The timing of the first appearance of lymph follicles and germinal centres in various lymph nodes, and the ways in which numbers of these and IgM-synthesising cells increase within the nodes, were investigated in male and female C57Bl/6N mice aged from 4 d to 16 wk. The lymphoid organs examined were the Peyer's patches, spleen, somatic (submandibular, deep cervical, brachial, axillary, inguinal and popliteal) and visceral (mesenteric and lumbar) lymph nodes. Primary follicles appeared in most somatic lymph nodes 6 d after birth. The number of follicles per node then increased rather sharply in larger lymph nodes and slowly in smaller nodes, up to 28 d of age, reaching a level which varied according to the location of the node. Thereafter, the number of follicles in the somatic lymph nodes increased only slightly to moderately, reaching a peak or plateau at 8–12 wk. In the mesenteric (ileocaecal) nodes, primary follicles first appeared at 12 d, then increased linearly during the suckling period and after weaning to reach a plateau at 8 wk of age. Germinal centres appeared in the submandibular and mesenteric nodes at 28 d and their numbers increased consistently in the latter, while remaining low in the former. The impact of possible ‘natural’ exogenous antigen stimulation of the various lymph nodes was estimated from the presence of IgM-synthesising cells and germinal centres. Differences between the patterns of age-dependent changes in the numbers of lymph follicles observed in the somatic and mesenteric lymph nodes during their ontogeny are discussed in relation to differences in the magnitude of the exogenous antigen stimulatory effect. We also found that the variations in the numbers of lymph follicles produced in somatic lymph nodes at different locations during the first 28 d after birth reflected differences in the dimensions of the body regions drained by a particular somatic lymph node at this stage of development.

The janus kinases (JAK) and signal transducers and activators of the transcription (STAT) pathway have been shown to be activated by a number of cytokines or growth factors and to play significant roles in the differentiation of various cell types. In the present study, we investigated the distribution of the JAK–STAT pathway using immunohistochemistry in the human epidermis. Each element of the pathway showed abundant and differential expression in the epidermis. The differential distribution of the elements was most strikingly observed in the horny keratinised cell and granular layers of the epidermis. JAK2, JAK3, STAT1 and STAT5 were expressed in high amounts, and JAK1, TYK2, STAT2, STAT3, STAT4 and STAT6 to a much lesser extent in the horny cell layer. JAK3, TYK2, STAT2, STAT3, STAT4 and STAT6 were more abundantly expressed in the granular layer than the lower layers of the epidermis. JAK1, STAT1 and STAT5 were expressed at almost the same levels in the various layers of the epidermis. These results show that elements of the JAK–STAT pathway are abundantly and differentially expressed in the epidermis. It is suggested that each element of the pathway may play a role at a distinct stage of keratinocyte differentiation.

The posterior hypothalamic nucleus has been implicated as an area controlling autonomic activity. The afferent input to the nucleus will provide evidence as to its role in autonomic function. In the present study, we aimed to identify the detailed anatomical projections to the posterior hypothalamic nucleus from cortical, subcortical and brainstem structures, using the horseradish peroxidase (HRP) retrograde axonal transport technique in the rat. Subsequent to the injection of HRP into the posterior hypothalamic nucleus, extensive cell labelling was observed bilaterally in various areas of the cerebral cortex including the cingulate, frontal, parietal and insular cortices. At subcortical levels, labelled cells were observed in the medial and lateral septal nuclei, the bed nucleus of stria terminalis, and various thalamic and amygdaloid nuclei. Also axons of the vertical and horizontal limbs of the diagonal band were labelled and labelled cells were localised at the CA1 and CA3 fields of the hippocampus and the dentate gyrus. The brainstem projections were from the medial, lateral and parasolitary nuclei, the intercalated nucleus of the medulla, the sensory nuclei of the trigeminal nerve, and various reticular, vestibular, raphe and central grey nuclei. The posterior hypothalamic nucleus also received projections from the lateral and medial cerebellar nuclei and from upper cervical spinal levels. The results are discussed in relation to the involvement of the posterior hypothalamic nucleus in autonomic function and allows a better understanding of how the brain controls visceral function.

