Alpha-fetoprotein (AFP) is the major serum protein during development. AFP is one of the earliest proteins to be synthesised by the embryonic liver. The synthesis of AFP decreases dramatically after birth and only trace amounts are expressed in the adult liver. The tissue distribution of AFP in early human embryogenesis has not been defined. We have studied the expression pattern of AFP mRNA in human and mouse embryos by in situ hybridisation. In humans, AFP is expressed in the hepatic diverticulum at 26 d postovulation as it differentiates from the foregut endoderm (i.e. in the most primitive hepatocytes). It is also expressed in the endoderm of the gastrointestinal tract and in the yolk sac at this age. AFP is subsequently expressed in the mesonephros and transiently in the developing pancreas. In the mouse, no expression of AFP was observed in the mesonephros but other sites of expression were similar. Thus AFP has a distinct temporospatial expression pattern during the embryonic period and this differs between human and mouse species. It is interesting that AFP is expressed by tumours such as primitive gastrointestinal, renal cell and pancreatic tumours as well as those of hepatocyte origin. This distribution reflects the sites of AFP expression during development.

Newly developed concepts and methods for the quantification of fibre type regionalisation were used for comparison between all muscles traversing the ankle of the rat lower hindlimb (n = 12). For each muscle, cross-sections from the proximodistal midlevel were stained for myofibrillar ATPase and classified as type I (‘slow’) or II (‘fast’). For the 11 ‘fast’ muscles (i.e. all except soleus), the muscle outline and the position of each type I fibre were digitised for further computer processing. Two potentially independent aspects of type I fibre regionalisation were evaluated quantitatively: (1) the degree to which type I fibres were restricted to a limited portion of the total cross-sectional area (‘area-regionalisation’) ; (2) the extent and direction of the difference (if any) between the centre of the muscle cross-section and the calculated centre for the type I fibre cluster (‘vector regionalisation’). Statistical analysis showed that type I fibres were vector regionalised in practically all investigated muscles and area regionalised within most of them, the only consistent exceptions being peroneus brevis and peroneus digitorum 4, 5. In muscles with a high degree of area regionalisation the population of type I fibres also had a markedly eccentric intramuscular position (i.e. high vector regionalisation). A significant relationship was observed between the relative position of a muscle within the hindlimb (transverse plane) and the direction and degree of its type I fibre eccentricity. On average, the degree of type I fibre eccentricity was greater for muscles remote from the limb centre than for those situated more centrally. In addition, the intramuscular concentration of type I fibres was typically greatest towards the centre of the limb, the most striking exception being tibialis posterior. For the slow soleus muscle, which is centrally placed within the limb, our analysis concerned the type II fibres, which were found to be weakly vector regionalised but not significantly area regionalised. It is concluded that, within muscles of the rat's lower hindlimb, fibre type regionalisation is a general and graded phenomenon which may reflect differentiating (embryological?) mechanisms of a transmuscular significance. Furthermore, the analysis demonstrated the usefulness of our new methods and concepts for the quantification of fibre type regionalisation.

The glycan composition of the N- and O-linked oligosaccharides of the follicle (Sertoli) cells of the urodele amphibian Pleurodeles waltl testis were identified by lectin histochemistry, performed alone or in combination with enzymatic and chemical deglycosylation methods. The follicle cells were shown to contain: (1) Fuc, Galβ(1,4)GlcNAc, GalNAc and Neu5Acα(2,3)Galβ(1,4)GlcNAc in both N- and O-linked oligosaccharides; (2) Man in N-linked glycans; and (3) Galβ(1,3)GalNAc in O-linked sugar chains. The follicle cells at the pre- and postmeiotic stages showed some differences in the UEA-I-positive Fuc characterisation, suggesting differences in the glycan composition. In addition, the sequence Neu5Acα(2,6)Gal/GalNAc was shown in the follicle cells only after spermiation, in the sperm-empty lobules of the developing glandular tissue. These results suggest that the follicle cells modify their glycoprotein content, probably for the performance of new roles, as the spermatogenetic cells develop. Thus the follicle cells surrounding male germ cells at different spermatogenetic stages would contain different glycoproteins involved in specific roles during male germ cell proliferation and maturation.

