The vascular and nervous structures and their relations with the spinal nerve roots were examined in the 2nd, 3rd and 4th posterior sacral foramina in relation to percutaneous needle insertion for neuromodulation. A foraminal branch provided by the lateral sacral artery to each foramen entered the inferior lateral quadrant of each foramen, adjacent to the nerve root medially. Facing the posterior sacral aperture and around the sacral nerves, there was no venous plexus. A venous plexus was sometimes present near the median line, and always around the proximal part of the spinal ganglion. The sacral nerve roots, especially the 3rd, had a long extradural course in the foramen, presenting a potential risk of nerve lesions during procedures involving needle insertion.

Conventional carbohydrate histochemistry and the binding patterns of 21 lectins were analysed to characterise the glycoconjugate content in the components of the vomeronasal organ of the armadillo Chaetophractus villosus. The mucomicrovillous complex of the sensory epithelium bound most of the lectins studied. No reaction was observed with Con A, PSA, S-Con A and SBA, and the sustentacular cells were stained with UEA-I, DSL, LEL, STL and Con A. The vomeronasal receptor neurons were labelled with S-WGA, WGA, PNA, UEA-I, STL, Con A, S-Con A, ECL and RCA120. The basal cell layer reacted with S-WGA, WGA, LCA, UEA-I, DSL, LEL, STL, Con A, JAC and VVA. The nonsensory epithelium exhibited a differential staining in relation to the different components. The mucociliary complex stained with ECL, DBA, JAC, RCA120, STL, LCA, PHA-E, PHA-L, LEL, BSL-I and VVA. However, SJA and UEA-I stained the mucus complex lining a subpopulation of columnar cells. The cytoplasm and cell membranes of columnar cells was labelled with DBA, DSL and LCA. The apical region of these cells exhibited moderate reactivity with LEL and SJA. None of the lectins bound specifically to secretory granules of the nonsecretory cells. Basal cells of the nonsensory epithelium were labelled with DSL, LEL, LCA, BSL- I and STL. The vomeronasal glands showed a positive reaction with WGA, DSL, LEL, LCA, DBA, PNA, RCA120 and SBA. Subpopulations of acinar cells were observed with ECL, S-WGA, Con A, S-Con A and DBA. PNA and RCA120 stained the cells lining the glandular ducts. In comparison with previous results obtained in the olfactory mucosa of the same group of armadillos, the carbohydrate composition of the vomeronasal organ sensory epithelium differed from the olfactory sensory epithelium. This is probably related to the different nature of molecules involved in the perireceptor processes.

In certain Australian marsupials including the tammar wallaby (Macropus eugenii) and the brushtail possum (Trichosurus vulpecula), formation of the acrosome is not completed in the testis but during a complex differentiation process as spermatozoa pass through the epididymis. Using transmission and scanning electron microscopy this paper defined the process of acrosome formation in the epididymis, providing temporal and spatial information on the striking reorganisation of the acrosomal membranes and matrix and of the overlying sperm surface involved. On leaving the testis wallaby and possum spermatozoa had elongated ‘scoop’-shaped acrosomes projecting from the dorsal surface of the head. During passage down the epididymis, this structure condensed into the compact button-like organelle found on ejaculated spermatozoa. This condensation was achieved by a complex process of infolding and fusion of the lateral projections of the ‘scoop’. In the head of the epididymis the rims of the lateral scoop projections became shorter and thickened and folded inwards, to eventually meet midway along the longitudinal axis of the acrosome. As spermatozoa passed through the body of the epididymis the lateral projections fused together. Evidence of this fusion of the immature outer acrosomal membrane is the presence of vesicles within the acrosomal matrix which persist even in ejaculated spermatozoa. When spermatozoa have reached the tail of the epididymis the acrosome condenses into its mature form, as a small button-like structure contained within the depression on the anterior end of the nucleus. During the infolding process, the membranes associated with the immature acrosome are either engulfed into the acrosomal matrix (outer acrosomal membrane), or eliminated from the sperm head as tubular membrane elements (cytoplasmic membrane). Thus the surface and organelles of the testicular sperm head are transient structures in those marsupials with posttesticular acrosome formation and this must be taken into consideration in attempts to dissect the cell and molecular biology of fertilisation.