The present immunoperoxidase cytochemical study describes the distribution of glial intermediate filament molecular markers, glial fibrillary acidic protein (GFAP) and vimentin, in the brain and spinal cord of the adult lizard, Podarcis sicula. GFAP immunoreactivity is abundant and the positive structures are mainly represented by fibres of different lengths which are arranged in a rather regular radial pattern throughout the CNS. They emerge from generally immunopositive radial ependymoglia and are directed from the ventricular wall towards the meningeal surface. The glial fibres give origin to endfeet which are apposed to the blood vessel walls and subpial surface where they form the continous perivascular and subpial glia envelopes, respectively. In the optic tectum and spinal cord, star-shaped astrocytes coexist with radial glia. In the spinal cord, cell bodies of immunopositive radial glia are displaced from the ependyma. While vimentin immunoreactive elements are almost completely absent in the brain except for a few diencephalic radial fibres, the spinal cord ependyma exhibits a clearly vimentin positivity and no GFAP staining. In the Podarcis CNS the immunocytochemical response of the astroglial intermediate filaments appears typical of mature astroglia cell lineage since it fundamentally expresses GFAP immunoreactivity. Moreover, this immunocytochemical study shows that the Podarcis fibre pattern with predominant radial glial cells is morphologically more immature than in avians and mammalians, a condition suggesting that reptiles represent a fundamental step in the phylogenetic evolution of vertebrate astroglial cells.

We used scanning electron microscopy to count the number of mucous gland openings in the tracheae and lower portion of the larynges of the rat, guinea pig, hamster, mouse and rabbit. Cells of the airway surface epithelium were removed by protease digestion better to visualise the gland openings. The distribution of glands was further studied by conventional histology and by PAS/Alcian blue staining of whole mounts. In all rodent species, gland openings in the larynx occurred with a frequency of 1–2 per mm2. Mice had no gland openings in their tracheae, and hamsters, only a handful. Rat tracheae contained 126±42 gland openings (±S.D.; n = 6) at a frequency of ∼ 0.6 per mm2 at the top of the trachea and ∼ 0.15 per mm2 at the bottom. Guinea pig tracheae contained 153±90 gland openings (±S.D.; n = 5), with 54% being in the top 40% of the trachea. In both rat and guinea pig, tracheal glands were found in the ventral aspect between the cartilaginous rings, and were absent from the dorsal membranous portion. Gland openings in most species were simple circles of ∼ 50 μm diameter. However, glands in the rat trachea generally opened obliquely into shallow (∼ 20 μm deep) oval troughs (∼ 150×75 μm), which had their long axes oriented from head to tail. In the rabbit, there was no evidence of tracheal or laryngeal glands histologically. However, the tracheal and laryngeal surfaces contained numerous pits (∼ 30 μm diameter) distributed evenly over and between cartilages at a frequency of ∼ 4 per mm2. These may correspond to the ‘nests’ of goblet cells described by others.

Cortical cerebellar basket cells are stable postmitotic cells; hence, they are liable to endure age-related changes. Since the cerebellum is a vital organ for the postural control, equilibrium and motor coordination, we aimed to determine the quantitative morphological changes in those interneurons with the ageing process, using unbiased techniques. Material from the cerebellar cortex (Crus I and Crus II) was collected from female rats aged 2, 6, 9, 12, 15, 18, 21 and 24 mo (5 animals per each age group), fixed by intracardiac perfusion, and processed for transmission electron microscopy, using conventional techniques. Serial semithin sections were obtained (5 blocks from each rat), enabling the determination of the number-weighted mean nuclear volume (by the nucleator method). On ultrathin sections, 25 cell profiles from each animal were photographed. The volume density of the nucleus, ground substance, mitochondria, Golgi apparatus (Golgi) and dense bodies (DB), and the mean surface density of the rough endoplasmic reticulum (RER) were determined, by point counting, using a morphometric grid. The mean total volumes of the soma and organelles and the mean total surface area of the RER [SN (RER)] were then calculated. The results were analysed with 1-way ANOVA; posthoc pairwise comparisons of group means were performed using the Newman-Keuls test. The relation between age and each of the parameters was studied by regression analysis. Significant age-related changes were observed for the mean volumes of the soma, ground substance, Golgi, DB, and SN (RER). Positive linear trends were found for the mean volumes of the ground substance, Golgi, and DB; a negative linear trend was found for the SN (RER). These results indicate that rat cerebellar basket cells endure important age-related changes. The significant decrease in the SN (RER) may be responsible for a reduction in the rate of protein synthesis. Additionally, it may be implicated in a cascade of events leading to cell damage due to the excitotoxic activity of glutamate, which could interfere in the functioning of the complex cerebellar neuronal network.

