

The fixative was mostly formalin, and dou- ble-embedding in celloidin and paraffin was usually employed. The sections were generally 5-20 pm in thinness and most were stained with hematoxylin and eosin, alum cochineal, or “azan”. Twenty of the specimens were impregnanted with silver.

Twenty-four graphic reconstructions (at least one a t each stage) were prepared according to the point-plotting method (see Gaunt and Gaunt, 1978, Chapter 5, for de- tails). Every fourth section was projected, usually at a magnification of 50. For the de- tails of the hypoglossal and upper cervical nerves, however, every section was used. In most instances, the reconstructions were made from photographic paper onto which the histologic sections had been projected di- rectly. By this method, varying densities in mesenchymal areas are much more clearly distinguished.

DEFINITIONS AND LANDMARKS

To aid in clarifying this difficult region, a summary of certain terms used is first provided .

Somites. In agreement with usual procedure, the numeral associated with a somite indicates a pair. Thus a 25-somite embryo is one that possesses 25 pairs of somites. The numeral 1 refers to the most rostral somite pair visible. However, “it is believed that most of the specimens thus encountered in embryos with more than 20 somites actually are second somites” (Arey, 19381, because a more rostrally situated pair has already regressed.

EMBRYONIC HYPOGLOSSAL NERVE AND OCCIPITAL SOMITES 239

Occipital somites. The four somites situated rostral to the first cervical nerve are designated occipital. A very small cavity, here termed “somitocoele”, is found in the earliest stages (9 and 10).

Occipital dermatomes, myotomes, and sclerotomes. The term dermatome is used in embryology for the superficial epithelial wall of a somite. Myotomic elements are already present at stage 12 as cuboidal, eosinophil cells. At stage 13, medial to the middle third of the dermatome, a sheet of very elongated somitic cells is found. These cells run parallel to the longitudinal axis of the neural tube and are collectively designated the myotome. Most of the remaining somitic cells constitute the sclerotome, which can be identified between stages 10 and 12. The cells of the sclerotomes, as pointed out by Blechschmidt (19571, cannot be assumed to be migrating medially; and this has been confirmed in the rat by Gasser (1979): “sclerotomal cells do not migrate during normal embryonic development. ”

Hypoglossal cord. The dermatomyotomic material of a t least the rostralmost three occipital somites forms a densely cellular prolongation that arises caudal to the rostral cardinal vein. This prolongation has been termed the hypoglossal cord (Hunter, 193513). The derniatomic cells cannot be distinguished from the myotomic cells, nor can possible contributions from neural crest and placodal material of the vagus be excluded.

Intersegmental arteries. By stage 13, intersegmental arteries accompany the developing spinal nerves and lie between the somites. One to three similar vessels develop in the occipital region and accompany hypoglossal roots. The most caudal of these is the primary hypoglossal artery. It runs between the last two occipital somites and accompanies the last root of the hypoglossal nerve.

Rhombomeres. At least seven rhombomeres can be identified at stages 13 and 14. Vagal fibers emerge from rhombomere 7, caudal to both of which the first occipital somite is found.

Hypoglossal ganglion. Ganglionic material related to the rarely seen dorsal roots of the hypoglossal nerve in certain hoofed animals is known as Froriep’s ganglion. Its existence in the human is extremely doubtful.

Pharyigeat arches. Three pairs of arches are present by stage 12, four by stage 13 (O’Rahilly, 1978). Pharyngeal arch 4 is largely occupied by the superior and inferior vagal ganglia, and the rostral cardinal (fu-

ture internal jugular) vein. For reasons elab- orated by Frazer (1923), the “fishy nomenclature” involved in the term “bran- chial” is not appropriate to mammals.

Median thyroidprimordium. In the median plane, the epithelial bud that indicates the future thyroid gland (O’Rahilly, 1983) serves as a landmark for the migration of the occipital somitic material.

OBSERVATIONS

Although the main observations relating to this study are based on stages 11-22, certain features of stages 9 and 10 will first be reviewed in order to complete the account.

Stage 9 (one to three pairs of som.ites; 1.5-2.5 mm; approximately 20 days)

Somites are first visible a t stage 9 (O’Ra- hilly, 1973). The one to three pairs of somites present a t this stage are presumed to be occipital. The first pair appears immediately caudal to the midpoint of the notochordal plate.

Stage 10 (4-12 pairs of somites; 2-3.5 mm; approximately 22 days)

As the number of somites increases during this and the next stage, it is no longer possible to assign a certain number to the occipital region. Somitocoeles are present but are no longer visible at the next stage. Sclero- tomic cells are distinguishable a t the ventromedial angle of the somite (see Fig. 25 of Corner, 1929).

Stage 11 (13-30 pairs of somites; 2.54.5 mm; approximately 24 days)

One embryo at the end of stage 11 (20 pairs of somites) was studied before proceeding to stage 12. A monograph on this specimen was published by Davis (1923), and certain disa- greements with his account will be mentioned in the Discussion below.

The rostral neuropore has barely closed. The hindbrain shows six distinct rhombomeres. The site of the seventh is indicated by the vagal-accessory neural crest. No indication of the hypoglossal nerve was seen.

At least three and possibly four pairs of somites are considered to be occipital, although it is only in the next stage that the neural crest for the cervical region is clearly delineated and can be used as a criterion for distinguishing the occipital from the cervical region.

Somite 1 is small and makes almost no contact with the surface ectoderm. It is situ-


ated immediately caudal to the vagal-accessory crest (Fig. la). Numerous mitotic figures are visible in its walls. Sclerotomic material is found in its caudal half. The dermatome of somite 2 is epithelioid and is in contact with the surface ectoderm. Mitotic figures are present in all regions of the somite. Sclero- tomic material is present between the com- pact part of the somite and the neural tube. Somite 3 is more regular in shape and resembles the following somite. Dermatomic and sclerotomic parts are evident (Fig. la). Medi- ally, the sclerotomic material follows closely the lateral wall of the neural tube (Fig. la’). Intersegmental arteries are present between somites 2 and 3, and between somites 3 and 4.

Stage 12 (21-29 pairs of somites; 3-5 mm; approximately 26 days)

Seven rhombomeres are usually present. Of the 22 specimens, three showed fine cellular strands arising from the basal lamina of the hindbrain and ending at the ventral border of occipital myotomes 2,3, and 4 (Fig. lb’). These strands, although they do not yet show nerve fibers, are in line with beginning nerve fibers within the neural tube, as seen in a silver preparation. Moreover, comparison with later stages indicates conclusively that these cellular strands represent the first signs of the hypoglossal nerve (Fig. lb, black dots).

