Biomedical subjects

G Danscher

Publications and source records attributed to G Danscher.

At least 55 records · Page 3Linked to original sources

PubMed1998

Computer-assisted visualization of the rat epididymis: a methodological study based on paraffin sections autometallographically stained for zinc ions.

A concept for the computer-assisted visualization of tubular organs is presented. Unmarked histological zinc-stained serial sections from the epididymis of the Wistar rat were aligned to demonstrate the concept. Virtual images were made through the aligned sections and served as controls for the alignment process. Animation of the serial sections and the virtual images revealed new information about the structure of the organ under investigation. The analysis was used to upgrade the anatomical knowledge of rat epididymis by describing how the epididymal duct runs through the structure. The proximal parts of the epididymis contain large communicating septa of connective tissue dividing the caput and the upper part of the corpus epididymidis into segments. The tortuousness was high in the caput with many turns within a small area of the epididymis, whereas longer loops were found in the lower part of the corpus and cauda epididymidis. The tube of the vas deferens was found to become an integrated part of the ductal system in the cauda epididymidis, although it was histologically easy to distinguish from the epididymal duct. The total number of cross-sections of the ductus epididymidis in the 2254, 15-microm-thick, tissue sections analysed was 104700, giving a minimum length of the ductal system of 1.5 m.

Animals↗

PubMed1998

Histochemical tracing of zinc ions in the rat testis.

To detect free zinc ions in the rat testes four rats were transcardially perfused with Na2S, and the seminiferous tubules from two other rats were incubated in Na2S. Sections from the two sources were autometallographically (AMG) developed, whereby zinc sulphide crystal lattices created in the tissue by the sulphide treatment were silver enhanced. Light microscopical analysis showed zinc ions in primary spermatogonia until the zygotene primary spermatocytes (stage I), in late pachytene spermatocytes (stages XII and XIII), and in late spermatids from step 15 to step 19 (stages I-VIII). The highest intensity of AMG grains was detected in the residual bodies and tails of step 19 spermatids. Grains were occasionally found in the cytoplasm of Leydig cells. Sections from animals treated with the chelator diethyldithiocarbamate prior to sulphide treatment showed a complete lack of AMG staining. At ultrastructural levels the AMG grains were found in smooth-surfaced endoplasmic reticulum of all spermatogonial stages, and in the acrosome, midpiece, and tail of late spermatids. The presence of zinc ions in preleptotene spermatocytes and cytoplasmic lobes of late spermatids suggests a specific role of free zinc at the onset of meiosis and at spermiation.

Animals↗

PubMed1997

Demonstration of vessels in CNS and other organs by AMG silver enhancement of colloidal gold particles dispersed in gelatine.

We present a new autometallographic technique for demonstrating vessels and other small cavities at light microscopy (LM) and electron microscopy (EM) levels. It is possible to obtain detailed knowledge of the 3-D appearance of the vascular system by exchanging blood with a 40 degrees C, 8% gelatine solution containing colloidal gold particles (gold gelatine solution, GGS) and ensuing silver enhancement of the gold particles by autometallography (AMG). The GGS-AMG technique demonstrates the vascular system as a dark web that can be studied in cryostat, vibratome, methacrylate, paraffin and Epon sections at all magnifications. The infused GGS becomes increasingly viscous and finally becomes rigid when the temperature falls below 20 degrees C. An additional advantage of this technique is the fact that none of the tested counterstains or immunotechniques interfere with this AMG approach. The GGS-AMG technique is demonstrated on rat brains but can be applied to any organ. We believe that the present technique is valuable for both experimental studies and routine pathology.

Animals↗

PubMed1997

Increased amount of zinc in the hippocampus and amygdala of Alzheimer's diseased brains: a proton-induced X-ray emission spectroscopic analysis of cryostat sections from autopsy material.

Zinc has been implicated as a contributing cause of the neuropathology of Alzheimer's disease (AD), but consensus on the zinc content of AD brains has not yet been established. In the present study, multi-element PIXE was used to measure zinc in cryostat sections of brain tissue from AD patients and from normal control subjects. Compared to their age-matched controls, the AD patients showed an increase in zinc in the hippocampal and amygdalar regions. The instrumental PIXE assays do not show whether the zinc changes are due to altered zinc in the boutons of Zinc-ENriched (ZEN) neurons, i.e., zinc ions in synaptic vesicles, or to changes in the amount of zinc tightly bound to macromolecules. We hypothesise that the increased zinc level is caused by an increase in the amount of ZEN terminals. Such an increase could be the result of a sprout of ZEN terminals in diseased areas of the brain.