Both the fetal and maternal microvasculature of bovine placentomes was examined by scanning electron microscopy of vascular casts. So far the development of the vascular architecture of the bovine placentome in early gestation has only been studied 2-dimensionally due to technical difficulties arising from the fragility of the early placental blood vessels. Repeated experiments led to the selection of the microvascular corrosion casts presented here. The vasculature of the maternal compartment is supplied by large caruncular stalk or spiral arteries, which release short maternal stem arteries. In the 3rd month of gestation, these arteries branch into several arterioles at their base, thus providing the vascular framework for the lower part of the septal walls of the primary crypts. In the 4th month, due to progressive longitudinal growth of the stem arteries, branching into arterioles occurs not only at the base, but over the whole length of the stem arteries. These arterioles supply the capillary complexes of the septa which resemble the major part of the septal vasculature and face the secondary crypts. Further indentation results in the formation of tertiary crypt capillary complexes, encircling the earlier secondary unit. From the 6th month of gestation the architecture resembles the fully developed maternal placenta with stem arteries running directly to the fetal side to branch into 4 to 6 arterioles, which turn back to enter secondary and tertiary septa. Maternal venules, collecting the blood from the capillary bed of secondary and tertiary septa, converge onto stem veins leaving the caruncle via branches of the uterine vein. The fetal part of the placentome is supplied by the cotyledonary arteries, which branch into fetal stem arteries that are the tributary to single villous trees. Over their whole course towards the maternal side, these give off arterioles entering secondary villi. The tertiary or terminal villous vasculature consists of capillaries, which are organised in serial capillary loops. This system is progressively elaborated in the course of gestation. In the 4th month there are only finger-like loops, whereas from the 6th month large fan-like structures can be observed. In early gestation the maternal and fetal blood vessels meet predominantly in a countercurrent fashion, changing to the less efficient crosscurrent exchange when the tertiary unit develops. These results indicate the development of a highly elaborated fetomaternal villous-crypt exchange system, already established in the 1st half of gestation, thus meeting the increasing needs of the fetus.

Motor neurons are lost during embryonic development, but it remains controversial whether motor neuron cell death occurs during postnatal life. In this study we investigated the effect of postnatal maturation on the number of intact spinal motor neurons in the rat using retrograde labelling with model-based counting, and an unbiased stereological counting technique. To determine the number of motor neurons innervating a specific forelimb muscle in rats of different postnatal ages FluoroGold was injected into the flexor carpi radialis. Before postnatal day 21 there were higher numbers of retrogradely labelled motor neurons than in adult rats, suggesting a ‘loss’ with postnatal maturation. This loss may be attributed to tracer diffusion to adjacent muscles and to the permeability of the muscle spindle capsule in younger animals. To obtain an unbiased estimate of the number of motor neurons in the C7 and C8 segments of the postnatal rat cervical spinal cord the fractionator/optical disector counting technique was used. This method did not show a loss of spinal motor neurons between birth and adulthood. The main conclusion from this study is that there is no loss of spinal motor neurons during postnatal maturation.