Four occipital somites are present, as determined by relationships to (1) the concentra- tions of the cervical neural crest (which are caudal to the occipital region), and (2) the primordia of the hypoglossal nerve.

Somite 1 may still show a dermatome, which is frequently not in contact with the surface ectoderm but is always in intimate relationship to the vagal crest. In most instances, numerous mitotic figures are present, especially near the crest material. Somite 1, for example, is being transformed into loose mesenchyme with very dense cy- toplasm. The topographical relationships are shown in Figure l b (open arrow). Myotomic material is distinguishable.

Somites 2, 3, and 4 show the characteristic somitic features: dermatome, myotome, and sclerotome. A somitocoele is not evident in any of the occipital somites. The dermatome is still intact dorsally, whereas the ventral two-thirds are being transformed into mesenchyme. Sclerotomic material is wedged between the myotome and the neural tube but is not yet related to the notochord. The hypoglossal artery is visible between somites 3

and 4, and a second intersegmental vessel is present between somites 2 and 3.

An apparent rotation of the somites takes place between stages 11 and 12. The longitudinal axis of the dermatomyotome, as seen in cross section (Fig. lb’), comes to make a more acute angle with the median plane. Associ- ated with this, the dorsal surface of the body, as seen in cross-section, is changing from a gentle to a steeper curvature.

Stage 13 (30 or more pairs of somites; 4-6 mm; approximately 28 days)

At least seven rhombomeres are found. The efferent roots of cranial nerves 5,7,9,11, and 12 are present and are readily seen in the four silver-impregnated specimens. The neural crest of the accessory nerve is present for the first time (Fig. 8 of Miiller and O’Ra- hilly, 1980a), as are also the spinal ganglia. The motor nuclei of these nerves form a more or less continuous longitudinal sheet in the medial part of the floor plate.

The hypoglossal nerve consists of three to five roots (Fig. 2a). The cellular sheath of the first root is less distinct and hence this root can readily be missed unless silver impregnation has been used. The roots proceed laterally, penetrate the sclerotomic material, and end in or near the myotome. The cells of the hypoglossal nucleus occupy the same area as in the previous stage and form part of the general nuclear sheet (Fig. 3).

Four occipital somites were found in each of the reconstructed specimens (Figs. lc, 2a). In the less advanced embryos of this stage, dermatomes, myotomes, and sclerotomes are distinguishable. In 17 of the 22 specimens, the epithelioid character of the dermatomes has become lost. The myotomes are clearly differentiated and, in sagittal sections, are seen to be composed of longitudinal fibrils with central nuclei. The first myotome is present and is best seen in sagittally sec- tioned embryos. In the reconstructed embryo (Fig. lc) it is of reduced size in comparison with myotomes 2 to 4, which are fused. In the more advanced embryos, septa are beginning to form between occipital myotomes 2, 3, and 4. In some specimens, there appears to be a continuity between fibrils of adjacent myotomes. The descent of material from the occipital myotomes (the hypoglossal cord of the next stage) is beginning (Fig. 4). Sclero- tomic material is emerging from all the occipital somites, but a gap is still present between those of the right and left sides. In the sagittal series, at least the last occipital sclerotome is beginning to show dense and


loose zones, such as are found in the cervical region. Even in the advanced embryos, sclerotomic cells are not visible around the notochord. The latter, however, is surrounded by a cellular sheath that appears to arise from the perinotochordal mesenchyme. The longitudinal extent of the cellular sheath varies from one occipital somite (no. 4) to four occipital somites (nos. 4-1). Rostra1 to that level, no cellular sheath is found around the notochord. The cervical flexure of the brain is appearing and, associated with it, the notochord is beginning to bend.

Stage 14 (5-7 mm; approximately 32 days) The cervical flexure of the brain is more

pronounced (Fig. 2b). The accessory nerve is still surrounded by neural crest cells which are in contact with the material of the spinal ganglia. Dorsally, the spinal ganglia form a continuous column. The dorsal roots of a t least the cervical nerves are developing.

In the less advanced seven embryos, the roots of the hypoglossal nerve are still com- pletely separated and run almost laterally to the myotomes. In the remaining specimens, the roots andor rootlets unite, sometimes only unilaterally. In many instances, three components result, the most caudal of which is the thickest and accompanies the hypoglossal artery (Fig. 2c). The main trunk of the hypoglossal nerve remains behind the rostral cardinal vein and the fourth pharyngeal arch. The cells of the efferent nuclei of cranial nerves 5, 7, 9, 10, and 11 have separated from the neurons that give rise to the abducent and hypoglossal nerves. The cells of the former, visceral group lie medially and basally and constitute the ventromedial column (see Windle, 1970). The cells of nerves 6 and 12 form a ventrolateral column, and the latter (Fig. 2c) is in line with the spinal motor nuclei.

Indications of dermatomes can still be found in some specimens. In sagittally sec- tioned embryos, the first myotome is seen to be immediately adjacent to the superior vagal ganglion. Hence the first occipital somite produces a myotome. This and the following occipitocervical myotomes contain typically arranged myoblasts. In transverse series, the material of the dermatomyotomes can be followed as it migrates as far as the lateral pericardial wall (hypoglossal cord, HC in Fig- ure 2b, c; see also Figures 5, 7, and 9). The cord, which can be identified already in advanced embryos of stage 13, contains both myoblasts and undifferentiated cells. The

ventral tip of the cord extends rostrally for a variable distance into the lateral part of the future tongue. The longitudinal extent of the cord is best seen in coronal sections. The hypoglossal nerve may end blindly or may already send silver-impregnanted fibers into the hypoglossal cord. The mesenchyme of the fourth pharyngeal arch is looser than that of the cord.

The sclerotomes are now situated antero- laterally with reference to the neural tube (Fig. 2b, c), whereas they were located more strictly laterally a t the previous stage (Fig. 2a). The occipital sclerotomic material is organized into two portions, the larger of which (here designated a) is situated rostrally and seems to be derived from occipital somites 1- 3, whereas the smaller portion (p) represents sclerotome 4 (Fig. 2b, c). The rostral portion (a) has no connection with the mesenchyme of the otic capsule, which is now becoming differentiated. The two sclerotomic portions (a and 0) are separated by a cleft through which the caudalmost, thick root of the hypoglossal nerve and the hypoglossal artery run. The right and left caudal portions of the sclerotomic mass (p) are united across the median plane (Fig. 7). This sclerotomic contribution to the cellular sheath of the notochord increases and is identifiable by silver impregnation. Moreover, the cellular sheath now extends rostrally as far as the apex of the notochord, which is now flexed to almost a right angle.

