Ultrastructural observations on myoepithelial cells and nerve terminals in the camel Harderian gland

Summary The myoepithelial cells and the nerve terminals of the Harderian gland in the one-humped camel were examined by using the transmission electron microscope. The myoepithelial cells are well developed, and composed of a cell body containing the nucleus, and many cytoplasmic processes. The cytological features are consistent with the function of the myoepithelial cell mainly as a contractile element, and possibly as a regulator for fluid transport. They are attached to the glandular cells of the end-pieces with desmosomes and interdigitating cytoplasmic processes. Densely packed myofilaments fill most of the cytoplasm, and micropinocytosis vesicles on the inner and outer borders are prevalent. The glandular end-pieces are innervated with unmyelinated nerve terminals which have been observed in the interstitial connective tissue. Nerve terminals without neurolemmal sheath penetrating the basal lamina and forming a direct neuroglandular contact with the glandular cells were observed. These intraglandular nerve terminals were found in direct contact with the myoepithelial cells, and contained small clear vesicles and a few larger dense granules.     

Production of interleukin-1 in a South American opossum (Monodelphis domestica)

A homologous thymocyte costimulatory assay using thymocytes from a South American opossum (Monodelphis domestica) detected and measured interleukin-1 (IL-1). Opossum IL-1 was obtained from lipopolysac-charide-stimulated macrophage and skin cultures and its molecular weight was determined to be 15,000 to 17,000. Opossum IL-1 did not stimulate proliferation of murine thymocytes; conversely, neither human nor murine IL-1 stimulated opossum thymocytes. Anti-human IL-1 antibodies were also not reactive with opossum IL-1. These observations indicate that there is no serological and functional crossreactivity between the opossum (marsupial mammals) and human and mouse (eutherian mammals) in an IL-1/thymocyte system. This is unusual because such crossreactivity occurs between rodents and distant, nonmammalian species. M. domestica has been used in our laboratory as a model for photobiological research; studies on IL-1 may provide insight into the relationship of immunosuppression and tumorigenesis induced by ultraviolet radiation.     

An ultrastructural study of myoepithelium maturation during postnatal development of the hamster Harderian gland

Summary This study reports the ultrastructural cell modifications in the myoepithelium of the Harderian gland during the postnatal development of the Syrian hamster. Tissues were obtained from male and female hamsters at days 1, 3, 5, 7, 10, 12, 15, 17, 20, 27, 37, 46 and 90 after birth, and processed for transmission electron microscopy. Electron microscopy was coupled with point counting methods to quantitate changes in several subcellular organelles during the course of myoepithelial cell maturation. The myoepithelial cells in this gland remained immature at birth. The earliest age of development when organized bundles of microfilaments were observed was 7 days. By the 12th day, the myoepithelial cells had developed most of their specific characteristics and resembled the mature form. Myoepithelial cells mature synchronously with each other and with the secretory cells. No undifferentiated myoepithelial cells were found in adult glands. In addition to epithelial and myoepithelial cells, the secretory endpieces of the adult hamster Harderian gland contain a third cell type which resembles the myoepithelial cell in shape and has an extremely electron-lucent cytoplasm lacking microfilament bundles.     

G-protein activation enhances Ca+2-dependent lipid secretion of the rat Harderian gland

We studied the secretory mechanism of the Harderian gland of rats. After perfusion with HEPES-buffered Ringer's solution containing NaF (10 mM) with AlCl₃ (10 M), a G-protein activator, the glandular cells of the Harderian gland showed massive exocytosis and apocrine-like protrusions on the luminal surface. Some of the secretory vacuoles aggregated within the cytoplasm, and large vacuoles were formed. Contraction of the myoepithelial cells covering the glandular endpieces caused a narrowing of the glandular lumina, which contained cytoplasmic fragments, and deformation of the basal contour of the glandular end-pieces. The basal regions of the glandular cells also bulged between the myoepithelial cells. Secretory vacuoles were also discharged to the lateral cell surface, and the intercellular spaces were dilated. The enhanced secretory activities of the glandular cells and the contraction of the myoepithelial cells were similar to those in rats stimulated with 10 M carbachol (CCh). However, dilatation of the endoplasmic reticulum in glandular cells (type A cells), which leads to the formation of small vesicles, was observed in those glands stimulated by NaF+AlCl₃, but not in those stimulated by CCh. Removal of Ca⁺² from the perfusing HR or addition of EDTA (0.5 mM) diminished and inhibited NaF+AlCl₃- or CCh-enhanced secretory activity of the glandular cells and also allayed the deformation of glandular cells caused by myoepithelial cell contraction. The present results demonstrate the involvement of G-proteins and Ca²⁺-influx in the lipid secretion of glandular cells and in the contraction of myoepithelial cells of the Harderian gland in rats.     