Biomechanical experiments on isolated hip joints have suggested that the transverse ligament acts as a bridle for the lunate articular surface of the acetabulum during load bearing, but there are inherent limitations in such studies because the specimens are fixed artificially to testing devices and there are no modifying influences of muscle pull. Further evidence is thus needed to substantiate the theory. Here we argue that if the horns of the lunate surface are forced apart under load, the ligament would straighten and become compressed against the femoral head. It would thus be expected to share some of the features of tendons and ligaments that wrap around bony pulleys and yet previous work has suggested that the transverse ligament is purely fibrous. Transverse ligaments were removed from 8 cadavers (aged 17–39 y) and fixed in 90% methanol. Cryosections were immunolabelled with antibodies against collagens (types I, II, III, VI), glycosaminoglycans (chondroitins 4 and 6 sulphate, dermatan sulphate, keratan sulphate) and proteoglycans (aggrecan, link protein, versican, tenascin). A small sesamoid fibrocartilage was consistently present in the centre of each transverse ligament, near its inner surface at the site where it faced the femoral head. Additionally, a more prominent enthesis fibrocartilage was found at both bony attachments. All fibrocartilage regions, in at least some specimens, labelled for type II collagen, chondroitin 6 sulphate, aggrecan and link protein, molecules more typically associated with articular cartilage. The results suggest that the ligament should be classed as containing a ‘moderately cartilaginous’ sesamoid fibrocartilage, adapted to withstanding compression. This supports the inferences that can be drawn from previous biomechanical studies. We cannot give any quantitative estimate of the levels of compression experienced. All that can be said is that the ligament occupies an intermediate position in the spectrum of fibrocartilaginous tissues. It is more cartilaginous than some wrap-around tendons at the wrist, but less cartilaginous than certain other wrap-around ligaments, e.g. the transverse ligament of the atlas.

Ischaemia and reperfusion phases represent critical events during liver transplantation. The purpose of this study was to describe morphological alterations of both vascular and parenchymal compartments after ischaemia and reperfusion and to evaluate the possible relationship between morphometric parameters and biochemical/clinical data. Three needle biopsies were drawn from 20 patients who underwent orthotopic liver transplantation. The first biopsy was taken before flushing with preservation solution, and the second and the third to evaluate respectively the effects of cold ischaemia and of warm ischaemia/reperfusion. Biopsies were examined by an image analyser and morphometric parameters related to the liver parenchyma were evaluated. At the second biopsy we observed a decrease of the endothelium volume fraction while the same parameter referred to the sinusoidal lumen achieved a peak value. The hepatocytes showed a lower surface parenchymal/vascular sides ratio. This parameter was reversed at the end of the reperfusion phase; furthermore the third biopsy revealed endothelial swelling and a decreased volume fraction of the sinusoidal lumen. The results quantify the damage to the sinusoidal bed which, as already known, is one of the main targets of cold ischaemia; warm ischaemia and reperfusion accentuate endothelial damage. The end of transplantation is characterised by damage chiefly to parenchymal cells. Hepatocytes show a rearrangement of their surface sides, probably related to the alterations of the sinusoidal bed. In addition, the fluctuations of morphometric parameters during ischaemia/reperfusion correlate positively with biochemical data and clinical course of the patients.

Anatomical Society of Great Britain and Ireland

Future meeting

11–13 July 2001. University College, Dublin. Summer meeting, including a symposium on ‘The respiratory system’.

