The distribution of glycoconjugates was examined in the nonsensory regions of the rat cochlea during postnatal development using biotin-conjugated lectins. Temporal bones of rats at postnatal d 1 and at wk 2, 4 and 6 were fixed in 4% paraformaldehyde and 0.1% glutaraldehyde and processed for paraffin wax embedding. The dewaxed sections were incubated with 7 biotinylated lectins, followed by avidin-biotin- peroxidase complex. A different staining pattern was observed in the stria vascularis, spiral ligament and spiral limbus in the age groups examined. The staining intensity varied between lectins and the reaction product exhibited limited disparity. The staining intensity for WGA increased with age in all the 3 nonsensory regions. The staining patterns for the other lectins differed in the various nonsensory regions examined indicating tissue specificity. The limited variations in the lectin binding patterns after 2nd wk of postnatal life also indicate that the changes in the carbohydrate moieties are established during the fetal period of cochlear development and limited changes take place during postnatal maturation of the nonsensory regions.

Cortical spreading depression (CSD) and peri-infarct depolarisation (PID) are related phenomena that have been associated with the human clinical syndromes of migraine (CSD), head injury and stroke (PID). Nevertheless the existence of CSD in man remains controversial, despite the detection of this phenomenon in the brains of most, if not all, other animal species investigated. This failure to unambiguously detect CSD clinically may be at least partly due to the anatomically complex, gyrencephalic structure of the human brain. This study was designed to establish conditions for the study of CSD in the brain of a gyrencephalic species using the noninvasive technique of magnetic resonance imaging (MRI). The 3-dimensional (3D) gyrencephalic anatomy of the cat brain was examined to determine the imaging conditions necessary to detect CSD events. Orthogonal transverse, sagittal and horizontal T1-weighted image slices showed that the marginal and suprasylvian gyri were the most appropriate cortical structures to study CSD. This was in view of (1) their simple geometry: (2) their lengthy extent of grey matter orientated rostrocaudally in the cortex: (3) their separation by a sulcus across which CSD spread could be studied and (4) the discontinuity in the grey matter in these regions between the right and left hemispheres dorsal to the corpus callosum. The structure suggested by the T1-weighted images was corroborated by systematic diffusion tensor imaging to map the fractional anisotropy and diffusion trace. Thus a single horizontal image plane could visualise the neighbouring suprasylvian and marginal gyri of both cerebral hemispheres, whereas its complex shape and position ruled out the ectosylvian gyrus for CSD studies. With the horizontal imaging plane, CSD events were reproducibly detected by animating successive diffusion-weighted MR images following local KCl stimulation of the cortical surface. In single image frames, CSD detection and characterisation required image subtraction or statistical mapping methods that, nevertheless, yielded concordant results. In repeat experiments, CSD events were qualitatively similar in appearance whether elicited by sustained or transient KCl applications. Our experimental approach thus successfully describes cat brain anatomy in vivo, and elucidates the necessary conditions for the application of MRI methods to detect CSD propagation.

Metatarsal growth was studied in 780 human fetal metatarsal bones, crown-rump length ranging from 120 to 260 mm and gestational age ranging from 14 to 23 wk postconception. Total length, diaphyseal length and diameter were measured and statistical analysis performed. Means and standard deviations for each measurement were calculated at 1 wk intervals. These data were correlated with crown-rump length and gestational age and an equation was constructed in an attempt to predict gestational age.

Mammalian Langerhans cells (LC) are epidermal dendritic cells which originate in bone marrow and migrate toward the T cell area of lymph nodes, where they act as professional antigen-presenting cells. A variety of cell surface markers, such as the ectoenzyme adenosine triphosphatase (ATPase), Ia and CD1a antigens, have been used extensively to identify LC. Ultrastructural identification of this cell type in the mammalian epidermis is made by the demonstration of a typical and unique cytoplasmic organelle, the Birbeck granule (BG). Although we had earlier demonstrated the coexpression of ATPase and Ia antigens on epidermal dendritic cells of the chicken epidermis, the presence of the BG has not previously been documented. The aim of the present study was to investigate whether chicken epidermal LC-like cells possess an organelle similar to the BG, and thus to complete their identification. Our findings are the first demonstration of characteristic rod-shaped, racket-shaped and disc-shaped intracytoplasmic organelles, morphologically similar to the mammalian BG, in avian LC.