Stage 15 (7-9 mm; approximately 33 days) The cervical nerves are formed by dorsal

and ventral roots, and they divide into dorsal and ventral rami. Cervical nerves 1-3 are interconnected and join the descending part of the hypoglossal nerve (Fig. 1Oa).

The rootlets of the hypoglossal nerve still possess a sheath of neural crest cells. More than twenty rootlets are present, and they are arranged in a variable number of groups (Fig. 10). The thick caudal root is still accom- panied by the hypoglossal artery. One of the more rostral roots runs dorsocaudally to occipital myotomic material. These silver-impregnated fibers are also detectable in the previous and in the following stage. Some of the descending rootlets give fibers to the occipital sclerotomic material. Frequent connections occur between the hypoglossal rootlets and the superior vagal ganglion (Fig. 10a). The rootlets converge to form two or three roots, which enter the sclerotomic material and unite after leaving it. The hypoglossal


Hypoglossal cord

Neural crest/ganglion

Notochord Dense par t of sclerotome


nerve now lies medial to the hypoglossal cord and accompanies it into the tongue (Fig. lob).

Typical myotomic cells are no longer visible a t the site of myotome 1 and are greatly reduced in number in myotomes 2-4. In the reconstructed embryos, the material of myotomes 2, 3, and probably 4 can be followed into the hypoglossal cord. In one instance, it is possible that a contribution from the first cervical myotome joins the hypoglossal cord (Fig. 10a). The cord descends lateral to the rostral cardinal vein and vagus (Fig. ll), and

then passes rostrally (Fig. 6) into the tongue (Fig. 10). Its density distinguishes i t from the mesenchyme of the pharyngeal arches and from that of the tongue. The hypoglossal cord does not yet reach the median plane.

The two former portions (a and 0) of the sclerotomic material have united (Figs. 10, 11) along the line of the caudalmost hypoglossal root and the hypoglossal artery. The longitudinal extent of the occipital sclerotomic material becomes shorter in the more advanced (Fig. 10a) as compared with the less- developed (Fig. 1Oc) specimens. The caudal

A 1-4 Pharyngeal arches

AL Alar lamina Ao Dorsal aorta Em Emissary foramen

Epiphysis cerebri % Hypochordal bow HCan Hypoglossal canal HC Hypoglossal cord J F Jugular foramen JT Jugular tubercle M Myotome N Notochord NP Neural process of

vertebra (occipital arch, or pila occipitalis in the occipital region)

1-4

Abbreviations Used in Figures

01 Olfactory disc or

Op Optic vesicle or cup Ot Otic disc or vesicle.

pit

Used for otic capsule in Figure 14

Parac Paracondylar process (transverse process)

Para- Part of the basal chor- plate dal surrounding

the notochord Ph Pharyngeal cavity PL Parietal lamina

Fig. 1. Graphic reconstructions of occipital somites and their constituents a t stages 11-13. The scheme of shading shown here applies also to Figures 2 and 10-12. In drawings a and b, the small circles represent individual neural crest cells, which are identifiable by their location and by their elongated shape. Drawings a, b, and c are from reconstructions of the left side but, in order to facilitate comparison with later figures, are shown as if they represented the right-hand side. Draw- ings a, a' , b, and b' are at the same scale, whereas drawing c is reduced. a. Stage 11 (no. 2053, 20 somites, 3.1 mm) with at least three occipital somites (1,2, 3) and two pharyngeal arches (Al , A2). The sclerotomic parts of the somites are shaded. a' . Transverse section (3-5-6) through line I in (a). The dermatomyotome lies immediately under the surface ectoderm. The sclerotomic material lies parallel to the wall of the neural tube. b. Stage 12 (no. 6097, 25 somitcs, 3.4 mm) with four occipital somites (1,2,3,4) and three pharyngeal arches (e.g., A2,

Post- Part of the basal otic plate between

the otic capsule and the parachordal part

S Somite TA Truncus arteriosus Th Median

primordium of thyroid gland

V Rostra1 cardinal vein

x Plane of section X, Y, The three Z successive,

immediately postcranial, loose sclerotomic areas

01 Sclerotomic material of the first three occipital somites

p Sclerotomic material of the fourth occipital somite

9 Glossopharyngeal neural crest

10 Vagal neural crest or ganglion

11 Accessory nerve 12 Hypoglossal

nucleus or roots or nerve

junction * Occipitocervical

A3). Myotomes arc now evident in addition to sclerotomes. An open arrow indicates a mass of mesenchyme derived from somites 1 and 2, and which is limited ventrally by the rostral cardinal vein (V). The material in question is believed to be derived from somites 1 and 2 because it is in continuity with them, but other origins cannot be excluded. Four roots of the hypoglossal nerve arc indicated by black dots. b'. Almost transverse section (1-6-3) through line I in (b). The somite has expanded and now shows a myotome (M). The dermatome is indis- tinct ventrally. A hypoglossal root (12) is seen in part of its course through the sclerotomic material. A solid arrow indicates the site of the future hypoglossal nucleus. c. Stage 13 (no. 836, more than 30 somites, 4 mm) with four occipital somites (1, 2, 3, 4) and four pharyngeal arches (e.g., A2, A3, A4). The sclerotomes and myotomes are shaded. Four roots of the hypoglossal nerve are shown.


, 0.5mrnI


region of the sclerotomic mass displays three dense zones (Fig. 13) that are more evident laterally. The intermediate zone separates the two groups of hypoglossal fibers. The dense areas are in line with similar zones in the cervical region (Figs. 8 and 13).

The notochord is ventral with respect to the fourth occipital and cervical sclerotomes, running between approximately their ventral third and dorsal two thirds. The notochord is surrounded by sclerotomic material in /3 and in the cervical region.

The rostral region of the hypoglossal cord lies lateral to aortic arches 3 ,4 , and 6 and to the truncus arteriosus (Fig. 1Oc). The vertebral artery is beginning to develop.

Stage 16 (8-11 mm; approximately 37 days) The roots and rami of the cervical nerves

are now better defined. The first three cervical nerves communicate with the hypoglossal nerve, and the communications of C.2 and 3 are presumed to be the precursors of the ansa cervicalis and its roots, which are better seen in the next stage (Fig. 12b). The trunk of the vagus between its superior and inferior ganglia includes accessory fibers that separate and descend parallel to the notochord.

