GABA (gamma-amino-butyric acid) is the predominant neurotransmitter in the mammalian suprachiasmatic nucleus (SCN), with a central role in circadian time-keeping. We therefore undertook an ultrastructural analysis of the GABA-containing innervation in the SCN of mice and rats using immunoperoxidase and immunogold procedures. GABA-immunoreactive (GABA-ir) neurons were identified by use of anti-GABA and anti-GAD (glutamic acid decarboxylase) antisera. The relationship between GABA-ir elements and the most prominent peptidergic neurons in the SCN, containing vasopressin-neurophysin (VP-NP) or vasoactive intestinal polypeptide (VIP), was also studied. Within any given field in the SCN, approximately 40–70% of the neuronal profiles were GABA-ir. In GABA-ir somata, immunogold particles were prominent over mitochondria, sparse over cytoplasm, and scattered as aggregates over nucleoplasm. In axonal boutons, gold particles were concentrated over electron-lucent synaptic vesicles (diameter 40–60 nm) and mitochondria, and in some instances over dense-cored vesicles (DCVs, diameter 90–110 nm). GABA-ir boutons formed either symmetric or asymmetric synaptic contacts with somata, dendritic shafts and spines, and occasionally with other terminals (axo-axonic). Homologous or autaptic connections (GABA on GABA, or GAD on GAD) were common. Although GABA appeared to predominate in most neuronal profiles, colocalisation of GABA within neurons that were predominantly neuropeptide-containing was also evident. About 66% of the VIP-containing boutons and 32% of the vasopressinergic boutons contained GABA. The dense and complex GABAergic network that pervades the SCN is therefore comprised of multiple neuronal phenotypes containing GABA, including a wide variety of axonal boutons that impinge on heterologous and homologous postsynaptic sites.

The accessory olfactory bulb (AOB) is the primary target of the sensory epithelium of the vomeronasal organ (VNO), and thus constitutes a fundamental component of the accessory olfactory system, which is involved in responses to behaviour-related olfactory stimuli. In this study we investigated the characteristics of the AOB, VNO, vomeronasal nerves (VNNs) and caudal nasal nerve (CdNN) in pigs and sheep, species in which olfaction plays a key behavioural role both in the neonatal period and in adulthood. The patterns of staining of the AOB by the Bandeiraea simplicifolia and Lycopersicon esculentum lectins were the same in the 2 species, whereas the Ulex europeus and Dolichos biflorus lectins gave different patterns. In both species, lectin staining of the AOB was consistent with that of the VNNs, while the CdNN did not label any of the structures studied. The entire sensory epithelium of the pig was labelled by Ulex europeus and Lycopersicum esculentum lectins, and all 4 lectins used labelled the mucomicrovillar surface of the sensory epithelium in sheep.

The medial cortical surface of the mandible can be involved by tumour infiltration from the floor of the mouth. A detailed study of spread via accessory foramina through the edentulous alveolar crest has been previously undertaken, but no similar study has been carried out for the medial surface. In order to gain further appreciation of the mode of tumour spread, a study of the number and distribution of accessory foramina on the medial mandibular surface was performed on 89 mandibles. The number of foramina varied greatly from specimen to specimen. In the ascending ramus above the inferior dental foramen, 3 mandibles showed no foramina; the condylar section possessed the greatest proportion followed by the sigmoid and the coronoid. On the rest of the medial surface below the inferior dental foramen, all specimens showed at least 1 accessory foramen; the greatest concentration was in the middle third along the path of the inferior dental canal, followed by the upper third and the lower third section. Accessory foramina were repeatedly present at certain dedicated sites. The medial facing wall of the inferior dental foramen was found to be the commonest dedicated site (98.3%) followed by foramina on either side of the genial tubercles (71.9%), the digastric fossa (71.9%) and the median foramen above the genial tubercles (64%). The findings of this study are in keeping with the current observation that the lower border is least commonly involved in tumour spread. In view of the presence of accessory foramina along the inferior dental canal and especially on the medial facing wall of the inferior dental foramen, it is imperative to preclude tumour spread in this region prior to undertaking the conservative rim resection procedure. Medial to the symphysis the alveolar mucosa dips down almost to the level of the dedicated foramina in the vicinity of the genial tubercles. As a general rule the attached muscle forms a barrier to tumour spread except in the later stages, however, in irradiated mandibles resistance to spread has been previously reported to be diminished. Under these circumstances, it is possible that the numerous accessory foramina reported in this study could facilitate a direct pathway into the cancellous bone.