There is an accumulating body of evidence suggesting that the periaqueductal grey (PAG) is involved in the pathophysiology of migraine. Positron emission tomography (PET) studies in humans have shown that the caudal ventrolateral midbrain, encompassing the ventrolateral PAG, has activations during migraine attacks. The PAG may well be involved not only through the descending modulation of nociceptive afferent information, but also by its ascending projections to the pain processing centres of the thalamus. In this study the intranuclear oncogene protein Fos was used to mark cell activation in the PAG following stimulation of the trigeminally-innervated superior sagittal sinus (SSS) in both cats and in nonhuman primates (Macaca nemestrina). Fos expression in the PAG increased following stimulation to a median of 242 cells (interquartile range 236–272) in the cat and 155 cells (range 104–203) in the monkey, compared with control levels of 35 cells (21–50) and 26 cells (18–33), respectively. Activation was predominantly in the ventrolateral area of the caudal PAG suggesting that the PAG is involved following trigeminally-evoked craniovascular pain.

Knowledge of the orientation of facet joints in the cervical and upper thoracic region is important for understanding the biomechanical properties and clinical conditions relevant to the neck. The study was undertaken on dry macerated bones from 30 adult male human vertebral columns. The orientation of the superior articular facets in relation to their inclination with the sagittal and transverse planes was examined between C3 and T3 vertebrae in each column. The linear dimensions of the superior articular facets and the width/height ratio were also calculated. The results show that all vertebrae at C3 level and 73% at C4 level displayed posteromedially facing superior articular facets. Similarly at T1 level (C7/T1 joint) and below, all columns showed posterolaterally facing superior articular facets. The level of change in orientation, from posteromedial to posterolateral facing superior facets, was not constant and occurred anywhere between C4 (C3/C4 joint) and T1 (C7/T1 joint). The change in orientation followed 2 different patterns, i.e. sudden or gradual. The C6 vertebra (C5/C6 joint) was the most frequent site to show the transition. The shape of the superior articular facets was circular to oval at C3, C4 and C5 levels and gradually changed to a transversely elongated surface at C7 and T1. These findings correlate well with various cervical movements and associated clinical conditions.

The purpose of this study was to investigate how didelphid marsupials have diversified in morphology of their claws and digital pads as they evolved different foraging preferences such as terrestrial, aquatic, and arboreal feeding behaviours. Both arboreal and more terrestrial didelphids have papillary ridges on the digital pads of the fore and hindfoot. In contrast, the papillary ridges on the pedal digital skin of the water opossum Chironectes minimus have been replaced by nonoverlapping, thickened epidermal scales. Chironectes also differs from the other didelphids studied in having finger tips with reduced claws and digital pads that are covered with raised epidermal scales having projecting, finger-like cones arranged radially around the perimeter of each scale. The reduced claws and unusual digit skin microstructure of Chironectes likely improve this animal's ability to recognise and identify live animal prey under water using only its sense of touch.

Retinoic acid, when administered to pregnant mice on d 11·0 of gestation, causes limb skeletal abnormalities consisting of reduced digital number, shortening of the long bones and delayed ossification. We show here that these effects are correlated with a decrease in cell proliferation within 5 h of retinoic acid administration, specifically in the posterior half of the distal limb bud mesenchyme, from which the distal skeletal elements are generated. There is a specific downregulation of Fgf4, a gene known to be involved in limb bud outgrowth and expressed only in the posterior part of the apical ectodermal ridge; Fgf8, which is expressed throughout the apical ectodermal ridge, is unaffected. The reduction in Fgf4 expression is not accompanied by downregulation of Shh, nor of its receptor and downstream target gene Ptc, suggesting that the skeletal reduction defects induced by retinoic acid are mediated specifically by FGF4-induced skeletogenic mesenchymal cell proliferation.