Fig. 2. Graphic reconstructions of occipital somites and the hypoglossal nerve at stages 13 and 14. a. Stage 13 (no. 8066, 32-33 somites, 5.3 mm) with four occipital somites and four pharyngeal arches. The occipital sciero- tomic material forms a continuous mass laterally, indicated by an arrow, whereas the cervical sclerotomes remain separate, at least in their more condensed portions. The myotomes are not shown. Three hypoglossal roots and four spinal ventral roots (C.2-5) are evident. b. Stage 14 (no. 4245-6, 7 mm). The caudalmost three sclerotomes shown are cervical, whereas the two rostralmost masses (/3 and 01) belong to the occipital region. An arrow indicates an irregularly shaped mass of dense mesenchyme derived from occipital dermatomyotomes. Ven- trally, this hypoglossal cord (HC) extends onto the lateral side of the pericardium and is prolonged rostrally into the lateral portion of the future tongue. The vagus shows superior and inferior ganglia. Several hypoglossal rootlets unite and are then joined by a thick caudal root to form a trunk, which descends and ends close to the first cervical nerve. In Figures 2b and c and IOa, the hypoglossal cord is shown as if it were transparent, so that the underlying hypoglossal nerve and sclerotomes can be seen. c. Stage 14 (no. 6502,6.7 mm), more advanced. The hypoglossal cord (HC) is basically similar to that of the previous embryo, but its rostral prolongation is not as pronounced. The hypoglossal nerve descends further and is beginning to curve rostrally. The first cervical nerve is commencing to descend but retains its close relationship to the hypoglossal nerve. The lines marked x indi- cate the planes of section of Figures 5, 7, and 9.

Some little distance ventral to the rhomb- encephalon, the dura mater is becoming identifiable and surrounds the hypoglossal rootlets. The rootlets unite to form usually three main roots, which then penetrate the sclerotomic mass. In one instance, some ganglionic material was associated with the caudalmost rootlet but it is believed to be accessory in origin. The hypoglossal nerve is crossed laterally by the accessory nerve. At its termination, the hypoglossal nerve forms several small buds and, in some cases, the nerve reaches almost the tip of the tongue.

Between the occipital sclerotomic material and the first cervical sclerotome is a muscle primordium which is innervated by two hypoglossal rootlets. However, the other muscle primordia innervated by the hypoglossal are now situated in the tongue.

The dense zone (/3) seen in the caudal region (Fig. 13) of the sclerotomic mass at the previous stage now reaches the median plane in a restricted area, which lies ventral to the notochord and was named hypochordal bow (hypochordale Spange) by Froriep (1886). This condensed area appears also in the dense zones of the cervical region, but it was not present in stage 15. The more rostral dense area in Figure 13 is seen only in lateral sections; the intermediate dense area represented for stage 15 in Figure 13 and lying between the roots of the hypoglossal nerve is observed in some embryos only. The two sclerotomic portions (a and 0) are united more extensively. The rostral portion (a) appears to have grown further rostrally and becomes lost in the extradural mesenchyme.

The notochord may lie either relatively ventrally (Fig, 12a) with respect to the sclerotomes (as in the previous stage) or centrally as in advanced embryos of the next stage. Sclerotomic material surrounds the notochord in the entire occipital area.

Stage 17 (11-14 mrn; approximately 41 days) The hypoglossal nerve has changed only in

that the rootlets have lost their cellular sheaths. The nerve emerges from the occipital sclerotomic mass as either one (Fig. 12b) or two trunks. In the three reconstructed embryos, the ventral ramus of the first cervical nerve joins the hypoglossal nerve and, in one instance, its fibers were distinguishable as far as the level of the larynx but not as far as the superior root of the ansa (see below). In the embryo illustrated (Fig. 12b), the first cervical nerve lacks a ganglion. Ventral rami


2 and 3 unite to form the inferior root of the ansa, and the loop is completed by the superior root, which appears to descend from the hypoglossal nerve. Although it is difficult, even in silver preparations, to trace the course of the individual fibers, nevertheless it appears that, in the specimen illustrated (Fig. 12b), the superior root of the ansa is probably largely composed of C.2 and/or C.3 fibers.

Muscle groups are now beginning to take form in the tongue. Moreover, the portion of the hypoglossal cord seen formerly in relation to pericardium is being transformed into the infrahyoid muscles.

The arrangement of the sclerotomic material resembles that seen at the previous stage except that chondrification is beginning and that short neural processes are evident (Fig. 13). The caudal dense zone in the last occipi-

Figs. 3-9. Photomicrographs of silver preparations showing occipital somites and the hypoglossal nerve at stages 13-15. The planes of section of Figures 5, 7, and 9 are indicated on Figure 2c by the letter x.

Fig. 3. Stage 13 (no. 6473, 5 mm). The hypoglossal nucleus is marked (12), and occipital somite 1 is visible at the right. ~ 8 5 .

Fig, 4. Stage 13 (no. 6473,5 mm). At the right margin of the figure, the hypoglossal cord can be seen proceeding toward the rostral cardinal vein (V). x 51.

Fig. 5. Stage 14 (no. 6502, 6.7 mm). The dark band a t the right consists of dermatomyotomic material of occipital somites 1-3. The sclerotomic material is more medial. Various hypoglossal rootsirootlets are seen in cross- section. The rostral cardinal vein is adjacent to the superior vagal ganglion (10). ~ 8 5 .

Fig. 6. Stage 15 (no. 6504, 7.5 mm). Sagittal section through otic vesicle, pharyngeal arches, and pericardium. The termination of the hypoglossal nerve (12) is adjacent to the hypoglossal cord (HC). ~ 3 3 .

Fig. 7. Stage 14 (no. 6502,6.7 mm). The left and right vertical bands are dermatomyotomic material of occipital somites 1-3. A faint transverse, sclerotomic condensation reaches the notochord. X85.

Fig. 8. Stage 15 (no. 6504, 7.5 mm). Sagittal section through the pharyngeal region, a little medial to that of Figure 6, and showing a succession of dense areas (scler- atomic condensation) alternating with nerves. The last hypoglossal root is thick and can be seen above the first cervical nerve ((2.1). ~ 3 3 .