It is currently thought that New World monkeys, prosimians, and humans are the only primates to possess vomeronasal organs (VNOs) as adults. Recent studies of the human VNO suggest that previous investigations on Old World primates may have missed the VNO. We examined nasal septa from the chimpanzee (Pan troglodytes) grossly and histologically for comparison with nasal septa from humans, Old World monkeys (Macaca fascicularis, M. nemistrina) and prosimian primates (Microcebus murinus, Otolemur garnettii). Grossly, chimpanzees had depressions on the nasal septum similar to fossae reported anterior to the VNO openings in humans. Histologically, chimpanzees and humans had bilateral epithelial tubes which were above the superior margin of the paraseptal cartilages (vomeronasal cartilage homologue). The epithelial tubes had a homogeneous ciliated epithelium. These structures were thus positionally and structurally identical to the human VNO and unlike the well-developed prosimian VNOs which were surrounded by vomeronasal cartilage. Macaques had no structures which resembled the VNO of either the prosimians or humans. The results demonstrate that the VNO is present postnatally in the chimpanzee and is almost identical to the human VNO in its anatomical position and histological structure. This in turn suggests that the reported absence of the VNO in at least some adult Old World primates is artifactual, and that further study may provide evidence for its existence in other species.

During surgical exposure of the carpal tunnel it is possible to injure the neurovascular structures closely related to the flexor retinaculum, such as the superficial palmar arch and the communicating branch between the ulnar and median nerves. Because of the importance of these structures and with the purpose of increasing knowledge of anatomical details concerning to their location, a biometric study was performed on the retinaculum and the communicating branch, and between the communicating branch and the distal wrist crease, as well as between the retinaculum and the superficial palmar arch. We dissected 56 hands from 28 Brazilian formalin-preserved cadavers of both sexes (24 male) at the Federal University of São Paulo–Escola Paulista de Medicina, Brazil. The communicating branch was observed in 96.4% of cases and the superficial palmar arch in 78.6%. The communicating branch was found between the common palmar digital nerve of the 4th interosseous space (from the ulnar nerve) to the homonymous nerve of the 3rd interosseous space (from the median nerve). In males, the distance between the distal wrist crease and the site where the communicating branch originates from the ulnar component had an average of 33.9±5.5 mm on the right side and 30.2±8.2 mm on the left. The distance between the distal wrist crease and the junction of the communicating branch with the common palmar digital nerve of the 3rd interosseous space was 43.6±6.9 mm on the right and 40.2±6.2 mm on the left side. Conversely, in 14.8% of cases (1 female), the communicating branch was observed to emerge from the common palmar digital nerve of the 3rd interosseous space. The distance between the retinaculum and the superficial palmar arch in the axial line of the 4th metacarpal bone was on average 7.3±4.3 mm on the right and 8.3±3.5 mm on the left side. At the same level, the distance between the retinaculum and the communicating branch was 6.2±3.7 mm on the right side and 5.1±2.8 mm on the left. These results can be used as a reference during surgical procedures in the palmar region.

The expression of carbonic anhydrase in the chorioallantoic membrane (CAM) of the chick embryo was investigated by means of the histochemical localisation of the enzyme catalytic sites and the immunohistochemical identification of its isoenzymatic forms. The results show that carbonic anhydrase is developmentally expressed in a subset of cells both in the ectodermal and the endodermal epithelium. The distribution patterns from both methodological approaches indicated that carbonic anhydrase is a marker of the villus cavity cells and the mitochondria-rich cells in the ectodermal and the endodermal epithelium, respectively. Such a cell-specific pattern of the enzyme expression provides a further contribution to characterising the heterogeneous cell population of the chick CAM and supports specific functional involvement for the distinct cell types in CAM-mediated processes, such as calcium transport, maintenance of acid-base balance and water and electrolyte reabsorption, during chick embryogenesis.

The pulmonary intravascular macrophages (PIMs) of domestic ungulates are recognised by their specific surface coat, consisting of linearly arranged globules along the external leaf of the plasma membrane. The coat is sensitive to in vitro digestion with lipolytic lipase (LPL), intravenous heparin and clinical exposure to halothane anaesthesia. The sensitivity to these experimental manipulations suggests that the globules of the coat are predominantly composed of lipoproteins (LDL). The present administration of oestradiol proprionate in castrated male calves potentiated the translocation of the surface coat into the endocytotic pathway of the PIMs. Concurrently with mobilisation of the coat, the plasma membrane was thrown into prominent arrays of lamellipodial extensions. The sprawling macrophages made extensive adhesive contacts with the lining endothelium of the capillaries. Consequently, the endothelial cells were highly attenuated and precariously maintained the integrity of the vascular wall. At some focal points, the vascular wall was penetrated by the filopodial processes of PIMs, which protruded into the perivascular space. Furthermore, there were signs of neovascularisation in the form of overt mitotic changes, sprouting and precursor capillary formation. It is conceivable that the evolving profile of angiogenesis is due to the vascular endothelial growth factor (VEGF) paracrine function of PIMs. Endothelial cell specificity has been considered an important advantage of VEGF for neovascularisation. It allows pleotrophic response of endothelial cells to proliferate and to assemble into endothelial tubes.