The human vomeronasal organ is of interest because of its potential role in sex pheromone detection. Due to the scarcity of early human material, studies of its development have concentrated on fetal rather than embryonic stages. The availability of embryonic specimens in the Walmsley Collection has enabled us to study the development of the vomeronasal organ (VNO) in human embryos between Carnegie Stages 17 and 23. Embryos at Carnegie Stage 17 or below showed no evidence of a VNO. One embryo with characteristics intermediate between Carnegie stages 17 and 18 was the earliest to show evidence of a VNO, in the form of a shallow indentation. All embryos at Carnegie Stages 18 or later had VNOs. Three-dimensional computer reconstructions were made of the VNO in each specimen where this was possible. This in part depended on the plane of section. The total volume and lumen volume were measured from these reconstructions and the volume of the vomeronasal epithelium was calculated by subtraction. A generally consistent increase in total volume and epithelial volume was observed with increasing developmental stage. The lumen contributed rather little to the total volume at these stages.

The effect of systemic NGF treatment on loss and shrinkage of dorsal root ganglion cells was studied in adult male rats after permanent axotomy. Nineteen 16 to 18-wk-old rats had their right 5th lumbar spinal nerve ligated and cut approximately 7 mm peripheral to the ganglion. Two days before the operation, treatment with subcutaneous injections of human recombinant NGF (1.0–0.5 mg/kg/day) was started in 9 test rats; 10 controls were given saline injections. After 1 mo the levels of substance P (SP) and calcitonin gene related peptide (CGRP) were significantly increased in intact sciatic nerve. The number and mean volume of perikarya were estimated using assumption-free stereological techniques including vertical sections, the Cavalieri principle, optical disectors, the planar rotator and systematic sampling techniques. Systemic NGF administration had no influence on survival of primary sensory neurons after axotomy. The number of perikarya was 14300 (S.D.=1800) in axotomised ganglia in control rats versus 14700 (S.D.=2100) in axotomised ganglia of NGF treated rats. The reduction of perikarya volume after axotomy was significantly less after NGF treatment (11600 μm3 in the control group versus 8000 μm3 in the NGF treated group). However, the apparent protection of NGF-treatment on perikaryal volume is explained by a hitherto unrecognised size effect on nonaxotomised dorsal root ganglion cells. The untreated rats had a mean volume of 24700 μm3 (S.D.=2700 μm 3) whereas rats treated with NGF had a volume of 20400 μm3 (S.D.=1700 μm3) on the nonaxotomised side. In conclusion, systemic NGF treatment in adult rats has no effect on dorsal root ganglion cell loss in permanent axotomy whereas perikaryal size of intact nonaxotomised cells is reduced.

In the tammar wallaby (Macropus eugenii), post-testicular acrosomal shaping involves a complex infolding and fusion of the anterior and lateral projections of the scoop-shaped acrosome into a compact button-like structure occupying the depression on the anterior end of the sperm nucleus. The present study has generated cytochemical and histological evidence to demonstrate that the occurrence of actin filaments (F-actin, labelled by Phalloidin-FITC) in the acrosome of tammar wallaby spermatozoa is temporally and spatially associated with the process of acrosomal shaping in the epididymis, through a pool of monomeric actin (G-actin, labelled by Rh-DNase I) present in the acrosome throughout all stages of epididymal maturation. F-actin was not detected in the acrosome of testicular spermatozoa, but was found in the infolding and condensing acrosome of caput and corpus epididymal spermatozoa. When the spermatozoa completed acrosome shaping in the cauda epididymidis, F-actin disappeared from the acrosomal area. The strong correlation between the occurrence of F-actin and the events of acrosomal shaping suggested that the post-testicular shaping of the acrosome might depend on a precise succession of assembly and disassembly of F-actin within the acrosome as the spermatozoa transit the epididymis. Thus, actin filaments might play a significant role in the acrosomal transformation, as they are commonly involved in morphological changes in somatic cells.