Information abut the existence and anatomy of arterial anastomoses with the porcine bronchial artery is lacking in the literature. Prior to basic physiological investigations in a porcine model related to lung transplantation with bronchial artery revascularisation, this study was designed to examine the anatomy of systemic arterial anastomoses with the bronchial artery system. Twenty pigs were studied in 3 groups. In 2 groups the heart–lung block was removed with all mediastinal structures. One group served for investigation of coronary–bronchial artery anastomoses and one for investigation of oesophageal–bronchial artery anastomoses. The systemic arteries to be examined were cannulated. The inflated heart–lung block was examined macroscopically with Evans blue, and radiographically after contrast injection. In the 3rd group intercostobronchial artery anastomoses were studied radiographically with the heart–lung block in situ. Coronary–bronchial artery anastomoses were demonstrated in 3 of the 5 pigs with an aortic ‘pouch’ technique, but contrast was very limited in 2 of these 3. Oesophageal arterial anastomoses with bronchial arterial branches and/or the pulmonary veins were demonstrated in 6 of the 7 pigs and more markedly than the coronary–bronchial anastomoses. Intercostobronchial artery anastomoses could not be demonstrated angiographically. It was concluded that the existence of coronary–bronchial and oesophageal–bronchial artery anastomoses in the pig appear to establish an arterial net between the base of the heart and the distal oesophagus. The resemblance to human oesophageal–bronchial artery anastomoses supports use of a porcine model for experimental studies.

Studies of tetraploid[harr ]diploid (4n[harr ]2n) mouse chimaeras have demonstrated unequal contributions of 4n cells to different tissues of the midgestation conceptus. Such a pattern has also been reported in chimaeras as early as E3.5d, which show an enhanced contribution of 4n cells to the mural trophectoderm (Everett & West, 1996). In this study, sectioned 4n[harr ]2n and 2n[harr ]2n control chimaeric blastocysts were digitised and reconstructed in 3 dimensions (3-D). The 3-D images revealed only limited mixing of cells from the 2 contributing embryos of individual blastocysts in both chimaera groups. Consequently, the distribution pattern of the 2 cell types was dependent on the spatial relationship between the orientation of the blastocyst and the boundary between the 2 clusters of cells. The distribution patterns observed were not strikingly different for 4n[harr ]2n and 2n[harr ]2n chimaeras, each showing some transgenic positive cell contribution in all 3 identifiable developmental lineages. It was notable, however, that in all 4n[harr ]2n blastocysts at least some 4n cells were located adjacent to the blastocyst cavity. Such a consistent pattern was not evident in 2n[harr ]2n chimaeras. This study has demonstrated the value of 3-D reconstructions for the analysis of spatial relationships of 2 cell populations in chimaeric mouse blastocysts.

Adult muscle is highly vascularised, with blood vessels being essential for adequate oxygenation of the tissue and for supporting increased metabolic demands. Whether this is the case during muscle development has not been examined. Resin histology was used to map the muscle splitting process and conventional transmission electron microscopy to examine early muscle differentiation at the midlimb level or later at the mid radius/ulna level in the chick wing bud from stages 24 (4.5 d) to 36 (10 d) (Hamburger & Hamilton, 1951). Microinjection of India Ink into the extra-embryonic vasculature was used to visualise the patent muscle microcirculation. The results showed that the premuscle masses are present at stage 24 and initial splitting of the muscle masses commences at stage 28. The final muscle pattern is not established until stage 36. At stage 26 the cells within the premuscle masses exhibited a mesenchymal morphology, but at stage 28 overt muscle differentiation was evident with myofibrils present within myoblasts. Undifferentiated mononucleated cells were interspersed with the differentiating myoblasts. The ratio of mononucleated cells[ratio ]myoblasts decreased and the myoblasts became plumper and increasingly packed with myofibrils with age. There was no evidence of secondary myotube formation at any of the stages examined. Vascular invasion of the limb occurred at stage 35 just prior to the establishment of the final muscle pattern. This was surprising as it was assumed that myogenic differentiation would be both oxygen and nutrient dependent. The results of this study provide descriptions of the splitting of the premuscle masses through to the establishment of the final muscle pattern at the midlimb or mid radius/ulna level of the chick wing bud together with the differentiation of the myogenic cells within the developing muscles. However, the relationship between muscle patterning at the tissue level and muscle differentiation at the cellular level with vascularisation remains unclear. It is hoped that the results of the study may provide the basis for future investigations into mechanisms involved in muscle patterning and the signalling mechanisms for vascular invasion.