Aged↗

PubMed1997

Ultrastructural localization of zinc ions in the rat prostate: an autometallographic study.

BACKGROUND: The prostate contains high amounts of free zinc ions which are excreted into the seminal fluid. The extra- and intracellular distribution of zinc ions using the highly specific autometallographical (AMG) method is described. METHODS: Prostates from sulfide-perfused rats were excised, and the ZnS crystals were silver-enhanced to sizes detectable by the electron and light microscope. RESULTS: AMGZnS grains were found primarily in the acinic lumen of the lateral lobes. The dorsal lobe stained only faintly, while the ventral lobe was void of grains. At ultrastructural levels, the presence of zinc ions was confined to apical secretory vesicles and lysosome-like structure of the epithelium of mainly the lateral lobes. CONCLUSIONS: We suggest a constant secretion of zinc ions from the epithelial cells into both the acinar lumen and the intercellular canaliculi, and that the zinc enriched secretory cells in the prostate belong to a system of glandular cells that uses zinc ions to aggregate macromolecules to be excreted.

Animals↗

PubMed1997

Distinct differences in partial oxygen pressure at micrometer ranges in the rat hippocampal region.

A mapping at micrometer ranges of the partial oxygen pressure in the rat hippocampus was performed. The oxygen tension in the rat hippocampal region was measured using a glass oxygen microsensor in 30-microm steps along straight lines at a set of stereotactic coordinates. In the hippocampus the pattern of the oxygen tensions reflected the autometallographic zinc sulphide (AMG(ZnS)) pattern, i.e. the pattern of zinc enriched (ZEN) terminals. The highest levels of oxygen tension were recorded in the areas that are most heavily stained with the autometallographic zinc sulphide (AMG(ZnS)) method, like hilus fasciae dentatae. The zinc ions located in synaptic vesicles of the ZEN terminals can also be demonstrated by AMG silver amplification in brains from animals in vivo treated with sodium selenite. This method depends on the presence of a substantial reduction capacity of the tissues as selenite ions (SeO(2)(3)-) must to be reduced to selenide ions (Se2-) before the catalytic zinc selenide crystals can be formed. At some point, either during the transport from the infusion site to the actual target tissue or in the target tissue itself, selenium is reduced from Se(+ IV) to Se(- II). The importance of the reduction capacity of the target tissue in this process is demonstrated by the fact that areas found to have the highest concentration of zinc ions, e.g. hilus fasciae dentatae and the mossy fibres of CA3, are almost unstained after 1 h of i.p. Na2SeO3 exposure. An explanation of this phenomenon could be that the reduction process Se(+ IV) <==> Se(- II) leading to the formation of Se2- is moved to the left by the presence of oxygen, thus inhibiting the precipitation of ZnSe crystals. It is suggested that the subtle oxygen pressure pattern found in the rat hippocampus might also reflect essential biological zinc-related mechanisms vital to brain function.

Animals↗

PubMed1997

Histochemical demonstration of zinc ions in human epididymis using autometallography.

A recently described autometallographic technique, allowing demonstration of chelatable zinc in human biopsy material, was applied to cryostat sections from biopsies of human epididymis. Sections from the rat epididymis were used as control materials to examine the quality of the method compared with a previously used autometallographic method. The human epididymis exhibits heavy staining in the head of the epididymis and only small amounts of zinc in the body and tail of the organ. The zinc staining was found in the apical part of the ciliated cells and in the lumen. The present technique can be used to localize zinc ions at ultrastructural levels. Zinc grains were localized in lysosome-like bodies and secretory granules of the ciliated cells. The luminal staining was present as free, evenly dispersed zinc grains or attached to sperm cells and stereocilia in the lumen. The large differences in staining patterns along the epididymal tract in humans and rats suggest that zinc ions are important for the maturation of sperm cells.

Biopsy↗

PubMed1997

Optical slicing and 3-D characterization of hippocampal capillaries in the rat visualized by autometallographic silver enhancement of colloidal gold particles.

In order to visualize the vascular system of the rat brain, 10 Wistar rats were perfused transcardially with glutaraldehyde and a 40 degrees C gold-gelatine solution. The brains were post-fixed with glutaraldehyde and vibratomized into 100-micron-thick slices, and the gold particles were developed by autometallography. In this way, the colloidal gold particles in the vessels became encased in silver and thereby made visible. The developed gold staining is stable and does not interfere with further dehydration and counterstaining. Images were frame grabbed during optical slicing, and classic stereograms and 'shadow' 3-D images were produced. We found a high variation of capillary density in the hippocampal region reflecting known subregional structures. The silver-enhanced vessels acted as natural markers and made it possible to study and measure aspects of the complexity of dehydration and staining artifacts. We found a non-linear shrinking of 13-17% in the x- and y-directions and a spatial shrinking up to 50% in some regions after the dehydration and staining process. This observation may be of interest not only in relation to tissue subjected to this fixation protocol but also to other fixation procedures. The gold-gelatine autometallographic technique and the present stereograms can release data for stereological use as well.