The expression of carbonic anhydrase (CA) in the quail metanephros was investigated during embryonic development. The immunohistochemical localisation of the isoenzymes CAII and CAIII was compared with the distribution of enzyme activity visualised by a histochemical cobalt-precipitation procedure. The developmental profile of CA activity was also evaluated by means of a biochemical method. The occurrence of a moderate and diffuse CAII immunostaining from the first developmental appearance of the metanephros anlage testified to an early expression of carbonic anhydrase. This finding is discussed in relation to the involvement of the enzyme in the morphogenetic mechanisms leading to the establishment both of cell polarity and epithelial phenotype. CA expression in the renal sites that are positive in adults proved to be developmentally regulated. In the collecting duct system, enzyme activity could not be identified until the time of hatching. No CA was detected at any stage examined at the sites where, in adults, enzyme occurrence has previously been interpreted as a membrane-associated CA isoform. The differentiating renal tubules displayed no CAIII immunoreactivity. It can be argued that the bulk of the enzyme activity in the embryonic metanephros is due to the cytosolic isoenzyme CAII.

The vocal ligaments insert at the anterior and posterior commissures of the larynx. These structures fulfil biomechanical functions, balancing the different elastic moduli of tendon, cartilage or bone and undergo age-related changes that may be responsible for voice changes with increasing age. The aim of this study was to analyse the insertion structures of the vocal ligaments by means of macroscopic, histological, immunohistochemical and electron-microscopic methods and to draw conclusions from age-related structural changes on a functional basis. Investigations were carried out on the larynges of 22 males and 15 females (aged 1–95 y). In adolescence, the insertion zone of the vocal ligament tendon, a dense network of connective tissue rich in sulphated glycosaminoglycans at the thyroid cartilage, is characterised by a layer between tendon and cartilage comparable to fibrocartilage. The insertion zone lacks a perichondrium. Collagen fibrils of the vocal ligament tendon penetrate directly into the thyroid cartilage. In the insertion area, the chondrocytes are surrounded by collagen fibrils, which show positive reactivity to antibodies against type I and type III collagen. Sulphated glycosaminoglycans are integrated between the collagen fibrils. In the area of the posterior glottis, elastic cartilage rests like a cap on the hyaline base of the arytenoid cartilage. There is no distinctive border between the structures. With increasing age, ossification of the laryngeal skeleton occurs, involving hyaline cartilage at the posterior glottis and hyaline and fibrocartilage at the anterior commissure. At the same time, a loss of sulphated glycosaminoglycans is observed inside the vocal ligament tendon. Advanced ossification of the laryngeal skeleton, particularly in the area of the commissures, an increasing loss of glycosaminoglycans in the vocal ligament tendon and changes in the elastic tissue reduce the elastic modulus between tendon, cartilage and bone, thus ‘stiffening’ the insertion zones, which could be one factor among others favouring voice changes with advancing age.

There is evidence that the endothelial cell (EC) glycocalyx is a significant determinant of vascular permeability, acting as a charge-size filter to permeant molecules. We have therefore examined its oligosaccharide composition in 3 classes of microvessel with differing permeabilities. EC in rat brain, retina and myocardium were labelled with a panel of lectins and subjected to a semiquantitative analysis. Surprisingly, no substantial differences were evident for any lectin labelling between the 3 microvessel types despite their marked morphophysiological diversity. In particular, all showed substantial sialic acid expression, with Maackia amurensis (MAA) labelling sialic acid in an α2–3 linkage to β-galactose and Sambucus nigra (SNA) recognising sialic acid in an α2–6 linkage to β-galactose. Arachis hypogaea (PNA) binding after neuraminidase digestion indicated the presence of Gal β1–3GalNAc attached to terminal sialic acid. The results therefore show that the sequences NeuNAc α2–3Gal β1–3GalNAc and NeuNAc α2–6Gal β1–3GalNAc are strongly expressed in the 3 microvessel types irrespective of their permeability properties. This homogeneity suggests that these lectin ligands may be involved in a common set of EC functions, e.g. cell[ratio ]cell and cell[ratio ]matrix interactions. However, we cannot rule out the possibility that glycocalyx differences may exist between vessels in the paracellular cleft which may alter its filtration properties.