The fibrillar collagens associated with the articular cartilages, joint capsule and ligamentum teres of the rabbit hip joint were characterised from the 17 d fetus to the 2-y-old adult by immunohistochemical methods. Initially the putative articular cartilage contains types I, III and V collagens, but when cavitation is complete in the 25 d fetus, type II collagen appears. In the 17 d fetus, the cells of the chondrogenous layers express type I collagen mRNA, but not that of type II collagen. Types III and V collagens are present throughout life, particularly pericellularly. Type I collagen is lost. In all respects, the articular cartilage of the hip joint is similar to that of the knee. The joint capsule contains types I, III and V collagens. In the fetus the ligamentum teres contains types I and V collagens and the cells express type I collagen mRNA; type III collagen is confined mainly to its surface and insertions. After birth, the same distribution remains, but there is more type III collagen in the ligament, proper. The attachment to the cartilage of the head of the femur is marked only by fibres of type I collagen traversing the cartilage; the attachment cannot be distinguished in preparations localising types III and V collagens. The attachment to the bone at the lip of the acetabulum is via fibres of types I and V collagens and little type III is present. The ligament is covered by a sheath of types III and V collagens. Type II collagen was not located in any part of the ligamentum teres. The distribution of collagens in the ligamentum teres is similar to that in the collateral ligaments of the knee. Its insertions are unusual because no fibrocartilage was detected.

This study determines deformations of the midface that contribute to a class III appearance, employing thin-plate spline analysis. A total of 135 lateral cephalographs of prepubertal children of European-American descent with either class III malocclusions or a class I molar occlusion were compared. The cephalographs were traced and checked, and 7 homologous landmarks of the midface were identified and digitised. The data sets were scaled to an equivalent size and subjected to Procrustes analysis. These statistical tests indicated significant differences (P<0.05) between the averaged class I and class III morphologies. Thin-plate spline analysis indicated that both affine and nonaffine transformations contribute towards the total spline for the averaged midfacial configuration. For nonaffine transformations, partial warp 3 had the highest magnitude, indicating the large scale deformations of the midfacial configuration. These deformations affected the palatal landmarks, and were associated with compression of the midfacial complex in the anteroposterior plane predominantly. Partial warp 4 produced some vertical compression of the posterior aspect of the midfacial complex whereas partial warps 1 and 2 indicated localised shape changes of the maxillary alveolus region. Large spatial-scale deformations therefore affect the midfacial complex in an anteroposterior axis, in combination with vertical compression and localised distortions. These deformations may represent a developmental diminution of the palatal complex anteroposteriorly that, allied with vertical shortening of midfacial height posteriorly, results in class III malocclusions with a retrusive midfacial profile.

Structural changes of the ventral velum of Xenopus laevis tadpoles from late prometamorphosis (stage 58) to the height of metamorphic climax (stage 62) were examined by light and transmission electron microscopy. Special emphasis was given to the blood vessel regression. Early changes of velar capillaries were formation of luminal and abluminal endothelial cell processes, vacuolation, and cytoplasmic and nuclear chromatin condensation. At the height of metamorphic climax, transmission electron microscopy revealed apoptotic endothelial cells with nuclear condensation and fragmentation, intraluminal bulging of rounded endothelial cells which narrowed or even plugged the capillary, and different stages of endothelial cell detachment (‘shedding’) into the vessel lumen. These changes explain the ‘miniaturisation’ of the velar microvascular bed as well as the typical features found in resin-casts of regressing velar vessels which have been observed in a previous scanning electron microscopy study of the ventral velum.