Animals↗

PubMed1997

The effect of selenium on the localization of autometallographic mercury in dorsal root ganglia of rats.

The autometallographic technique was used to demonstrate the localization of mercury in dorsal root ganglia of adult Wistar rats. The animals were either exposed to mercury vapour, 100 micrograms Hg m-3, 6 h day-1, 5 days per week, or treated with organic mercury in the drinking water, 20 mg CH3HgCl per litre, for 4 weeks. The effect of orally administered sodium selenite on the pattern of intracellular distribution of mercury in these two situations was investigated. In rats exposed to mercury vapour alone, faint staining was present in ganglion cells. The selenite induced a conspicuous increase in the number of stained cells and in the intracellular staining intensity. In rats treated with organic mercury, mercury deposits were detected within ganglion cells and macrophages. The number of mercury-containing cells was increased by co-administration of selenite. In addition, satellite cells, the capsule and vessel walls were faintly stained. Twenty weeks after cessation of the organic mercury treatment, mercury staining was reduced. Again, selenite treatment enhanced staining intensity. When studied using the electron microscope, mercury was restricted to lysosomes, irrespective of treatments. The present study shows that the deposition of autometallographic mercury in the dorsal root ganglia depends on the chemical type of mercury, the co-administration of selenite and the length of the survival period.

Administration, Inhalation↗

PubMed1997

Histochemical demonstration of zinc ions in ejaculated human semen.

A revised in-vitro technique for autometallographic demonstration of chelatable zinc in the human ejaculate is presented, and the localization of the loosely bound pool of zinc ions is described in semen smears and at the ultrastructural level. In semen smears, black autometallographic (AMG) grains indicated the presence of zinc ions dispersed between the spermatozoa. These AMG grains have the same size as grains associated with the sperm tail and may have the same origin. EM analysis of AMG-developed smears fixed in osmium suggested that the detected zinc ions might be related to huge protein molecules present in semen and adhering to the surface of the spermatozoa. Spermatozoa in AMG-stained smears exhibited zinc ions in the midpiece and head, and also joined to the membrane of the tail. Washed spermatozoa exhibited zinc ions only within the midpiece. Ultrastructurally, they were found located in the helecine mitochondria. A few grains were found in the acrosome of the washed spermatozoa. Treatment with the chelating agent DEDTC resulted in complete bleaching of the zinc staining. These findings and the fact that calcium EDTA acid blocks the plasma and surface staining, but not the acrosomal and mitochondrial staining, suggest that chelatable zinc ions exist in two separate pools in human semen.

Adult↗

PubMed1997

Autometallographic demonstration of zinc ions in rat sperm cells.

An in-vitro technique for autometallographic (AMG) demonstration of chelatable zinc in electroejaculated sperm cells and spermatozoa from the epididymis is presented and the localization of zinc ions in rat spermatozoa is described. Sperm cells from caput epididymis showed zinc staining in all parts of the tail and a sparse, dispersed staining in the acrosome. Spermatozoa from cauda epididymis showed heavy staining in the acrosome but no staining in the tail, or post-acrosomal part of the sperm head. This distinct acrosomal AMG staining was also found in ejaculated spermatozoa, but additionally a segmentation of the tail was seen based on differences in staining intensity. The membrane penetrating chelator diethyldithiocarbamate (DEDTC) was found to block the AMG staining whereas calcium-EDTA, known not to pass through cell membranes, did not influence the staining, proving that the detected zinc ions are intracellularly located. Two different approaches for demonstrating the presence of a chelatable zinc pool at electron microscope levels are presented, and the ultrastructural presence of AMG grains located in the acrosome and in the mitochondria of the midpiece is demonstrated. It is postulated that an exchange of zinc ions takes place between the epididymal epithelium and the sperm cells as they pass along the epididymal duct.

Acrosome↗

PubMed1997

Dentinal and pulpal uptake of mercury from lined and unlined amalgam restorations in minipigs.