The infrequent mention of epipharyngeal pouches occurring in some species of bears indicates the scarcity of morphological and functional knowledge about these structures. In order to provide precise morphological data on the structure of these remarkable formations and to verify their taxonomic utility, the pharyngeal regions of 1 spectacled bear and 3 brown bears were examined. All these individuals possessed epipharyngeal pouches, which are tubular, blind-ending outpouchings of the caudodorsal pharyngeal wall equipped with respiratory epithelium and a thick layer of elastic fibres. While the spectacled bear and Ursus arctos syriacus possessed a single pouch on the caudodorsal wall of the nasopharynx, in Ursus arctos and Ursus arctos beringianus 2 unequally sized pouches were present. Two additional sacs of smaller size, representing outpouchings of the lateral pharyngeal wall, occurred in the spectacled bear. These findings prove epipharyngeal pouches to be constant and unique morphological features of the family Ursidae, the anatomical features suggesting involvement in the respiratory system most probably in important aspects of ursid phonation. This is the first description of epipharyngeal pouches in the spectacled bear.

The voltage-gated ‘glial’ sodium channel NaG belongs to a distinct molecular class within the multi-gene family of mammalian sodium channels. Originally found in central and peripheral glia, NaG has since been detected in neurons in rat dorsal root ganglia (DRG) and may play a role in Schwann cell-axon interactions. We have studied the presence of NaG-like immunoreactivity in the intact and injured human peripheral nervous system using a specific affinity-purified antibody. Nerve fibres in normal and injured peripheral nerves and normal skin exhibited intense NaG-immunoreactivity. Numerous NaG-immunoreactive nerve fibres surrounded neuronal cell bodies within postmortem control DRG, and in DRG avulsed from the spinal cord (i.e. after traumatic central axotomy). There were no significant differences in the pattern of NaG immunostaining between control and avulsed DRG, or with delay after injury. Generally, the neuronal cell bodies were only very weakly immunoreactive to NaG, indicating that the NaG immunoreactivity was predominantly in Schwann cells/myelin. In accord, we demonstrated NaG immunostaining in cultured human and rat Schwann cells, and in distal nerve after wallerian degeneration. NaG thus appears to be a useful new marker for Schwann cells in the human PNS, and a role in neuropathy deserves investigation.

The stimulation or ablation of cerebellar structures has produced a variety of visceral responses, indicating a cerebellar role in visceral functions. Studies using anterograde and retrograde tracing methods have revealed connections between the hypothalamus and cerebellar structures. The aim of this study is to investigate the cerebellar connections of the dorsomedial (DMH) and posterior hypothalamic nuclei using retrograde axonal transport of horseradish peroxidase (HRP). In the present study, micro-injection of HRP restricted within the borders of the DMH showed that the projections of this nucleus are not uniform throughout its extent. The posterior DMH receives projections from the cerebellum, whereas the anterior DMH does not. These projections were from the (greatest to least concentration) lateral (dentate), anterior interposed (emboliform), and medial (fastigial) cerebellar nuclei. In addition, both the anterior and posterior DMH receive projections from various areas of the brainstem which confirms earlier studies and provides detailed descriptions. This study also demonstrates the distribution of labelled neurons to cerebellar and brainstem nuclei following HRP injection into the posterior hypothalamic nucleus. It provides clear evidence for a direct cerebellar nuclei-posterior DMH and cerebellar nuclei-posterior hypothalamic nucleus connections. We suggest that the brainstem reticular nuclei and other connections, such as the solitary, trigeminal and vestibular nuclei, of both DMH and posterior hypothalamus may contribute to the indirect cerebellohypothalamic connections. These observations offer a new perspective on the question of how the cerebellum may influence autonomic activity.