We report a change in the vascularisation of the adipose depots surrounding the popliteal lymph node that has, and the contralateral node that has not, been exposed to a simulated immune challenge. The percentage of the depot that consists of vessels, as measured by image analysis, decreases over a period of 2 d after immune stimulus, then increases in a biphasic manner over the next 2–3 wk. By 1 mo after the stimulus, the vascularisation has returned to baseline values. The adipose tissue surrounding both the stimulated and the unstimulated lymph nodes shows a similar pattern, but the unstimulated depot lags by 3–6 d in reaching its maximum vascularisation. These data support the hypothesis that perinodal adipose tissue is involved in peripheral immune responses.

During development axons contact their target tissues with phenomenal accuracy but the mechanisms that control this homing behaviour remain largely elusive. A prerequisite to the study of the factors involved in hard-wiring the nervous system during neurogenesis is an accurate calendar of developmental events. We have studied the maxillary and mandibular components of the trigeminal system to determine the stages during embryogenesis when a gross somatotopic order is first established within the trigeminal ganglion and the axons projecting to the brainstem. The retrograde transganglionic fluorescent tracers DiO and DiI were injected into the maxillary and mandibular arches or their derivatives in fixed mouse embryos staged between 13 and 40 somites (E9–E11). After 1–4 wk, the distribution of the 2 tracers was determined using confocal laser scanning microscopy. The first maxillary nerve cell bodies and their developing axons were labelled at the 30 somite stage (E10). This was 2 somite stages earlier than the mesencephalic nucleus and the ganglion cell bodies of the mandibular nerve. The gross somatotopic division of cells within the trigeminal ganglion projecting to the maxillary and mandibular targets was established by the 32 somite stage (E10). This arrangement was evident as 2 groups of cell bodies occupying adjacent but separate regions of the trigeminal ganglion. The central branches of the maxillary and mandibular cell bodies entered the metencephalon as 2 distinct bundles at the same stage. The trigeminal motor nucleus was first detected at the 38 somite stage (E10.5).

Gross somatotopy in the major divisions of the trigeminal ganglion is established before outgrowing axons have contacted their peripheral target tissue at E10.5. This suggests that target tissues do not induce somatotopy.

The cellular ensheathment of capillaries in the 3 outer capillary layers of the central retina of the adult tree shrew Tupaia belangeri was studied quantitatively by transmission electron microscopy. Using a stereological approach, the relative surface of capillary basal lamina ensheathed by Müller cells and by nonmacroglial cells (collectively termed non-Müller cells) was estimated in 5 animals. The participation of Müller cells was distinctly different in the 3 capillary layers studied. In the outermost capillary layer 1, the mean (standard deviation) percentage surface coverage by non-Müller cell processes was 46.8 (15.3)%. Much less of the capillary basal lamina was ensheathed by non-Müller cells in capillary layers 2 and 3 (3.0 (2.1)% and 0.3 (0.3)% respectively). The observed total variation of the stereological estimates for the surface fraction of Müller cells (expressed as the between-subject coefficient of variation) was significantly higher in capillary layer 1 (28.8%) compared with capillary layers 2 (2.2%) and 3 (0.3%). In capillary layer 1, the high observed total variation was due to a high biological variation among animals for the fractions of both Müller cell and non-Müller cell ensheathment. The rare occurrence of direct contacts between the capillary basal lamina and the perikarya of either microglial cells (capillary layer 3) or amacrine cells (capillary layer 2) corresponded well to the low stereological values obtained for the relative capillary surface ensheathed by non-Müller cells in these capillary layers. Previously, extensive and frequent contacts between the basal lamina of capillaries belonging to capillary layer 1 and horizontal cells had been observed in single sections. The present study quantitatively demonstrates a marked paucity of macroglial investment of capillaries located in capillary layer 1 of Tupaia. It can be concluded that horizontal cells ensheath most of the capillary surface not invested by Müller cells.

Variations in the hamstring muscles are not common. We describe here a rare anomalous muscle in the popliteal fossa and speculate on its functional significance.