The immunohistochemical location of glial fibrillary acidic protein (GFAP) was used to study the state of maturation of retinal astrocytes from an 18-mo-old infant and to compare it with the situation in the adult. Infant astrocytes showed intense GFAP immunoreactivity in the perikarya and possessed spindle-like enlargements in their processes, while in the adult immunoreactivity in the perikarya was scarce and the spindle-like enlargements were not evident. Two types of astrocyte were observed in adult and child retinas: elongated and star-shaped. In the adult, the star-shaped type tend to be more stylised and to have longer processes than in the infant. In the infant, numerous astrocyte cell bodies were observed over vessels, while in the adult these were scarce. In the infant, the star-shaped astrocytes made up a honeycomb plexus, but this was not fully developed. These results suggest that at 18 mo of postnatal development the retinal astrocytes are still increasing and growing into the astroglial structure found in adults.

The structure and relative degree of mineralisation of antler and pedicle bone of yearling red deer stags exposed either to low or high levels of environmental fluoride were determined by digital quantitative backscattered electron (BSE) imaging. Bone fluoride content (BFC) in antlers (845±86 mg F−/kg ash, arithmetic mean±S.E.M.) and pedicles (1448±154 mg F−/kg ash) of deer from a highly fluoride polluted area in North Bohemia (Czech Republic) were significantly higher (P < 0.001) than those of controls from uncontaminated regions in West Germany (antlers: 206±41, pedicles: 322±52 mg F−/kg ash). Mean (56.5±4.5%) and maximum (84.9±2.1%) mineralised bone area of the control antlers significantly (P < 0.05 and P < 0.001, respectively) exceeded the corresponding values for the N. Bohemian deer (43.3±1.3 and 73.3±1.9%, respectively), while the pedicles from the 2 groups did not differ significantly. In the pooled antler samples (n = 18), negative correlations existed between BFC and mean (rs = −0.62, P < 0.01) as well as maximum (rs = −0.69, P < 0.01) mineralised bone area. Morphological imaging revealed a decreased width and an increased porosity of the antler cortex in the N. Bohemian specimens. Mean (148.5±1.7) and maximum (154.2±1.7) BSE-signal intensities (= grey levels; range between a monobrominated (grey level 0) and a monoiodinated (grey level 255) dimethacrylate resin standard) of the antlers from the controls were significantly higher than those of the N. Bohemian deer (140.7±2.1 and 145.7±2.2, respectively ; P < 0.05 for both comparisons). In the pooled antler samples, negative correlations between BFC and mean (rs = −0.51, P < 0.05) as well as maximum (rs = −0.52, P < 0.05) BSE-signal intensities were observed. No significant differences in mineralisation density parameters were found for the 2 pedicle samples, and BFC and mineralisation density of the pooled pedicles were uncorrelated. Morphological imaging revealed bone mottling (denoting increased remodelling activity) and frequent occurrence of apparently increased osteocyte lacunae in some of the pedicles from the N. Bohemian deer. It is concluded that the reduced amount of mineralised bone in, and the lower mineralisation density of, the N. Bohemian antlers resulted from a fluoride induced disturbance of bone mineralisation. The rapid growth of antlers leads both to a high mineral demand and a high rate of fluoride uptake during antlerogenesis. This, and the limited lifespan of antlers, which does not allow for a compensation of a delay in the onset or progression of the mineralisation process, renders antler bone particularly susceptible to fluoride. Antlers are therefore considered a useful model for studying fluoride effects on bone formation. Furthermore, analysis of cast antlers enables a noninvasive monitoring of environmental pollution by fluorides.