Bell-Magendie's law of separation of spinal function states that afferent and efferent fibres join the spinal cord separately in ventral and dorsal spinal nerve roots. For over 100 years there have been reports that challenge the exclusiveness of this law in mammals; very few studies have referred to man. We conducted a prospective morphological study in patients with preganglionic traction injuries of the brachial plexus to address this question. Avulsed ventral and dorsal roots were examined after variable intervals from the injury for histological and ultrastructural evidence for myelinated afferent fibres entering the cord via the ventral roots. Intact myelinated fibres were found in all ventral root specimens, but the majority of fibres in later biopsies are regenerative. A small number of fibres could be demonstrated that are likely to be ‘wrong way ventral afferents’. Their number is falsely low due to wallerian degeneration of dorsal and ventral afferents following the mechanical and ischaemic effects of traction injury. Our findings are the first morphological evidence in human material that Bell-Magendie's law might not entirely be correct and they underline the difficulties in comparing traumatic with experimental rhizotomy.

Vascular sympathetic neuroeffector associations have been examined in rat iris arterioles using serial section electron microscopy and reconstruction techniques. Examination of random sections showed that, of all profiles of varicosities (199) seen to lie closer than 4 μm to vascular smooth muscle cells, only a small proportion (29/199) were found in close association with vascular smooth muscle cells, where adjacent membranes were separated by less than 100 nm. However, serial section examination, from intervaricose region to intervaricose region, of 79 varicosities similarly observed lying within 4 μm of vascular smooth muscle cells showed that 54 formed close associations with vascular smooth muscle cells. In serial sections, all these varicosities were also closely associated with melanocytes and of the 25 remaining varicosities, 22 formed close associations with melanocytes alone, whilst 3 did not come into close association with any effector cell. The increased observation of close associations with vascular smooth muscle cells in serial sections, compared with random sections, is consistent with the demonstration that the area of contact only occupies, on average, a small percentage (5%) of the total surface area of the varicosity as seen in the 3-dimensional reconstructions. In both random and serial sections, close associations were observed between varicosities and vascular smooth muscle cells or melanocytes irrespective of whether fibres were present singly or in small nerve bundles. Three-dimensional reconstruction of associations of varicosities and vascular smooth muscle cells demonstrated several common features, such as accumulations of synaptic vesicles and loss of Schwann cell covering at the region of membrane facing the effector cell. The similarity in the appearance of the neuroeffector association seen in this study and those described in previous studies provides evidence for the existence of a common sympathetic neuroeffector association, irrespective of the receptor subtype involved in neurotransmission.

Wound healing has been studied in the surface ectoderm overlying the midbrain region of stages 16–20 chick embryos by light microscopy, scanning and transmission electron microscopy, and immunofluorescent techniques. The embryos were divided into 6 groups, i.e. stages 16–17 for groups I, V and VI, and stages 19–20 for groups II, III and IV. For groups I and II embryos, a longitudinal incision about 0.6 mm was made close to the dorsal midline and the embryos incubated for varying periods of time up to 24 h. To determine the role of actin in the process of healing, selected groups I and II embryos were stained with FITC phalloidin and the wound margins examined using a confocal microscope. Wounds of all embryos in group I and about 20% in group II healed completely within 24 h of reincubation. The process of healing involved a change in the shapes of the ectodermal cells at the wound ends. This appeared as a zipping-up of the wound from both ends. In about 80% of group II embryos where healing did not occur, wound gaping was marked. Intense actin staining (actin cable) was observed at the wound margins of groups I and II embryos suggesting that the actin purse-string mechanism may play a role during wound healing in this epithelial model. The role of tension in wound healing was also determined by placing 2 secondary wounds about 0.5–0.7 mm long close to, and at right angles to the ends of the primary wound in groups III and V embryos. The procedure decreased the tension within the ectodermal cells at the wound ends. Groups IV and VI embryos served as controls for groups III and V embryos, respectively. Healing of both primary and secondary wounds after reduction of tension was rapid. Most primary wounds in group V embryos healed completely within 3 h of reincubation and the rate of reepithelialisation after the reduction of tension was about 160% more than that in group VI (control) embryos. Similarly, most primary wounds in group III embryos were almost closed within 6 h of reincubation. Here, the rate of reepithelialisation was 80% more than that in group IV (controls). Thus tension is an important factor in wound healing in this model.