Apoptosis is a form of naturally occurring cell death during development and it is characterised by extensive DNA fragmentation. Apoptosis is easily detected in the gill epithelium of brown trout embryos in ultrathin sections (Rojo et al. 1997). Here we provide the first biochemical evidence for apoptosis in the gill epithelium of brown trout embryos, using in situ end-labelling of DNA breaks (Gavrieli et al. 1992). Embryos at d 57 of development as well as those at hatching, were processed to analyse the distribution of apoptotic cells in the gills. The extent of apoptosis revealed by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labelling method technique is considerably greater than apoptosis detected by nuclear morphology. This method revealed that apoptosis was frequent at hatching, although it was also present during embryonic development. The presence and distribution of stained nuclei were different depending on the developmental stage. In embryos of 57 d, apoptotic flattened nuclei were dispersed in the gill epithelium, whereas at hatching, they were mainly grouped in the tips of the filaments and in the gill arches. TUNEL also revealed a distinct pattern of nuclear staining: at hatching, the intense staining covered the entire cell, but in embryos it was restricted to the nucleus. These results show the functional relevance of apoptosis at hatching, when apoptosis seems to be the unique process by which cell numbers in the gill epithelium are adjusted, in order to prepare for the new extrinsic conditions affecting the free-living life of alevins.

Neurotisation involves transfer of nerves for the restoration of function following injury. A number of nerves have been used in different part of the peripheral nervous system. This study was undertaken to develop a practical and relatively safe surgical approach to the treatment of L4 root lesion's. We examined the effectiveness and safety of neurotisation of the deep peroneal nerve and its branches by the superficial peroneal nerve. Twelve legs of dissected cadavers provided for teaching purposes in the anatomy laboratory were used to display the common peroneal nerve and its branches. Each branch was measured using calipers and analysed to investigate the possibility of neurotisation of the deep peroneal nerve by the superficial peroneal nerve and its branches. It was found that of the measured branches, transposition was possible between those to peroneus longus and tibialis anterior on the basis of their diameter and length. In recent decades, advances in microsurgical reconstruction and understanding of the microanatomy have played major roles in improving the results of surgical treatment of nerve injuries. There is a need for further experimental studies on the feasibility of this surgical approach.

Cultures of rat thymic epithelium were used to measure the effect of thymulin secretagogues on dye-coupling and proliferation. Dye-coupling was assessed after the injection of lucifer yellow dextran which cannot permeate the connexin pore of gap junctions and the smaller, permeant cascade blue. In addition to gap junctional communication, larger intercellular bridges were demonstrated by the transfer of lucifer yellow dextran between cells. The extent of intercellular communication was found to be influenced by both cell density and the number of passages. In control cultures, intercellular communication was reduced in cell groups of low (<20 cells/group) or high cell densities (>100 cells/group) compared with groups of 20–60 cells. The highest coupling indices were found in subcultures 20–30. Taking these factors into account, significant decreases in coupling index were observed after pretreatment of test cultures with factors known to influence the secretion of thymulin (5 U/ml interleukin 1 (α and β), 1 μM progesterone, 1 μM oestrogen, 1 μM testosterone, 1 ng/ml adrenocorticotropic hormone, 100 nm rat growth hormone) but 7.5 ng/ml thymulin had no effect on dye-coupling. The nonspecific gap junction uncoupler, octanol, abolished dye-coupling. Cellular proliferation, as measured by the uptake of tritiated thymidine, showed that the same factors that reduced coupling also increased proliferation. None of these factors affected the number of multinucleate cells present, except interleukin-1β which caused a significant reduction in the average number of nuclei per cell. Thus rat thymic epithelium in vitro provides a model for the study of the direct action of factors on cells of the thymic microenvironment.