The purpose of this investigation was to determine the amount of epineurium surrounding the recurrent laryngeal nerve (RLN) compared with a limb nerve, that to flexor hallicus longus (NFHL). Nerve samples were obtained from 10 adult dogs and studied using scanning electron microscopy and light microscopy to measure the relative proportion of epineurium and the relative proportions of adipose and collagenous tissue comprising the epineurium in both nerves. Significantly greater relative epineurial cross-sectional areas and adipose content were found in the RLN than in the NFHL. Based on observations on noncranial peripheral nerves, the findings indicate that the RLN is better protected against deformational forces associated with compression than stretching forces. The RLN may not be structured well for successful reinnervation after injury. The patterns observed for adipose tissue in RLN epineurial tissue appeared unique compared with those previously reported in peripheral nerves. The primary role associated with adipose tissue is to ‘package’ the nerve for protection. The RLN is considered to be a vital nerve in the body, as are other cranial nerves. The large proportions of adipose tissue in the epineurium may relate to the importance of protecting this nerve from injury.

We examined numerous ovarian follicles from 32–35 d virgin mice by transmission electron microscopy and light microscopic immunohistochemistry. No macrophages were seen, but various stages of apoptotic granulosa cells were encountered. Presumably a granulosa cell or its debris in an advanced stage of apoptosis was destined to be phagocytosed by adjacent normal-looking granulosa cells. Other granulosa cells of normal appearance were seen in the region of the zona pellucida in contact with and apparently phagocytosing atrophic oocytes. Such granulosa cells were characterised by the presence of gap junctions with other cells and frequently contained annular gap junctions in the cytoplasm. To confirm the lack of involvement of macrophages in the process of follicular atresia and elimination, specially prepared ovarian sections were incubated with antimouse macrophage monoclonal antibodies (F4/80, Mac-1, Mac-2). None of the follicles examined showed positive immunoreactivity with these antibodies. Atretic follicles may shrink and eventually disappear from the ovary as a result of repeated apoptosis and phagocytosis by granulosa cells. There is no evidence for the presence or involvement of macrophages in the atretic follicles, at least in prereproductive mice as examined.

The aim of this study was to determine whether a relationship exists between decrease in fecundity and structural changes in the antimesometrial endometrium of the mouse. Fecundity was calculated as the number of animals showing a placental sign/number of copulated animals ×100 (%). Structural changes in the endometrium were examined by electron microscopy. A negative correlation between age and fecundity was found. Fecundity was 50% at 7 mo of age. At this age, amorphous material appeared in the region between the basement membrane deep to the luminal epithelium and the subepithelial cells. This material was sometimes attached to the basement membrane. It increased in amount with advancing age, as fecundity decreased. The structure of the uterine luminal epithelial cells did not alter with age. The results indicated that decrease in fecundity with advancing age is correlated with the appearance of amorphous material beneath the basal lamina of the endometrial epithelium. It is suggested that this could impair communication between the luminal epithelium and the endometrial stroma, which plays an important role in implantation.

Major variations of the principal arteries of the upper limb have long received the attention of anatomists. These variations can be summarised as the presence of 2 main arteries traversing the cubital fossa, a deep (or normal) brachial artery in coexistence with a superficial brachial, radial or ulnar artery. Anastomosis between these arterial trunks at elbow level has been reported in 1–6% of cases as an incidental finding in studies on the major arterial variations of the upper limb (Quain, 1844; Müller, 1903; Poynter, 1922; Adachi, 1928; McCormack et al. 1953; Wankoff, 1962; Rodríguez-Baeza et al. 1995). Only a single report (Ljubomudroff, 1927) has dealt specifically with the anastomosis.

The anatomical pattern of the anastomosis has been classified into 2 or 3 types depending on different morphological details. Three types have been described, taking into account its length, calibre and form (Quain, 1844) or the positions of the origin and number of recurrent radial arteries (Ljubomudroff, 1927). Two types have been described on the basis of whether the anastomosis coursed anterior or posterior to the bicipital tendon (McCormack et al. 1953). The aim of this study was to revisit these specific morphological details in a statistically reliable sample in order to catalogue the variations of the anastomosis and to provide an embryological explanation.