Four supernumerary ossicle variations—the ossicle at the lambda, the parietal notch bone, the asterionic bone, and the occipitomastoid bone—were examined for laterality differences, intertrait correlations, sex differences, and between group variations in the samples from around the world. Significant laterality differences were not detected in almost all samples. In some pairs of traits, significant association of occurrence were found. Several geographic samples were sexually dimorphic with respect to the asterionic bone and to a lesser extent for the parietal notch bone. East/Northeast Asians including the Arctic populations in general had lower frequencies of the 4 accessory ossicles. Australians, Melanesians and the majority of the New World peoples, on the other hand, generally had high frequencies. In the western hemisphere of the Old World, Subsaharan Africans had relatively high frequencies. Except for the ossicle at the lambda, the distribution pattern in incidence showed clinal variation from south to north. Any identifiable adaptive value related to environmental or subsistence factors may be expressed in such clinal variation. This may allow us to hypothesise that not only mechanical factors but a founder effect, genetic drift, and population structure could have been the underlying causes for interregional variation and possible clines in the incidences of the accessory ossicles.

The pattern of expression of a variety of placental nitric oxide synthase isoforms has contributed to elucidating the regulatory mechanisms of nitric oxide (NO) synthesis during gestation. The maintenance of vascular tone, attenuation of vasoconstriction, prevention of platelet and leukocyte adhesion to the trophoblast surface, and possible participation in uterine blood flow seem to be the main functions of NO generated at the fetal-maternal interface in humans and mice. Extending this knowledge to other rodent species commonly used as laboratory animals, in this study we focus on NADPH-diaphorase activity and the distribution of nitric oxide synthase isoforms (NOS) in the trophoblast cells of Calomys callosus during different phases of pregnancy. NADPH-diaphorase activity was evaluated cytochemically and the presence of NOS isoforms detected by immunohistochemistry. These techniques were performed on pre- and postimplantation embryos in situ and in vitro, as well as in placentae on d 14 and 18 of pregnancy. Neither NADPH-diaphorase activity nor inducible or endothelial NOS isoforms were found in pre-implanting embryos except after culturing for at least 48 h, when some of the embryonic cells were positive for the diaphorase reaction. On d 6·5 of pregnancy, trophoblast cells showed intense diaphorase activity both in situ and under in vitro conditions. A positive reaction was also found in the different placental trophoblast cells on d 14 and 18 of pregnancy. The inducible NOS (iNOS) isoform, but not the endothelial isoform, was immunodetected in trophoblast cells from the placenta and from postimplantation embryos in situ and under in vitro conditions. These results strongly suggest the production of NO by the iNOS isoform in the trophoblast of Calomys callosus after embryo implantation. The data also emphasise a possible role for the trophoblast in producing and releasing cytotoxic molecules at the fetal-maternal interface.

We carried out immunohistochemistry and western blotting of fresh frozen sections and crude extracts from adult rat thyroids. The histochemical and immunoblotting studies were performed with P2X receptor antibodies from 2 different sources. P2X-immunopositive cells were identified by fluorescence double labelling and confocal microscopy. Results of the western blotting experiments showed double bands of approximately 70 kDa and 140 kDa for all 7 P2X receptor subtypes with both sets of antibodies. Histochemical stains with antibodies from both sources also gave essentially identical results. P2X1, P2X2 and P2X6 receptors were detected exclusively in vascular smooth muscle; P2X5 and P2X7 receptors were also present on vascular smooth muscle. Endothelial cells stained for P2X3, P2X4 and P2X7 receptors. Thyroid follicular cells displayed immunoreactivity for P2X3, P2X4 and P2X5 receptors. No immunostaining for P2X receptors was observed on C-cells. Possible roles for the broad expression of P2X receptor subtypes in the rat thyroid are discussed.