Immunoreactivity to insulin (Ins), somatostatin (Som), glucagon (Glu) and pancreatic polypeptide (PP) was found in 70%, 22%, 15% and 11% respectively of Houbara pancreatic endocrine islet cells. Whilst Ins occurred centrally and SOM was observed both in peripherally and centrally located islets, the other hormones were localised in peripheral islet cells; Som was also observed in neuronal cell bodies and nerve fibres. In addition, the islet cells contained substance P (SP) (65%) in the centre and vasoactive intestinal polypeptide (VIP) (2%) at the periphery. Immunoreactivity to choline acetyltransferase (ChAT), VIP and galanin (Gal) occurred in the walls of blood vessels located mainly at the periphery of islets. Occasionally, VIP and Gal immunoreactive varicose nerve terminals and ChAT immunoreactive cell bodies were also observed in the centre of islets. SP neuronal cell bodies were not observed but prominent SP immunoreactive varicose terminals were discernible in capillary walls within the islets. Neuropeptide Y (NPY) immunoreactive neurons were detected in neuronal cell bodies located mainly peripherally. Neuronal nitric oxide synthase (nNOS) immunoreactivity occurred in neuronal cell bodies and nerve fibres mainly at the periphery and also in centrally located islet endocrine cells. Immunoreactivity to tyrosine hydroxylase (TH) was similar in distribution to that of ChAT. In comparison with other avian species, the islets of the dorsal pancreatic lobe of the bustard contain all the peptidergic hormones normally present in the islets of other avian species, but are not segregated into dark A and light B cells. Many of the insulin containing cells also contained SP. The islets also contained several neuropeptides which are probably involved in their regulation.

In the infant the region lateral to the sella turcica which is traditionally termed the ‘cavernous sinus’ is composed of 3 individual compartments. The aim of this study was to demonstrate a similar compartmentation of the adult cavernous sinus and to identify and quantify the adipose bodies which are located within the compartments. The region of the cavernous sinus in 136 adults was analysed by microdissection and histology. We demonstrate that in 66% of the cavernous sinus in adults is composed of at least 2 compartments and in 22% it is made up of 3. Assimilating the renaming in infants we termed the compartments ‘pterygopalatine’, ‘orbital’ and ‘lateral sellar’. The pterygopalatine and orbital compartments are connected with extracranial tissue spaces via the superior orbital fissure and contain characteristic adipose bodies. Exact topographic descriptions and measurements of the compartments and their adipose bodies are provided. Our study clearly defines compartmentation of the adult parasellar space in most individuals and thus changes the anatomical view of this space. The direct connection of 2 of these compartments with extracranial tissue spaces and the measurements of their adipose bodies are of interest for surgeons and neuroradiologists.

A quick-freezing and deep-etching method in combination with erythrocyte splitting was used to examine the cytoplasmic aspect of whole-mount human erythrocyte membranes. Various external forces induced alterations in membrane skeletal organisation during the splitting procedure. The initial change was elongation in the peripheral part of the membrane skeleton, examined by immunostaining with a monoclonal antispectrin antibody. Under severe stretching conditions, a linear rearrangement of filamentous components was evident; these were disposed parallel to the rim of the erythrocyte, while the central part of the concavity exhibited a more compacted structure. These changes resulted in a different distribution of membrane skeletal components between central rigid and peripheral flexible areas in biconcave erythrocytes. It is suggested that the reversible membrane skeletal changes in the flexible areas which resist the external forces are important for maintaining the normal framework of biconcave human erythrocytes.

Electron immunocytochemistry was used to examine perivascular nerves of hamster mesenteric and renal arteries during hibernation and 2 h after arousal from hibernation. Vessels from cold-exposed but nonhibernating, and normothermic control hamsters were also examined. During hibernation the percentage of axon profiles in mesenteric and renal arteries that were immunopositive for markers of sympathetic nerves, tyrosine hydroxylase (TH) and neuropeptide Y (NPY), were increased 2–3 fold compared with normothermic and cold control animals. This increase was reduced markedly only 2 h after arousal from hibernation. The small percentage of nitric oxide synthase-1-positive axon profiles found in mesenteric (but not renal) arteries was also increased during hibernation and returned towards control values after arousal. In contrast, the percentage of perivascular axons immunostaining for vasoactive intestinal polypeptide (VIP), a marker for parasympathetic nerves, was reduced in mesenteric arteries during hibernation. There was no labelling of perivascular nerves for substance P in either mesenteric or renal arteries. It is suggested that the increase in percentage of TH- and NPY-immunostained perivascular nerves may account for the increased vasoconstriction associated with high vascular resistance that is known to occur during hibernation. The reduction in the percentage of axons positive for VIP in hibernating animals would contribute to this mechanism since this neuropeptide is a vasodilator.