We recently reported the results of applying immunohistochemical methods to examine the distribution of neuropeptides and neurotransmitter synthesising enzymes in the autonomic nerve fibres of specimens of human vas deferens obtained at vasectomy (Jen et al. 1999). As anticipated from previous neurohistochemical and electron microscopic studies (Baumgarten et al. 1971), the majority of nerves supplying the muscle coat are noradrenergic in type as demonstrated by the presence of tyrosine hydroxylase (TH) and dopamine beta hydroxylase (DbH) (Dixon et al. 1998), both of which are enzymes involved in the synthesis of noradrenaline.

In addition to the profuse intramuscular plexus of nerves, a second nerve plexus occurs at the base of the epithelium lining the vas deferens. These subepithelial nerves are immunopositive for vesicular acetylcholine transporter (VAChT) and therefore cholinergic in type, presumably serving a secretomotor function (Jen et al. 1999). Our immunohistochemical studies have also shown the presence of neuropeptide Y (NPY) in most of these subepithelial nerves in the human vas deferens with minor populations containing vasoactive intestinal peptide (VIP) and nitric oxide synthase (NOS), an enzyme involved in the formation of nitric oxide.

The Summer Meeting of the Anatomical Society of Great Britain and Ireland was held at the University of Glasgow from 13th to 15th July 1999. It included a symposium on ‘Neurobiology of the basal ganglia’. The following are abstracts of communications and posters presented at the meeting.

Most nerves are attached to the neuraxis by rootlets. The CNS–PNS transitional zone (TZ) is that length of rootlet containing both central and peripheral nervous tissue. The 2 tissues are separated by a very irregular but clearly defined interface, consisting of the surface of the astrocytic tissue comprising the central component of the TZ. Central to this, myelin sheaths are formed by oligodendrocytes and the supporting tissue is astrocytic. Peripheral to it, sheaths are formed by Schwann cells which are enveloped in endoneurium. The features of transitional nodes are a composite of those of central and peripheral type. The interface is penetrated only by axons. It is absent at first. It is formed by growth of processes into the axon bundle from glial cell bodies around its perimeter. These form a barrier across the bundle which fully segregates prospectively myelinated axons. Rat spinal dorsal root TZs have been used extensively to study CNS axon regeneration. The CNS part of the TZ responds to primary afferent axon degeneration and to regenerating axons in ways which constitute a satisfactory model of the gliotic tissue response which occurs in CNS lesions. It undergoes gliosis and the gliotic TZ tissue expands distally along the root. In mature animals axons can regenerate satisfactorily through the endoneurial tubes of the root but cease growth on reaching the gliotic tissue. The general objective of experimental studies is to achieve axon regeneration from the PNS through this outgrowth and into the dorsal spinal cord. Since immature tissue has a greater capacity for regeneration than that of the adult, one approach includes the transplantation of embryonic or fetal dorsal root ganglia into the locus of an extirpated adult ganglion. Axons grow centrally from the transplanted ganglion cells and some enter the cord. Other approaches include alteration of the TZ environment to facilitate axon regeneration, for example, by the application of tropic, trophic, or other molecular factors, and also by transplantation of cultured olfactory ensheathing cells (OECs) into the TZ region. OECs, by association with growing axons, facilitate their extensive regeneration into the cord. Unusually, ventral motoneuron axons may undergo some degree of unaided CNS regeneration. When interrupted in the spinal cord white matter, some grow out to the ventral rootlet TZ and thence distally in the PNS. The DRTZ is especially useful for quantitative studies on regeneration. Since the tissue is anisometric, individual parameters such as axon numbers, axon size and glial ensheathment can be readily measured and compared in the CNS and PNS environments, thereby yielding indices of regeneration across the interface for different sets of experimental conditions.

Anatomical Society of Great Britain and Ireland

Future meeting

24–26 July 2000. Department of Anatomy, University of Cambridge; this will be a tripartite meeting with the Anatomische Gesellschaft and the Nederlandse Anatomen Vereniging. This will include a symposium on the new anatomical basis of the emergence of behaviour, and one entitled ‘Anatomy: the challenges for the 21st century.’