The aim of the present investigation was to examine dentin and pulp in amalgam-restored teeth in minipigs with respect to presence of mercury, and to evaluate whether lining of restored cavities with a resin-modified glass ionomer cement had any effect on the penetration of mercury. Class I and V cavities were cut in 49 posterior and canine teeth of 4 minipigs. 35 cavities were restored with amalgam and 14 with composite material. 26 of the amalgam cavities were lined with a resin-modified glass ionomer cement. Following observation periods of 17-28 months, the pigs were killed and the jaws sectioned. After decalcification, the teeth were processed for light or electron microscopy examinations. Mercury was visualized by autometallography. Silver-enhanced mercury was found in all teeth with amalgam, whereas teeth with composites were devoid of mercury. Mercury could be traced in the odontoblast processes, in the body of odontoblasts, and on rare occasions in the nerve tissue of the pulp from lined and from unlined amalgam restorations. The present study thus demonstrates transport of mercury through dentinal tubules to the pulp, and that lining of cavities with a resin-modified glass ionomer cement does not prevent penetration of mercury.

Animals↗

PubMed1997

Autometallographic silver enhancement of zinc sulfide crystals created in cryostat sections from human brain biopsies: a new technique that makes it feasible to demonstrate zinc ions in tissue sections from biopsies and early autopsy material.

We present a new technique that allows zinc ions in synaptic and secretory vesicles of biopsy and early autopsy material (< 2 hr post mortem) to be transformed to nanometer-sized zinc sulfide crystal lattices for subsequent autometallographic (AMG) development. Human brain biopsies, or other tissue samples containing zinc-enriched (ZEN) cells, are frozen in liquid nitrogen or by CO2 gas immediately after removal. The tissue blocks are cut in a cryostat and the sections placed on glass slides. The slides are transferred to an H2S exposure chamber placed in a -15 C freezer. After 1-24 hr of gas exposure the sections are removed from the chamber, fixed while thawing, and dehydrated. The sections are then exposed to an AMG developer. AMG causes silver enhancement of zinc sulfide crystal lattices created in the tissues through the H2S exposure, making them visible. It is imperative that the tissues are frozen instantaneously after removal, because loosely bound or free zinc ions start leaving their vesicular compartment soon after death. The AMG technique can, despite inadequate fixation and damage to the tissue caused by freezing, also be used to trace zinc ions at ultrastructural levels, and it is demonstrated that zinc ions in the human neocortex are located in synaptic vesicles. In the few human biopsies analyzed thus far, the light microscopic pattern created by the silver-enhanced ZEN terminals resembles that seen in the neocortex of rat brain. The technique has been applied to cryostat sections from neocortex biopsies of five individuals undergoing brain surgery. Biopsies from three patients resulted in satisfactory AMG-stained sections. Rat brains removed and frozen immediately after decapitation constituted the material on which the present technique was developed. Such material results in an almost uniform high quality of staining, and we found that unexposed sections can be stored for at least 5 months at -80 C without ensuing significant loss of AMG staining intensity.

Animals↗

PubMed1996

Electron Microscopical Autometallography: Immunogold-Silver Staining (IGSS) and Heavy-Metal Histochemistry

Immunogold-silver staining (IGSS) utilizes a histochemical method called autometallography (AMG) to amplify tiny gold particles to sizes easily visible both in light and electron microscopy. In both applications it is advisable to use the smallest possible gold diameters (1-6 nm) to obtain the highest sensitivity, thus, allowing minute amounts of the target substance to be demonstrated. Gold labels smaller than 10 nm in diameter have been clearly shown to give the highest labeling densities of antigen-antibody binding sites. AMG can be used for the detection of catalytic crystal lattices of metallic gold and silver, and sulfides or selenides of mercury, silver, copper, bismuth, and zinc. The method has its roots in "physical development" technique, transplanted from photography to histology by Liesegang at the beginning of this century. In 1981, a series of papers were published by one of us with the purpose of introducing a reliable and easy-to-handle technique for light microscopical and ultrastructural studies. AMG has a multitude of applications apart from its use in detecting tissue metals. These include the highly sensitive and efficient in situ colloidal gold tracing of peptides, proteins, and amines by immunocytochemistry using the IGSS method, of carbohydrates by lectin IGSS, and of nucleic acids by IGSS in situ hybridization, IGSS in situ polymerase chain reaction, and IGSS in situ self-sustained sequence replication-based amplification (in situ 3SR) techniques, the last two even performing with single-copy sensitivity. Applications of pre- and postembedding AMG for semithin and ultrathin tissue sections are described.

Journal Article↗

PubMed1996

The autometallographic zinc-sulphide method. A new approach involving in vivo creation of nanometer-sized zinc sulphide crystal lattices in zinc-enriched synaptic and secretory vesicles.