The acetabular point was analysed by studying human pelvic bones from 326 individuals ranging from newborns to age 97 y. The bones were categorised into 3 groups according to the degree of fusion for the 3 elements of the pelvis: nonfused (59), semifused (5) and fused (262). The acetabular point in immature pelvic bones is clearly represented by the point of the fusion lines for each bony element at the level of the acetabular fossa. In adult pelvic bones the acetabular fossa has an irregular clover-leaf shape, the superior lobe being smaller than the anterior and posterior lobes. Cross-sectional analysis of acetabular morphology suggested that the acetabular point in adult pelvic bones is always represented by the indentation between the superior and the anterior lobes of the acetabular fossa.

The aim of the present study was to estimate mean neuronal volume and absolute size distributions of the neocortical neurons in brains from controls and AD patients using stereological methods based on unbiased principles to determine whether changes in absolute cell size are part of the neuropathological pattern of Alzheimer's disease. The neocortex of 8 patients with Alzheimer's disease (AD), mean age 81·1 (68–94) y was compared with 9 nondemented controls, mean age 80·9 (65–101) y. The brains came from Johns Hopkins University Hospital (JHUH) in Baltimore, USA, the Netherlands Brain Bank (NBB), and from a large brain repository in Denmark. The rotator method was used to obtain an estimate of cell volumes providing absolute size distributions of the volume of both cell perikaryon and cell nuclei. The geometric mean volume of cell nuclei in neocortical neurons was 328 μm3 (interindividual CV = 0·15) in the Alzheimer group compared with 277 μm3 (interindividual CV = 0·17) in controls which was a statistically significant increase (P = 0·049). The perikaryal volume was 1117 μm3 in the Alzheimer group compared with 999 μm3 in controls which was a nonsignificant difference (P = 0·20). There was a highly significant correlation between the nuclear and perikaryal volumes in all individuals. The average slope of the regression lines was significantly higher in the Alzheimer patients than in the controls, illustrating that nuclear hypertrophy was more pronounced in the largest neurons.

Cardiac fibrosis is a characteristic feature of left ventricular hypertrophy. The aim of this study was to develop a simple and accurate method to analyse collagen accumulation, taking into account the variation in cardiac muscle fibre orientation and nonuniform collagen distribution. This technique was used to determine the amount and types of collagen that accumulate during pressure overload cardiac hypertrophy. These data were correlated with myocyte size, and with the diastolic stress–strain relationship of the intact myocardium. Myocyte size was significantly increased in the hypertrophied hearts, compared with age and sex matched controls (control 363±25 μm2 vs experimental 244μm2; mean±S.E., P < 0.05). No overall collagen accumulation was observed in the hypertrophied hearts, but a significant increase in collagen I was found with a reduction in the amount of collagen III in experimental animals. Since no increase in diastolic stiffness of the hearts was observed, these results indicate that an increase in the overall collagen content of the heart, rather than the upregulation of a specific type, may be necessary to cause diastolic dysfunction.

In 17 adult and 3 full term fetuses the hip joints were dissected to expose the insertion of gluteus minimus. An attachment of the deep surface of the tendon as it passes over the hip joint capsule was found in every case. Histological examination of this insertion confirmed the presence of short intramuscular tendons firmly anchoring the tendon to the capsule. It is suggested that this attachment retracts the capsule during hip joint motion, thereby preventing capsular entrapment.