Fig. 9. Stage 14 (no. 6502, 6.7 mm). A slightly more caudal transverse section than that of Figure 7. The left and right third aortic arches are distinguishable above (dorsal to) the pharyngeal cavity. The hypoglossal nerve terminates immediately lateral to the rostral cardinal vein (V). ~ 8 5 .

tal somite (p) now meets its fellow of the opposite side in its whole extent, thereby re- sembling the appearance of the first cervical area. The densest part, which is ventral to the notochord, is the hypochordal bow. The more rostrally situated dense area does not reach the median plane. The dense zones in the cervical region give rise to neural processes (Fig. 13), which extend as short septa between the spinal ganglia. The neural arches are entirely fibrous. The cartilaginous areas begin to form in the centra (future bod- ies) of the cervical vertebrae. In the occipital region, the caudal dense zone (0) behaves like a neural arch (Fig. 14); distally it is fibrous and, what is even clearer in later stages, extends dorsally into a septum-like structure. In most parts of the occipital component of the future basal plate, chondrification has begun except in the dense areas. Cartilage is sparse in the median plane. Ventral to the notochord it forms a thin sheath that extends rostral to the point where the notochord leaves the sclerotomic material to lie in the roof of the pharyngeal cavity. The skeleto- genic material for the future basal plate reaches more rostrally than the caudal end of the otic capsule (Fig. 14).

The point where the notochord leaves the occipital region varies from one embryo to another. The angle formed between its occipital and cervical parts may be a gentle curve inside the occipital sclerotomic material, a gentle curve where it leaves 0, or rostral to and therefore inside the occipital area. At this stage, the future parachordal area of the basal plate is not identical with the postotic area.

Summary of stages 18-23 (13-31 mm; approximately 44-57 days)

The hypoglossal nerve resembles basically that seen at stage 17. By stage 23, a number of rootlets unite to form three to five roots. Usually two trunks enter an undivided hypoglossal canal and join to constitute the hypoglossal nerve. The ansa cervicalis and its various branches are well developed (Fig. 3 of Muller et al., 1981).

Within the tongue at stage 19, groups of myoblasts run transversely, longitudinally, and obliquely. By stage 23, myotubes are identifiable in the hyoglossus, although striations are not evident in the intrinsic muscles of the tongue.

In stage 19 (Fig. 14), the basal plate of the chondrocranium is well established. Large portions of the occipital region and of the

248 R. O'RAHILLY AND F. MULLER

Fig. 10. Graphic reconstructions to show derivatives of occipital somites and their relationship to nerves and vessels at stage 15. a. No. 6506 (7.5 mm). The two rostralmost sclerotomic areas (a and 6) seen at the previous stage (Fig. 2b, c) have now united to form one mass (arrow), which is penetrated by the hypoglossal roots. The hypoglossal cord (HC) forms an acute angle on the lateral side of the pericardium and then extends as far rostrally as the first pharyngeal arch, where it occupies the lateral portion of the developing tongue. The hypoglossal nucleus (unshaded) lies partly under cover of the superior ganglion of the vagus, with which it is connected by two small roots. The hypoglossal nerve follows the angular course of the hypoglossal cord and ends in the root of the developing tongue. The first three cervical

nerves unite and join the hypoglossal nerve. The bra- chial plexus is forming. b. No. 6506 (7.5 mm). The isolated hypoglossal cord and hypoglossal nerve, seen in situ in (a). c. No. 6504 (7.5 mm). The sclerotomic arrangement is similar to that of the previous embryo. Only the distal portion of the hypoglossal cord is shown. The first cervical ventral ramus is seen to join the hypoglossal nerve; contributions from (2.2 and C.3 are not included in the reconstruction. Aortic arches 3, 4, and 6 are out- lined. The second cervical segmental artery has been cut short, and the first cervical segmental artery is seen to accompany the first cervical nerve. Two primitive occipital intersegmental arteries are present; the more rostral is incomplete, the other is the hypoglossal artery.


Fig. 11. Schematic representation of occipital somites and hypoglossal cord. The shaded band on the outline of an embryo of stage 15 represents the block of tissue shown in the main drawing and seen from above (solid arrow). Some of the segmental pattern shown schemati- cally on the right side of the drawing can still be detected in the sclerotomic material and myotomes indicated on the left side of the figure. Rostra1 to the first cervical nerve (C.l), the last root of the hypoglossal nerve (12) is seen beside the hypoglossal artery. The sclerotomic mass consists of two portions (a and p), one of which ((3) contributes a bridge across the median plane. The hypoglossal cord (HC) is derived from occipital somites 1-3.

cervical vertebrae are cartilaginous. The former dense segments of the sclerotomes are still indicated by the fibrous parts of the neural arches in the vertebrae and in the occipital region, as well as by the septa that began to form in stage 17. They are best seen laterally and in sagittal sections. In the occipital area two such septa are present. The caudal one is broad and is connected with part (3; the other septum in the occipital region leads to the former zone CY rostra1 to the most anterior root of the hypoglossal nerve.

The hypochordal bow of the occipital region is transformed into cartilage. It is visible from the ventral side of the basal plate as a transverse bar that is clearly related to the former part 6. In one of the embryos a second bar-like protuberance is present at the ventral side of the basal plate in the same area to which the above-mentioned septum runs. It corresponds to the intermediate dense area

seen in stage 15 (Fig. 13, stage 15) and in some embryos of stage 16. Area (3 begins to form the paracondylar process (Macklin, 1921; jugular or transverse process, Lewis, 1920). The lamina alaris and a foramen em- issarium are present. The notochord trav- erses the occipital region obliquely and leaves it opposite the epiglottic eminence. Most of its occipital part is not covered by cartilage. The cartilaginous material lies ventral to the notochord. In some areas the basioccipital cartilage seems bipartite in coronal sections. The angle of the notochord lies between its occipital and vertebral parts. The otic capsule has enlarged caudally. The primordium of the parietal plate is mesenchymal and has little or no connection with the otic capsule. A cartilaginous commissure (Macklin, 1921) is not yet present between the basal plate and the otic capsule. The hypoglossal nerve forms two bundles within the cartilaginous hypoglossal canal.

In stage 20 (Fig. 141, the transformation of the fibrous material into cartilage has pro- gressed. However, the most caudal part of the notochord is still without a cartilaginous covering above the basal plate. The neural arch of part 0, i.e., the occipital arch (Lewis 1920; de Beer, 1937), has grown faster than the neural arches of the cervical vertebrae. The occipital portion of the chondrocranium is establishing connections with (1) the parietal lamina of the otic capsule, (2) the cap- sular part of the otic capsule (occipitocapsular commissure), and (3) the cochlear part of the otic capsule (basicochlear commissure). The axis of the cartilaginous hypoglossal canal is dorsoventral. In one embryo (that described by Lewis, 1920), a bipartite canal was present on the right side (Fig. 14). The occipital condyles are beginning to form, although the foramen magnum is not yet complete. The myelencephalon and spinal cord are being enclosed laterally by part 0 (pilae occipitales) before a transverse bridge (tectum posterius) closes the foramen in later stages. A cartilaginous bar is still present a t the site of the former hypochordal bow, The optic capsule is now connected to the basal plate by cartilage (the basicochlear commissure). Hence it is now possible to distinguish a postotic part and a parachordal part of the basal plate.