Effects of chronic administration of cytidine-5′-diphosphate choline (CDP choline) on the aged female mouse Harderian gland

 The effect of chronic intake of cytidine-5′-diphosphate choline (CDP choline), a precursor of the membrane lipid phosphatidylcholine, was analyzed in secretory cells of the Harderian gland (HG) of aged (26-month-old) female Swiss mice. These mice were treated with the drug from 12 months of age until they were 26 months old. The results obtained were compared with those of an age-matched control group and another adult (12-month-old) control group. In both control groups, the HGs displayed secretory tubules formed by a single squamous epithelium of lipid-secreting cells. The tubule lumens were occupied by numerous small pleomorphic porphyrin accretions. In contrast, the tubules of the CDP-choline-treated animals consisted of a single columnar epithelium, with lipid-secreting cells filled with lipid vacuoles. The most striking feature in the HGs of the CDP-choline group was the absence of prophyrin deposits. These observations indicate that porphyrin biosynthesis in female Swiss mice is maintained even in very old animals, and that chronic administration of CDP choline prevents porphyrin deposition in the HG of aged animals. Accepted: 1 August 1996     

The development of monoaminergic brainstem-spinal systems in the North American opossum

Summary Evidence is presented for an early appearance of monoaminergic neurites within the spinal cord of the developing opossum. They are present within the marginal zone before hindlimb movements begin (stage I) and they start to grow into the intermediate zone by the time hindlimb movements are first observed (stage II). Monoaminergic neurites grow first into the dorsolateral intermediate zone and the intermediolateral cell column where they can be found by the beginning of stage II. Shortly thereafter, fluorescent varicosities can be traced into the area dorsal to the central canal presumed to become lamina X. Fluorescent processes extend in to the ventral intermediate zone (ventral horn) somewhat later in development. Monoaminergic axons have grown into all of the areas they occupy in the adult animal, except for laminae I and II, by the time immature hindlimb movements can be altered by cutting all brainstem projections to the lumbosacral cord (stage III). Monoaminergic innervation of laminae I and II is the last to develope, but it is present by the time thoracic transection produces complete spinal shock.This investigation was supported by U.S.P.H.S. Grant NS-07410     

Development and hormonal regulation of mast cells in the Harderian gland of Syrian hamsters

Summary The morphological features and relative number of mast cells per mm² were studied in the Harderian glands of male and female Syrian hamsters (Mesocricetus auratus) under different experimental conditions. The structural and ultrastructural characteristics of Harderian mast cells corresponded to those of connective tissue mast cells. The Harderian glands from female hamsters contained more mast cells than those of male hamsters. A subcutaneous implant of testosterone (2 mg/24 mg beeswax) resulted in a rapid decrease in the number of recognizable mast cells 6 h after the implantation. Neither orchidectomy nor ovariectomy significantly altered the relative number of mast cells. However, the daily subcutaneous injection of 20 IU of human chorionic gonadotropin during 20 days resulted in a significant decrease of identifiable mast cells. The administration of another steroid such as progesterone or the induction of states of hypo- and hyperthyroidism did not alter the distribution of mast cells in the Harderian glands of female Syrian hamsters.     

Magnetic resonance imaging of the Harderian gland in piglets

The main purpose of the present study was to investigate the value and effectiveness of functional and morphological magnetic resonance imaging (MRI), in order to assess the extent of brain injury in a hypoxic-ischaemic piglet model, and further to validate that the ischaemic injury was successfully induced. In this way, we also characterized the Harderian gland. MRI was performed at 1.5 T in anaesthetized piglets (n = 10, 12-36 h of age). Magnetic resonance perfusion and diffusion imaging were performed at different time points, before, during and after the induction of hypoxia-ischaemia. The effects following bilateral clamping of the carotid arteries were also assessed by contrast-enhanced magnetic resonance angiography. Morphological assessment included T1- and T2-weighted imaging, and fat-suppressed T1-weighted imaging before and after contrast medium enhancement. Morphological MRI revealed a prominent, well-defined structure located at the eyeball. Magnetic resonance angiography reconstructed with volume rendering showed this structure to be partially enclosed by large venous sinuses. At dissection, when compared with the magnetic resonance images, the deep gland of the third eyelid, the Harderian gland, corresponded to this structure both in topography and in size. By contrast, the lacrimal gland proper presented as a small, soft and pale structure that was difficult to distinguish from the surrounding connective tissue. At histological examination, the Harderian gland consisted mainly of compact areas of tubuloacinar glands with abundant eosinophilic granules. The present MRI demonstration of the Harderian gland was an accidental finding during an investigation to assess the extent of brain injury in a hypoxic-ischaemic piglet model. The combination of MRI and histology made it possible to detect and describe the Harderian gland in pig. It has generally been studied in rodents and lower vertebrates and is reported to possess various endocrine and exocrine functions.     