The motility of the avian cloaca is under neural control, but little is known about the neural network that accomplishes this function. This present study was designed to determine the distribution of nitric oxide-synthesising neurons in the pigeon cloaca by enzyme histochemistry for reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d). NADPH-d-positive staining was seen in the neurons and fibres in the cloaca. The highest density of nerve fibres was noted in the coprodeum and the lowest in the proctodeum. In the coprodeum, NADPH-d neurons were found singly, formed small groups of 2–10 neurons, or were seen in plexuses in the muscle layer, lamina propria, or around the arterioles. Several NADPH-d-positive neurons were also observed in the ganglia of the cloaca. NADPH-d fibres ran in the muscle layer, lamina muscularis mucosae and lamina propria, or surrounded blood vessels. The distribution pattern of acetylcholinesterase (AChE)-stained neurons and fibres in the cloaca was similar to that of NADPH-d. Double staining for NADPH-d and AChE showed colocalisation of the 2 enzymes in many neurons of the cloaca. Tyrosine hydroxylase (TH)-immunoreactive nerve fibres originating outside the cloaca were also noted. In the urodeum and proctodeum, neurons or fibres positive for NADPH-d, AChE or TH were scattered in the lamina propria. Nerve fibres immunoreactive for calcitonin-gene related peptide, galanin, methionine-enkephalin, substance P, and vasoactive intestinal peptide were found sparsely in the cloaca. Our results demonstrate that nitrergic neurons constitute a subpopulation which is closely associated with the cholinergic system in the pigeon cloaca.

It is well known that the internal carotid artery has no branches in the neck during postfetal life (von Lanz & Wachsmuth, 1955), but during embryonic development there are anastomotic connections between the internal carotid and basilar artery that rarely persist after intrauterine life (Hassen-Khodja et al. 1992; Widmann & Sumpio, 1992; De Caro et al. 1995). The most common anomaly found incidentally on angiography is a persisting trigeminal artery, but persistent hypoglossal, otic and proatlantal arteries can also be found (Reynolds et al. 1980; Ouriel et al. 1988; Salas et al. 1998). We report a rare example of a persistent hypoglossal artery in a 64 y old female cadaver, cause of death unknown, embalmed by standard mortuary procedures for dissection during the routine anatomy course. The persistent hypoglossal artery originated from the anteromedial wall of the left internal carotid artery 2.5 cm above the bifurcation (Fig.). It extended towards the hypoglossal canal. The diameter of its lumen was 1.5 mm. During its course in the neck, it was positioned anterior to the internal carotid artery and medial to the facial and lingual arteries, the posterior belly of the digastric muscle and the convexity of the hypoglossal nerve in the neck. At 1 cm above the convexity of the hypoglossal nerve in the neck, the artery gave rise to a branch that entered the skull through the carotid canal anteromedial to the internal carotid artery (Fig. 1) and joined the basilar artery. The medial branch of the hypoglossal artery crossed the internal carotid artery on its medial side and entered the skull together with the hypoglossal nerve and joined the basilar artery. We found no other anomalies in the cerebral arterial system.

The amphibian testis is a useful model because of its zonal organisation in lobules, distributed along the cephalocaudal axis, each containing a unique germ cell type. Sperm empty lobules form the so-called glandular tissue at the posterior region of the gonad. Androgen production is limited to the cells of the interstitial tissue surrounding lobules with spermatozoa bundles and to the cells of the glandular tissue. In this work, we have studied the distribution of terminal carbohydrate moieties of N- and O-linked oligosaccharides in the interstitial and glandular tissue of the Pleurodeles waltl testis, by means of 14 lectins combined with chemical and enzymatic deglycosylation pretreatment. Some differences in glycan composition between the interstitial and the glandular tissue have been detected. Thus in both tissues, N-linked oligosaccharides contained mannose, Gal(β1,4)GlcNAc, and Neu5Ac(α2,3)Gal(β1,4)GlcNAc, while O-linked oligosaccharides contained Con A-positive mannose, Gal(β1,3)GalNAc, Gal(β1,4)GlcNAc, Neu5Ac(α2,3)Gal(β1,4)GlcNAc, and WGA-positive GlcNAc. Fucose was also detected in both tissues. However, GlcNAc on N-linked oligosaccharides and GalNAc and Neu5Ac(α2,6)Gal/GalNAc on both N- and O-linked oligosaccharides were found only in the interstitial tissue. As glandular tissue cells arise from the innermost cells of interstitial tissue that surround lobules, the differences in the glycan composition of interstitial and glandular tissue shown in this work may be related to the start of androgen synthesis when steroid hormone (SH)-secreting cells develop.