In stage 21, the neural arch of part (3 is shorter than those of the vertebrae. The former, which is the occipital arch (pila occipitalis), still forms the lateral border of the future foramen magnum without being connected dorsally to its fellow of the opposite side by a tectum posterius. The hypoglossal


Fig. 12. Graphic reconstructions to show the hypoglossal nerve a t stages 16 and 17. a. No. 6510 (10.1 mm). The first cervical nerve communicates by its ventral branch with the hypoglossal. The full extent of the accessory nerve (11) is shown. The hypoglossal cord is not

included. b. No. 6520 (14.2 mm). The ansa cervicalis is now beginning to form. The accessory nerve and hypoglossal cord are not included in this reconstruction, but the dense zone of the first cervical scierotome is shown.


,0.5mm,

Fig. 13. Schemes to summarize the relationship of the hypoglossal nerve to the derivatives of the occipital sclerotomes at stages 15-17. At stage 15, the areas associated with myotomes 2-4 are indicated (M2-4); somite 1 has lost its myotome. The hypoglossal artery and SUC- ceeding segmental arteries are also shown at this stage.

Subsequently, these vessels usually become lost. The future hypoglossal foramen (which is not yet a canal) gradually incorporates more of the hypoglossal roots, although it i s possible that the rostralrnost root disap- pears. Neural processes, NP (the precursors of the neural arch) become evident by stage 17.

canal contains two bundles of the hypoglossal nerve in most instances, as well as a venous plexus. In one case a bipartite hypoglossal canal was found. The lamina alaris is generally pierced by an emissary vein.

In stage 22 the jugular tubercle and tectum posterius form.

The occipital part of the basal plate at stage 23 has already been described in a detailed study of the chondrocranium (Muller and O’Rahilly, 1980b), and its relationship to the cervical column has been given particular attention (O’Rahilly et al., 1983).

DISCUSSION Hypoglossal nerve

The cellular sheaths of the hypoglossal roots are first identifiable during stage 12, a t and after 25 pairs of somites. These sheaths were seen also by Reiter (1944) in an embryo of 29 somites. No traces were found in an embryo (no. 2053) of 20 somites (stage 111, although Davis (1923) marked three areas as the primordium of the hypoglossal nerve in the same embryo (his Fig. 1).

The hypoglossal nucleus appears a t the end of stage 12 (Fig. lb’). According to Windle (1970), neurofibrillation in primary efferent neuroblasts, including the hypoglossal, occurs before 30 somites (stages 12 and 13).

The hypoglossal roots a t stage 13 contain silver-impregnated fibers. Each occipital myotome receives usually one root, as was found also by Hunter (1935a) in the rabbit. The

first hypoglossal root can easily be over- looked but is always identifiable in silver preparations. The roots begin to unite and form a hypoglossal nerve during stage 14 (Fig. 2b, c). The hypoglossal nucleus becomes separated from the general efferent column and lies more laterally, as also was pointed out by Windle (1970) who stressed the result- ing problems, here as elsewhere, in classifi- cation of functional components.

By stage 15, an idenfinite number of rootlets as well as roots are present. In some instances, one or more of the rostralmost rootlets from the hypoglossal nucleus enter the superior vagal ganglion. The caudalmost hypoglossal root (Fig. 13) is separated from the more rostra1 roots by a dense area (p). This relationship may have a bearing on the divided hypoglossal canal sometimes encountered in the adult: “ils sont quelquefois dou- ble” (Winslow, 1766). A complete bony division is found in about 15% of skulls, and tripartite and quadripartite canals have been recorded (Augier, 1931).

At stage 16, the hypoglossal and abducent nuclei are still separated. The formation of the general visceral (or dorsolateral) nuclei of nerves 5, 7, and 9-11 occurs, as was noted also by Windle (1970). Rostrally, a t stages 15-17, one or two hypoglossal rootlets inner- vate a muscle primordium that lies between the last occipital sclerotomes and the developing arch of the atlas. This primordium, which was observed also by Reiter (19441,


Fig. 14. Graphic reconstructions to summarize the further development of the occipital sclerotomes at stages 17-20. The otic capsule (mesenchymal at stage 17, cartilaginous at the later stages) is stippled. The upper left- hand drawing (a), stage 17, is a superior view. The next two drawings below, stage 19, are (b) superior and 6‘) inferior views. The basal plate has grown rostrally (as far as the adenohypophysis), and the hypochordal bow

appears to have been interpreted by him as representing the entire lingual musculature. That this is not the case is clear because, a t this time, the hypoglossal nerve itself has already reached the tongue (Fig. 12a) where muscle groups are forming.

Ganglionic material related to the rarely seen dorsal roots of the hypoglossal nerve has been described in certain hoofed animals and is generally known as Froriep’s ganglion. In the human, however, even Froriep (in 16

PL Parac

(HB) and paracondylar processes (Parac) have become evident. The right-hand drawings, stage 20, are (c) superior and (c’) inferior views. The otic capsule has united with the basal plate, thereby establishing the jugular foramen (JF). The right hypoglossal canal is bipartite superiorly [in drawing (c)] but not inferiorly [HCan in drawing (c’).

adults) found “in no case even the smallest sign of dorsal hypoglossal roots or of a related ganglionic rudiment” (Froriep and Beck, 1895). The ganglionic swelling found on the accessory nerve in this study and by Pearson (1939) is clearly not a Froriep’s ganglion, although considered as such by Streeter (1904, his Figs. 7, 11, and 12; 1912) and by Sensenig (1957). Windle (1970), however, refers to “a hypoglossal ganglion (Froriep)” at 8 mm, as well as to “a false appearance of . . . dorsal


rootlets.” Because spinal dorsal roots do not appear until stage 14, the identification of Froriep’s ganglion would not be possible before that time. Hence, the ganglionic material seen by Reiter (1944) a t 29 somites (stage 12) cannot be designated Froriep’s ganglion.

Some fibers leave the hypoglossal nerve and end in the hypoglossal cord lateral to the pericardial wall. Later, this part of the cord appears to form the infrahyoid muscles, which may perhaps receive a contribution from the hypoglossal nerve, as has been maintained for the adult by several workers (Testut and Latarjet, 1948).

The ansa cervicalis is found at stage 17 (Fig. 12b). Streeter (1904, his Figs. 8 and 9) illustrated the descendens cervicalis at 10 mm, and Pearson (1939) showed the ansa at 17 mm.