Cerebello-olivary fibers: Their origin,course and distribution in the North American opossum

Summary Although degeneration techniques suggest that cerebello-olivary fibers are limited in their origin and distribution, horseradish peroxidase and autoradiographic experiments make it clear that they arise within all cerebellar nuclei and project to most, if not all, areas of the contralateral inferior olive. Autoradiographic preparations show that cerebello-olivary fibers are highly ordered and suggest that the dentate nucleus projects primarily to the principal olive, the interpositus anterior relays particularly heavy to the dorsal accessory nucleus and the interpositus posterior distributes extensively to the medial accessory complex. Evidence for a small projection from the fastigial nucleus to the caudal medial accessory nucleus is also available. However, it appears clear that neither the dentate nor the interpositus nuclei project to just one subdivision of the olive. For example, although dentate fibers end extensively within the principal nucleus some of them also distribute to portions of the medial accessory nucleus and perhaps the dorsal accessory nucleus as well. The medial accessory olive is particularly complex and at rostral levels receives input from both interposed and dentate nuclei, whereas more caudally it receives a projection from the fastigial nucleus. Olivary fibers from both the interposed and dentate nuclei traverse the brachium conjunctivum descendons and distribute primarily to the rostral 2/3 to 3/4 of the olive, whereas those from fastigial neurons take a different route and end more caudally. Experiments utilizing horseradish peroxidase as a retrograde tracer suggest that cerebello-olivary fibers from both the interpositus anterior and dentate nuclei take origin from a population of generally small neurons.This investigation was supported by United States Public Health Service grants NS-07410 to Dr. Martin and NS-08798 to Dr. King.These results were presented in part at the 88th Meeting of the American Association of Anatomists, Los Angeles, California.     

Effects of ovariectomy and ageing on the structure and ultrastructure of the female Syrian hamster Harderian gland: a stereological analysis

The effects of ovariectomy and ageing on the structure and ultrastructure of the Syrian hamster Harderian gland were investigated by techniques of quantitative stereology. Tissues were obtained from intact 6-month-old, sham-operated 6-month-old, ovariectomized 6-month-old, intact 18-month-old and ovariectomized 18-month-old female hamsters. Glands from both ovariectomized and aged hamsters showed comparable qualitative and quantitative characteristics. They showed histological alterations that included thinning of the tubule walls, lowering of luminal porphyrins, invasion of lumina by neutrophils and the occurrence of interstitial porphyrins. Glands from both ovariectomized and aged hamsters showed statistically significant differences from control animals in relation to numerical density and cellular size. Finally, quantitative studies with the electron microscope revealed significant decreases in the volume densities of the cytoplasmic organelles concerned with secretion. These results support the hypotheses that the secretory activity of the female hamster Harderian gland is influenced, directly or indirectly, by ovarian hormones, and that many of the age-related modifications of the Harderian gland reflect alterations in ovarian function.     

A physiological role for protoporphyrin IX photodynamic action in the rat Harderian gland?

Aims: The lipid‐secreting exocrine Harderian gland contains a large amount of porphyrins (mainly protoporphyrin IX, PPIX) in the glandular cells, the physiological significance of which is rather poorly understood. Methods: In the present study, the possibility of using Fura‐2 to measure intracellular calcium ([Ca²⁺]_c) changes in these cells was assessed. Results: It was found that when Fura‐2‐loaded cells were excited by light at 340/380 nm, [Ca²⁺]_c increased spontaneously, indicating a photodynamic action powered by light at 340/380 nm. In contrast, with the visible spectrum calcium probe Fluo‐3 (λ_ex = 475 nm), carbachol at 10 μm induced [Ca²⁺]_c increase; [Ca²⁺]_c did not change without carbachol stimulation. Brief illumination with light at 340/380 nm induced a large [Ca²⁺]_c increase in Fluo‐3‐loaded cells. Photodynamic stimulation of [Ca²⁺]_c increase was confirmed with an exogenous photosensitizer sulphonated aluminium phthalocyanine (SALPC) and visible light (>580 nm). The wavelength‐dependence of the [Ca²⁺]_c increase correlates well with the excitation spectrum of the isolated Harderian glandular cells. Conclusion: These data suggest that PPIX present in rat Harderian glandular cells plays the role of a photosensitizer which upon activation by UVA and blue components of daylight and subsequent singlet oxygen generation, triggers [Ca²⁺]_c increase and secretory response. The PPIX photodynamic action may also play a potential role in photic entrainment of the central circadian clock.     