Conventional histochemistry and the binding patterns of 22 biotinylated lectins were examined for characterisation of glycoconjugates in the components of the olfactory mucosa of the armadillo Chaetophractus villosus. The mucous lining the olfactory epithelium showed binding sites for DSL, WGA, STL, LEL, PHA-E and JAC. Only the basilar processes of the supporting cells stained for Con-A and S-Con A. The olfactory receptor neurons stained with LEL, LCA, Con A, S-Con A, JAC and PNA. The layer of basal cells did not react with any of the lectins studied. Bowman's glands in the lamina propria showed subpopulations of acinar cells reacting with SBA, S-WGA, WGA, STL, Con A, PSA, PNA, SJA, VVA, JAC and S-Con A, but in our optical studies with lectins we were unable to differentiate between mucous and serous cells in the way that is possible on electron microscopy. The ducts of Bowman's glands were labelled with S-WGA, STL, LEL, PHA-E, BSL-I and JAC. This histochemical study on the glycoconjugates of the olfactory mucosa in the order Xenarthra provides a basis for further experimental investigations.

The aims of the current investigation were (1) to establish an efficient procedure for the isolation of rodent harderian gland cells and to define conditions for maintenance of viable differentiated cells; (2) to compare the in vitro growth pattern of cultured epithelial cells; and (3) to characterise the cultured epithelial cells from 3 rodent species: Wistar rats, Syrian hamsters and Djungarian hamsters. We have established primary culture conditions that permit the maintenance of viable and differentiated secretory cells from adult rodent harderian gland. This study demonstrates that the cell growth pattern is faster in hamsters than in rats and despite morphological changes, epithelial cells reestablish their distinctive (biochemical/metabolic) phenotype as indicated by lipid-containing vacuoles, porphyrin pigment and serotonin and tryptophan hydroxylase labelling.

The distribution of neurons containing neurotensin in the central nervous system of the chicken was studied immunohistochemically. The majority of the neurotensin-immunoreactive (-ir) cell bodies were located in the hypothalamus. Extensive groups of labelled perikarya were found in the hypothalamic periventricular nucleus and in the magnocellular periventricular nucleus. In addition, ir-perikarya were scattered throughout the lateral hypothalamic area and in the ventromedial hypothalamic nucleus. The only extrahypothalamic site of ir-perikarya was in the region immediately under the lateral forebrain bundle. Immunoreactive fibres were detected in the hippocampus, the parahippocampal area, the hypothalamus, the region of the tractus corticohabenular and corticoseptal tracts, the median eminence, the region above the posterior commissure and in the intercollicular nucleus. The distribution pattern of the neurotensin-ir neurons suggests that neurotensin-like peptides are involved in the hypophysiotropic functions.

The British Universities Human Embryo Database has been created by merging information from the Walmsley Collection of Human Embryos at the School of Biological and Medical Sciences, University of St Andrews and from the Boyd Collection of Human Embryos at the Department of Anatomy, University of Cambridge. The database has been made available electronically on the Internet and World Wide Web browsers can be used to implement interactive access to the information stored in the British Universities Human Embryo Database. The database can, therefore, be accessed and searched from remote sites and specific embryos can be identified in terms of their location, age, developmental stage, plane of section, staining technique, and other parameters. It is intended to add information from other similar collections in the UK as it becomes available.

Tooth germs of upper first molars of 1, 3, and 5-d-old rats, fixed in formaldehyde, were stained for the detection of apoptosis by the TUNEL method, and by the azo-dye method for the demonstration of acid phosphatase. For conventional light and electron microscopy the specimens were fixed in glutaraldehyde- formaldehyde and embedded in glycol methacrylate and Araldite. Results showed that macrophages, present in the stellate reticulum, contained basophilic bodies and TUNEL-positive globules, i.e. apoptotic bodies, in their interior. Macrophages also possessed strong acid phosphatase activity. Electron microscopy showed the presence of large vacuoles inside the macrophages containing dense fragmented material. Taken together these results suggest that the intra-epithelial macrophages of the stellate reticulum engulf apoptotic bodies.

The pattern of variation of certain vertebral measurements along the vertebral column is known to differ in man and mouse. This paper investigates changes in this pattern in 7 species of small mammals and attempts to correlate them with locomotor adaptations and limb dimensions.

In tonsils of the soft palate of pigs high endothelial venules (HEV) occur throughout the parafollicular region. In some regions of the venules the endothelial lining is uniformly high; in other areas, mainly the nuclear region of the endothelial cell protrudes into the lumen and the cytoplasmic region is attenuated. In this portion of the venule endothelial cells may traverse the lumen of the vessel to form intravascular bridging processes. The endothelial cells are characterised by pinocytotic vesicles, many surface microvilli, cytoplasmic processes and local cell-cell contacts with adjacent endothelial cells. These specialised HEV may play a role in the unusual pattern of lymphocytes from tonsillar tissue into blood. In the tonsils of the soft palate of pigs, HEV may mediate both lymphocyte migration to and from the lymphoid tissue. Further investigation is required to determine the importance of these 2 migration pathways.