Head mesenchyme The mesenchyme of the head is derived

from (1) apparently’ unsegmented mesoderm, which arises in large part from the prechordal plate and probably also from the neural crest, and (2) segmented mesoderm, arranged as somites. Both types, however, form skeletal muscles, innervated by soma- tomotor nerves.

Boyd (1960) modified Tello’s account of myo- genesis as follows: (1) the premyoblast, not distinguishable from associated fibroblasts; (2) the myoblast, an elongated, uni- or mul- tinucleated cell without transverse striation; (3) the myotube, which shows some periph- eral striation but in which the nuclei are still central; and (4) the muscular fiber, with fully established transverse striation and periph- eral nuclei. In the present study with routine stains, myoblasts were detected in the tongue by stage 19; and, by stage 23, myotubes were seen in the hyoglossus, although striations were not observed in the intrinsic muscles of the tongue. Early in the fetal period (105- 170 mm Crown-Rump), a few small muscle spindles are present in the extrinsic muscles; and later (260-390 mm C-R), more and larger spindles are visible in both the extrinsic and the intrinsic muscles of the tongue (Kleiss and Kleiss, 1980).

The cellular notochordal sheath, which begins to appear during stage 13, is derived from the head mesoderm. This sheath would seem to correspond to Gasser’s (1976, his Figs. 2-5) “parachordal condensation”.

As a result of scanning electron microscopy of mouse embryos, it has recently been found that, prior to the formation of somites, the

presomitic mesoderm becomes organized into segmental units termed somitomeres (Tam et al., 1982). The morphogenesis of the somitomeres appears to be related to neurulation and, in the mouse, “the eight pair of somitomeres are the first to separate themselves from the first seven and form the first pair of somites visible a t the light-microscope level” (Meier and Tam, 1982). Although similar in- formation is not available in the human, it is of interest to note that the first occipital somite appears opposite the eighth neuromere.

Occipital somites At stage 9 (one to three pairs of somites),

all the somites present are presumed to be occipital. In stages 10 and 11, where more than three pairs are present, the somites cannot be given a regional designation; hence it is not possible to speak of occipital somites (because the number may not be constant). Of a 20-somite (stage 11) embryo (no. 2053, studied also by the present writers; Fig. la), Davis (1923) stated without further explana- tion that “the segments may be classified” as 3 occipital, 7 cervical, and 10 thoracic.

A number of authors have assigned the somites of early embryos to regions but have by no means always provided their justifica- tion for doing so. For example, in a 24-somite embryo (stage 12), Johnson (1917) described 2 (or possibly 3) occipital, 8 cervical, 12 thoracic, and 2 lumbar somites, but the reason for this interpretation is unclear, Bardeen and Lewis (1901) used the bases of the limb buds as their criterion: the upper limb opposite C.5-T.l, and the lower limb opposite L.l-S. 1. This procedure, although sometimes valid (e.g., in a 25-somite, stage 12 embryo, Figure 7 of Muller and O’Rahilly, 1980a), is not consistently reliable (e.g., a t stage 13, their Figure 8).

Ingalls (1907), in a 35-somite, stage 13 embryo, assigned the somites in relation to the spinal ganglia. This procedure may work well at stage 13, but at stage 12 (e.g., Fig. 7 of Muller and O’Rahilly, 1980a) the neural crest is merely irregular in outline and not segmented.

By the time that 25 somite pairs are present in stage 12, the first four somite pairs are here assigned to the occipital region because (1) the continuous column of neural-

‘The cranial extent of metamerism in vertebrates is unclear. In the mouse it has been possible by scanning electron microscopy “to count seven somitomeres in the cranial region” rostra1 to the first somite, “which forms from the eight somitornere” (Meier and Tam, 1982).


crest cells begins between somites 4 and 5 (Fig. 7 of Muller and O’Rahilly, 1980a), and (2) cellular sheaths for nerve roots are present in relation to somites 2 4 . It is believed that these sheaths are for hypoglossal roots, because even in less advanced specimens of stage 13 (Fig. lc), nerve fibers (as detected by silver) are present within these sheaths only. What are considered to be cervical nerve roots are not seen until later in stage 13 (Fig. 2a).

By stage 14, occipital sornites as such can no longer be distinguished. In sagittal sections, however, four occipital myotomes are visible rostral to the first cervical nerve. This nerve now indicates clearly the occipitocervical junction (asterisk in Fig. 2b, c).

An alternative interpretation is possible on the assumption that an additional, rostrally placed occipital somite has disappeared by stage 12. Thus, the first somite seen during stage 11 (Fig. l a ) and earlier could be transformed (dedifferentiated?) into dense mesenchyme (Fig. lb , open arrow). Some support for this view can be found in the literature.

It has been shown by Arey (1938, his Table 3) that the first somite is large (equals the second in size) in stages 9 and 10 up to nine somites. From 10 somites to the end of stage 11, it is generally much smaller than the second somite. In stages 12 and 13, Arey described it as “presumably missing through regressive loss”; hence, “in embryos with more than 20 somites”, the first ones “actually are second somites”.

Reiter (1944) maintained that “altogether five occipital somites could be shown” (“Ins- gesamt konnten funf Kopfsomiten nachge- wiesen werden”), a viewpoint shared also by Ludwig (1957). Sensenig (1957) believed that “the occipital bone is formed from at least four somites, with some indication that a fifth may be involved.’’

It should be kept in mind that important species differences occur. For example, the first somite may be incompletely separated from the unsegmented mesoderm, as in the rat (Butcher, 1929), and may therefore not even be included in the enumeration, as in a study of the rabbit (Hunter, 1935a).

Occipital myotomes and hypoglossal cord Four occipital myotomes are found at

stages 12 to 14, as was also observed by Sen- senig (1957) at stage 13. Reiter (1944), in agreement with his acceptance of five occipital somites, lists five occipitaI myotomes. By stage 15 in the present study, the first my-

otome is no longer visible in situ, and myotomes 2 to 4 are reduced in size.

The term “hypoglossal cord” was used by Hunter (1935b) in the chick for the “ventral extension from an area of condensed meso- dermal material” lying a t the ventral edges “of occipital and upper cervical myotomes.” (A further term, “sub-myotomic tract”, does not appear to be applicable to mammals; Bates, 1948.) In the human, the dense mesenchyme that forms the hypoglossal cord begins to spread out already in stage ll. In stage 12, it can be seen that most of the mesenchyme is derived from the dermatomes of the first two somites. By stage 14, it can be observed (Figs. 2b, 11) that the material emanates from the first three occipital somites. By stage 15 (Fig. 10a, b), it has received contributions from the last occipital and probably the first cervical somites. As the hypoglossal cord is elongating, the myotomes are disappearing and are probably contributing to the cord. Most of the cells of the cord are not yet myoblasts. The general topography and angulation of the hypoglossal cord in the human (Fig. 10a, b) resemble those of the chick (Fig. 4 of Hunter, 193513) and even more so those of the cat (Figs. 1, 2 of Bates, 1948). The importance of growth movements, such as those of the pharynx and heart, for the changing relations of the hypoglossal cord has been emphasized by Hunter (1935b) and Bates (1948).