The sperm head structural organisation of the koala and hairy-nosed wombat, and the effects of varying concentrations of the ionic detergent, Triton X-100, on its component parts, were determined by electron microscopy. Although alike in form between the 2 species, the sperm nucleus of the former, but not the latter, had nuclear vacuoles and appeared to be more easily dispersed by the Triton X-100. The structure of the acrosome of the spermatozoon was similar between the 2 species and, in both, previously undescribed thin posterior and lateral segments were found to be present. It is suggested that this thin segment may facilitate sperm-zona and/or sperm-oolemma binding and fusion.

Five discrete hypostotic cranial traits, tympanic dehiscence, ovale-spinosum confluence, metopism, transverse zygomatic suture vestige, and biasterionic suture, were investigated in 81 human population samples. Except for ovale-spinosum confluence, marked asymmetric occurrences of the bilateral traits were not detected in the majority of the samples. Significant intertrait association was observed mainly between the biasterionic suture and other sutural variations including accessory ossicles. The traits showing relatively consistent sex differences across diverse populations were tympanic dehiscence, which is predominant in females, and biasterionic suture in males. On a world scale, the 5 hypostotic cranial traits showed distinctive patterns of geographical variation. Different clinal variations within and between macrogeographical areas such as western and eastern parts of the Old World were found for the frequencies of the traits. The Ainu may be the most distinct outlier in the eastern Asian region on the basis of the incidence of the traits, especially the transverse zygomatic suture vestige. The interregional variation without reasonable adaptive value and nonadaptive shift of the possible outliers presented in this study suggest that the genetic background for the occurrence of these traits cannot be excluded completely.

A Tripartite Meeting of the Anatomical Society of Great Britain and Ireland, the Nederlandse Anatomen Vereniging and the Anatomische Gesellschaft was held at St John's College, University of Cambridge, from 24th – 26th July 2000. It included symposia on ‘The neuroanatomical basis of the emergence of behaviour’ and on ‘Anatomy, the challenges ahead’ An Anatomical Society Review Lecture was given by Professor Martin Johnson of the University of Cambridge, and a European Federation for Experimental Morphology (EFEM) Lecture by Professor Karl Zilles of the University of Düsseldorf. The following are abstracts of communications and posters presented at the meeting.

Facet capsule injury has been hypothesised as a mechanism for neck pain. While qualitative studies have demonstrated the proximity of neck muscles to the cervical facet capsule, the magnitude of their forces remains unknown owing to a lack of quantitative muscle geometry. In this study, histological techniques were employed to quantify muscle insertions on the human cervical facet capsule. Computerised image analysis of slides stained with Masson's trichrome was performed to characterise the geometry of the cervical facet capsule and determine the total insertion area of muscle fibres into the facet capsule for the C4–C5 and C5–C6 joints. Muscle insertions were found to cover 22·4±9·6 % of the capsule area for these cervical levels, corresponding to a mean muscle insertion area of 47·6±21·8 mm2. The magnitude of loading to the cervical facet capsule due to eccentric muscle contraction is estimated to be as high as 51 N. When taken in conjunction with the forces acting on the capsular ligament due to vertebral motions, these forces can be as high as 66 N. In that regard, these anatomical data provide quantitative evidence of substantial muscle insertions into the cervical facet capsular ligament and provide a possible mechanism for injury to this ligament and the facet joint as a whole.