The development of commissural connections of somatic motor-sensory areas of neocortex in the North American opossum

Summary The North American opossum does not have a corpus callosum; neocortical commissural axons are contained entirely within the anterior commissure. We have used axonal transport techniques to study the origin and distribution of commissural axons from somatic motor-sensory cortex in developing and adult opossums. Neocortical axons grow into the anterior commissure by postnatal day (PND) 12, the contralateral external capsule by approximately PND 19, the area deep to the contralateral homotypic cortex by approximately PND 26 and the cortex proper by approximately PND 35. Commissural neurons were first demonstrated at about PND 26, when they form a fairly continuous band in the cortical subplate (presumptive layers V–VI). By at least PND 37, commissural neurons are also present in layers II and III, where they form a continuous band, and in layer IV, where they are sparse. In older pouch young and adult opossums the bands of commissural neurons, especially in layers V–VI, are interrupted, and commissural neurons are rare in layer IV. In general, commissural axons in both pouch-young and adult opossums innervate areas containing commissural neurons as well as layer I.In the acallosal opossum as well as in the callosal rat, the development of commissural connections from somatic motor-sensory cortex is characterized by pauses during the growth of axons into the opposite cortex, by a general inside-out-gradient, and by a transition from continuous bands to patchy, radial columns of commissural neurons and axons. This suggests that similar mechanisms govern the formation of commissural connections in the two species.This investigation was supported by U.S.P.H.S. Grants BNS-8309245 and NS-10165-10     

Ultrastructural observations on the shell membrane of the North American opossum (Didelphis virginiana)

Each developing opossum embryo is surrounded by a shell membrane which completely separates embryonic and maternal tissues. During the eighth and ninth prenatal days, the embryos together with their limiting shell membranes float freely within the uterine lumen, surrounded only by the secretions of the uterus. The shell membrane is transparent, nonelastic, tough, and capable of extreme deformation. It consists of a mat of interwoven fibers which vary in external diameter, are electron dense, and show no apparent substructure. The morphology and arrangement of component fibers are similar throughout the width of the shell membrane.     

The early development of subcortical projections to presumptive somatic sensory-motor areas of neocortex in the North American opossum

Summary We have studied the early development of subcortical projections to presumptive somatic sensory-motor areas of neocortex in the North American opossumDidelphis virginiana. The opossum is born in a very immature state, 12–13 days after conception, and climbs into an external pouch where it is available for experimental manipulation. Using the retrograde transport of wheat germ agglutinin conjugated to horseradish peroxidase, we have obtained evidence that axons from the dorsal raphe and superior central nuclei, the substantia nigra, the locus coeruleus and the parabrachial nuclei reach presumptive somatic sensory-motor areas of neocortex by at least postnatal day (PND) 10. Axons showing serotonin-like immunoreactivity, presumably from the dorsal raphe and/or superior central nuclei, and axons containing tyrosine hydroxylase immunoreactivity, presumably from the substantia nigra and/or locus coeruleus, are present in the same areas at birth or shortly thereafter. Thalamic axons do not grow into comparable areas of neocortex until after PND 10. Such axons reach the subplate region of ventrolateral neocortex first and then proceed dorsomedially; by estimated PD (EPND) 21, they are present in presumptive layers I, V and VI, but they do not innervate an identified layer IV until EPND 48. The developmental sequences suggested by our study are compared with those reported for other species and are discussed in light of their importance in the formation of major sensory and motor circuits.Abbreviations qq cerebral aqueduct - ca anterior commissure - Cb cerebellum - CcD dorsal cochlear nucleus - Cd caudate nucleus - CeS superior central nucleus - CI interior colliculus - Coe locus coeruleus - CP cortical plate - CS superior colliculus - CxA cortex ammonis - DB nucleus of the diagonal band - Dien diencephalon - Dor dorsal - EP ependyma - Fac facial nucleus - GLD dorsal lateral geniculate nucleus - GM medial geniculate body - HVM ventromedial hypothalamic nucleus - IC internal capsule - Lat lateral - Med medial - ML medial lemniscus - mlf medial longitudinal fasciculus - OSL superior olivary nucleus - PB parabrachial complex - ped cerebral peduncle - PFP parafascicular nuclei - Pu putamen - RaD dorsal raphe nucleus - rfl fasciculus retroflexus - RN red nucleus - SN substantia nigra - SNc substantia nigra, pars compacta - SNr substantia nigra, pars reticulata - STh subthalamic nucleus - Tect tectum - TgV ventral tegmental area - TrMo motor trigeminal nucleus - xVB ventrobasal nucleus of thalamus     