The detailed studies of Hunter (1935b) on the chick and of Bates (1948) on the cat led these authors to believe that the hypoglossal musculature is derived from rostral somites, at least the occipital and probably the upper cervical. The present investigation is the first in which this basic conclusion has been adequately supported in the human. The part of the hypoglossal cord lying on the lateral wall of the pericardium comes to be innervated mostly by cervical nerves, and it later forms, or a t least contributes to, the infrahyoid musculature. It has not been possible to evaluate whether this component is derived from cervical somites only or also receives an occipital contribution. Moreover, it is not certain whether all the lingual musculature is derived from the hypoglossal cord or whether a component develops in situ.

The results of detailed morphologic studies of chick and cat embryos have here been confirmed in the human. Although experimental investigations have been largely con- fined to the chick embryo, the basic similarity in the early development of the hypoglossal


musculature in both avian and mammalian species renders it highly likely that this musculature is largely, if not entirely, derived from occipital (or occipital and cervical) somites.

Although the nuclei of cranial nerves 3 ,4 , 6, and 12 may be arranged in linear order (the somatic efferent column), the muscles supplied by these nerves have little in com- mon. The muscles of the bulb arise from three independent mesenchymal condensations: the premandibular (from the prechordal plate) and two from the maxillomandibular mesoderm (Gilbert, 1957). The lingual musculature, however, arises from occipital myotomes.

lntersegmerztal arteries The hypoglossal artery is the last occipital

intersegmental artery (Padget, 1954). It is clearly identifiable late in stage 12, where occipital somites and hypoglossal roots can be determined. It begins to disappear a t the time when the vertebral artery commences to form (stage 15). In rare instances the hypoglossal artery persists postnatally. It is usually part of a large branch of the internal carotid that forms the basilar artery, and one or both vertebral arteries are frequently either hypoplastic or absent (Nakayama et al, 1970; Carbonin et al., 1976; Khodadad, 1977). A persistent hypoglossal artery may even be associated with an aneurysm (Ko- dama et al., 1976).

Occipital sclerotornes The derivatives of the occipital sclerotomes

are the first portions of the skull to form. At stage 11 the lateral wall and dorsal rim

of the somite represent the dermatome (Fig. la’). At stage 12 the elongated ventral portion of the somite is not as clear-cut as the dorsal region, in which the myotome can be distinguished on the medial aspect of the dermatome (Fig. lb‘). By stage 13, the dermatome loses its epithelioid character in the occipital region, although in other areas it is still well developed (Blechschmidt, 1957; Fig. lob).

Occipital sclerotomes were found arising from each of the four occipital somites, and it is likely that these correspond to the four sclerotomes listed by Reiter (1944) for his somites 2-5. Initially (stage 13) they form a continuous column on each side. Each column then (stage 14) becomes subdivided into a rostral (a) and a caudal (0) part, and the right and left caudal portions unite in the

median plane. The caudalmost root of the hypoglossal nerve, as well as the hypoglossal artery, pass between a and 0. Part shows the essential features of a vertebra except a complete centrum: chondrification posteri- orly is limited to the tissue ventral to the notochord including the hypochordal bow. In addition, part 0 shows right and left neural processes (which arise from the dense zone) and paracondylar processes (which represent transverse processes). Bardeen (1908) also found occipital chondrification ventral to the notochord; and, although he did not mention a hypochordal bow for the occipital, he re- ferred to and illustrated a “hypochordal brace” for the atlas.

The future hypoglossal canal is situated rostral to part 0. The hypoglossal roots have become arranged very closely by stage 17 (Fig. 131, so that a persistent septum in the hypoglossal canal would have developed prior to that stage (e.g., a t stage 15, Fig. 13). This loss of material between the roots makes it impossible to identify the former limits between the first three sclerotomes.

In the occipital area of later stages, the basioccipital cartilage corresponds to the incomplete centrum of 0 and the chondrifying parts of a. The exoccipital cartilage includes the pilae occipitales (which correspond to the neural arches of 01, the jugular process (which is derived from the embryonic paracondylar process), and the jugular tubercle (probably from a). The occipital condyle arises from and may receive material from the loose area caudal to 0. Such a dual origin appears to correspond to that described by Sensenig (1957). It should be stressed that both components belong to the future exoccipital seg- ment, and hence a possible basioccipital contribution to the condyle (O’Rahilly et al., 1983) is a different question. The origin of the squamous part of the occipital bone is uncertain: alar lamina (transverse process, Lewis, 19201, neural process (occipital hem- iarch, Macklin, 1921), or perhaps from the parietal lamina. The possible contributions of neural crest to the human skull are not known. Although neural crest material is noted in the proximity of occipital sclerotomes at stage 11 (Fig. la), it is not certain whether it will contribute to the skull or form the sheath of the hypoglossal nerve, or both.

The occipital part of the basal plate is derived from sclerotomes and hence is segmental in origin. Nevertheless, segmentally arranged “focal enlargements” (Marin-Pad-


illa, 1979) were not observed in the present study.

In summary, Goethe’s vertebral theory of the skull (O’Rahilly et al., 1983) is basically correct in the posterior cranial region. Fea- tures comparable to a portion of a centrum, transverse processes, and neural arches can be identified even in an adult skull. It is perhaps not entirely certain that the neural arches reach the median plane at the tectum posterius (which ossifies differently).

ACKNOWLEDGMENTS

This work was supported by research grant HD-16702, Institute of Child Health and Hu- man Development, National Institutes of Health.

LITERATURE CITED

Arey, L.B. 1983 The history of the first somite in human embryos. Contrib. Embryol. Carneg. Inst., 27t233-269.

Augier, M. 1931 Squelette ckphalique. In: Traite d’Ana- tomie humaine, Vol. 1. P. Poirier and A. Charpy, eds. Masson, Paris, pp. 89-654.

Bardeen, C.R. 1908 Early development of the cervical vertebrae and the base of the occipital bone in man. Am. J. Anat., 8t181-186.

Bardeen, C.R. and W.H. Lewis 1901 Development of the limbs, body-wall and back in man. Am. J. Anat., I:1- 35.

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