A new version of Timm's sulphide silver method involving in vivo binding of zinc ions in zinc enriched terminals is presented. By injecting sodium sulphide into the vena cava of deeply anaesthetized animals, it is possible to bind chemically the vesicular zinc, i.e. chelatable zinc (zinc ions), in secretory and synaptic vesicles, in the form of zinc sulphide crystal lattices. Four minutes after the intravenous injection the animal is perfused transcardially with a phosphate-buffered solution of glutaraldehyde, glutaraldehyde and formaldehyde, or with a saline solution. The nanometer-sized catalytic crystals can then be silver-amplified in cryostat and vibratome sections by exposure to an autometallographic developer. It is demonstrated that contemporaneously with silver enhancement, the zinc sulphide crystals are transformed to the corresponding silver sulphide crystals. For ultrastructural studies, autometallographic development of vibratome sections is recommended. From these sections tissue blocks are cut from the areas of interest, blockstained with osmium tetroxide and embedded in Epon. This approach results in a zinc-specific autometallographic staining of the sections of a hitherto unseen, high technical quality.

Animals↗

PubMed1996

Histochemical localization of zinc ions in the epididymis of the rat.

In the present study, the autometallograpic zinc sulphide technique, an improved version of the original Timm sulphide-silver method, was used. This technique reveals a particular pool of ionic zinc that is chelatable by diethyldithiocarbamate. At the light microscopical level, no reaction for zinc was found in tissues of young prepubertal rats. In adult mating and non-mating rats low zinc staining was found in the head and intermediate epididymis whereas the tail of the epididymis demonstrated high levels of zinc ions. Sections from the epididymal tail revealed a 'compartmentalization', based on pronounced differences in staining intensity along the epididymal ducts. At higher magnification zinc ions were found in the apical part of the principal cell and in the lumen. At the ultrastructural level autometallographic grains were located in vesicles and in lysosome-like structures of the apical parts of the principal cells. The luminal grains were found either associated with sperm cells, with the surface of the large microvilli (stereocilia), or free in the seminal fluid. The variation in content of zinc ions in the epididymal epithelium and lumen suggests that zinc ions are secreted into the lumen from the epididymal tail and may somehow be involved in maturation of the sperm cells.

Animals↗

PubMed1995

Argyrophilic nucleolar organizer regions. A revised version of the Ag-NOR-staining technique.

Silver staining techniques developed to demonstrate argyrophilic nucleolar organizer regions (Ag-NORs) have been widely applied in a variety of cell kinetic studies, using the mean number of AgNORs in tumour cells as a marker for malignancy of certain types of neoplasms. However, the AgNOR techniques currently available are not entirely satisfactory, as unspecific silver precipitates readily form in the sections. On the other hand, the contrast staining may be so weak as to render identification of the AgNORs difficult. In the present study, some of the key factors influencing the outcome of AgNOR staining were evaluated in a more systematic way. A modified AgNOR staining procedure is now proposed, giving highly contrasting AgNORs with minimal unspecific silver precipitation, thus facilitating both manual and computerized counting. The new technique involves the use of microwave irradiation in order to shorten the processing time, the use of gelatin as a protective colloid, and a Farmer's solution to optimize the specificity of the technique.

Adrenal Cortex Neoplasms↗

PubMed1995

Quantitative and qualitative distribution of mercury in organs from arctic sledgedogs: an atomic absorption spectrophotometric and histochemical study of tissue samples from natural long-termed high dietary organic mercury-exposed dogs from Thule, Greenland.

Organs from 10 sledgedogs fed methyl mercury-containing organs and meat from predatory marine animals also eaten by humans in the Thule district of Greenland, were examined histochemically for cellular distribution of mercury, and the organ concentrations of mercury were quantified by atomic absorption spectrometry (total Hg). In selected organs the methyl mercuric level was determined by gaschromatography. The highest concentration of total mercury was found in mesenterial lymph nodes followed by liver and kidneys, which indicates that the lymphatic system might play an important role in the regulating transport of mercury to target organs. The concentrations were age-related, and the results suggest that demethylation takes place in all organs except skeletal muscles, but lowest in CNS. The distribution of mercury at cellular and subcellular levels was studied by the autometallographic technique. The atomic absorption spectrometric and autometallographic results were in good agreement. The brain mean concentration in the oldest group was 438 micrograms/kg, a level much lower than what has been reported to cause effects in the human central nervous system. However, if humans over a period of e.g. 50 years eat Arctic marine meat and accumulate mercury in the same way as dogs, the possibility that this may have health implications cannot be entirely excluded.

Aging↗