The harderian gland and thermoregulation in the gerbil (Meriones unguiculatus)

Seven experiments with Meriones unguiculatus assessed the thermoregulatory consequence of Harderian secretion and fur anointment with lipids by autogrooming. Harderianectomy reduced the animal's capability to withstand cold (3–5°C) and wetness. During a 9 to 15 min exposure to cold-wet stress glandless animals lost an average of 4.6°C core body temperature, compared to an average loss of 1.6°C among intact animals. A comparable loss in body temperature occurred in animals whose fur lipids were removed with shampooing. Shaved animals coated with petroleum jelly withstood the cold-wet stress with a lesser decrease in body temperature than animals without the exogenous coat of jelly. The in vitro insulating qualities of fur lipids were demonstrated by blocking a cold air stream with various layers of lipids. The degree of insulation was related to the thickness of the lipid layers. Fur lipids were extracted from Harderianectomized and shampooed animals. Relative to controls, both procedures resulted in approximately 40–50% loss of fur lipds. The loss seen in gerbils following Harderianectomy was not found in the rat, golden hamster or C57 and DBA strains of inbred mice. Thus, in the gerbil the behavioral spread of Harderian lipids during a thermoregulatory groom insulated the gerbil from cold and wetness. The use of insulating lipids may be related to the gerbil's ecology in the cold deserts of Mongolia and Northeast China.     

Analysis of constant tissue remodeling in Syrian hamster Harderian gland: intra‐tubular and inter‐tubular syncytial masses

The Syrian hamster Harderian gland (HG) has a marked sexual dimorphism and exhibits an extraordinary rate of porphyrinogenesis. The physiological oxidative stress, derived from constant porphyrin production, is so high that the HG needs additional survival autophagic mechanisms to fight against this chronic exposure, provoking the triggering of a holocrine secretion in female glands that forms two types of secretory masses: intra‐tubular‐syncytial and inter‐tubular‐syncytial masses. The aim of this work was to study the development of this inter‐tubular holocrine secretion. To approach this task, we have considered that the steps developed during the formation of the so‐called invasive masses consist of the growth of epithelial cells, cell detachment from the basal lamina and invasion of surrounding tissues. The presence of these masses, particularly in the female HG, are closely linked to sexual dimorphism in redox balance and to alterations in the expression of certain factors such as cytokeratins, P‐cadherin, matrix metalloproteinases, cathepsin H, proliferating cell nuclear antigen, p53, CD‐31 and vascular endothelial growth factor, which seem to be involved in tissue remodeling. The results document unusual mechanisms of secretion in Syrian hamster HG: an extraordinary system of massive secretion through the conjunctive tissue, disrupting the branched structure of the gland.     

Anatomy, histology, histochemistry and fine structure of the Harderian gland in the South American armadillo Chaetophractus villosus (Xenarthra, Mammalia)

The anatomical, histological, histochemical and ultrastructural characteristics of the Harderian gland of the armadillo Chaetophractus villosus were described. The gland is the largest structure in the bony orbit. It is situated in the anteroventral region of the orbit. Obvious structural differences are not observed between males and females. The gland is compound-branched tubulo-alveolar, being characterized by a single layer of columnar cells surrounded by myoepithelial cells. It possesses a single excretory duct opened into the inner canthus. All glandular cells show yellow-green autofluorescence and additionally some glandular lumen may contain dense autofluorescent solid accretions. There are two peculiar and outstanding cytoplasmic features. One is represented by the smooth endoplasmic reticulum (SER), forming a closely woven meshwork. The other one is represented by membranous bodies apparently derived from the SER, RER and cytoskeleton with a Star of David configuration situated in the supranuclear region. Three types of vesicles are detected in the cytoplasm. Histochemical staining methods reveal lipids, proteins, neutral and acidic containing glycoconjugates in secretory vesicles. The mechanism of secretion appears either merocrine or apocrine. The epithelium of the intra- and inter-lobular excretory ducts suggests secretory activity. Tubulo-acinar glands similar to those seen in the lacrimal gland and nictitans glands are found related to the intralobular and main excretory ducts. The capillary network is characterized by fenestrated endothelium. The stroma possesses unmyelinated axons and plasma cells. The normal secretion of the secretory endpieces, particularly lipids, proteins and glycoconjugates, is complemented by mucous and serous secretions released by ductal cells and glands associated to the ducts.