Proper assessment of outcome in clinical trials of neural transplantation requires both biochemical and imaging indices of graft survival, and behavioural and physiological indices of graft function. For transplantation in Huntington's disease, a variety of ligands that are selective for striatal degeneration and graft-derived replacement are available, notably ligands of dopaminergic receptors on striatal neurons. However, the validity of such ligands is potentially compromised by adjunctive drug therapies (e.g. neuroleptics) given to patients in the course of normal clinical care. We review the present state of experimental and clinical understanding of the selectivity of available ligands for striatal imaging, their interaction with other drug treatments, and strategies for refining valid assessment protocols in patients.

The disector is the only objective method for quantifying particles of variable size in a given volume. With this method, cell organelles are identified on adjacent sections, but only those present in one section are counted. When counting extremely rare structures in transmission electron microscope sections (physical disector), the usual procedure of counting on electron micrographs is limited for economic reasons (e.g. micrographs highly outnumbering the investigated structures). Hence, to apply this unbiased stereological method, a modification of the physical disector concerning 3 aspects has been developed. (1) The prerequisite of screening large corresponding tissue areas (here ∼65000 μm2) was fulfilled by examining tissue areas along the edges of ultrathin sections. (2) The size of the counting frame was determined by measuring the lengths of the section margins (minus a guard area) by means of a Morphomat. This value was multiplied by the width of the investigated tissue zone, corresponding to the diameter of the electron microscope viewing screen. (3) Disector counting was carried out simultaneously on both sections (bidirectional disector) to improve efficiency. In the present study tiny synaptic bodies (SBs) were quantitated by disector in a rat pineal gland, yielding ∼30 SBs/1000 μm3. By contrast, single section profile counts of SBs amounted to 90 SBs/20000 μm2. Since the presently described adaptation of the disector is time-consuming, it is proposed to determine a proportion factor allowing to estimate number of structures per volume based on single section profile counts. This would decrease the evaluation time by more than 50%.

The density and distribution of nerve fibres immunoreactive to antisera for PGP 9.5 (general neuronal marker), calcitonin gene related peptide (CGRP) and substance P (SP) (markers for sensory neurons), as well as neuropeptide Y (NPY), vasoactive intestinal peptide (VIP) and tyrosine hydroxylase (TH) (markers for autonomic fibres), were examined in the temporomandibular joint (TMJ) of late gestation fetal sheep. This work formed part of a project investigating the influence of age and osteoarthritis on the innervation of the TMJ, and was undertaken to determine whether the innervation of the joint at 140 d gestation (17 d before birth) differed from that in the mature adult. Immunofluorescence microscopy was applied to serial sections of the capsule, disc and synovial membrane of 10 joints from 5 fetuses and image analysis was used for the quantitative assessment. The capsule, synovial membrane and the disc contained fibres immunoreactive (IR) to antisera for PGP 9.5, SP and CGRP. NPY-IR fibres were only visible in the loose connective tissue of the capsule. No VIP- or TH-IR nerve fibres were detected in the fetal TMJ. There was no statistically detectable difference between the density of nerve fibres immunoreactive to CGRP or PGP 9.5 antisera in the capsule or disc. Substance P-immunoreactivity (IR) was relatively weak in all samples examined. Scattered branches of CGRP-IR fibres were found deep in the disc proper. The lack of receptor endings, other than free nerve endings in the TMJ of the late fetal sheep, might be a reflection of the functional and anatomical immaturity of the TMJ, as reflected in the immature, gross and microscopic appearance of the disc, the inferior joint compartment and articular surface of the condyle at this stage. These results demonstrate that the capsule, synovial membrane and disc in the TMJ of fetal sheep at 140 d gestation age are innervated with sensory fibres, while autonomic fibres are located in the capsule only. The findings also support the view that the disc is innervated at an early stage of life but at a later stage the density of innervation in the central part of the disc regresses and the innervation remains only peripherally in the adult TMJ disc. Further work is required to determine (1) at what stage sympathetic fibres innervate the disc and the synovium, and (2) when the mechanoreceptive nerve endings develop.