Techniques for converting Golgi precipitate in CNS neurons into stable electron microscopic markers.

Direct electron microscopy of nervous tissue stained with the Golgi impregnation method is unsatisfactory because the cytoplasm of the cell bodies and processes of the impregnated neurons are completely filled with a compact precipitate of electron dense silver chromate. This precipitate entirely obscures the cytological details of the impregnated neurons. Because of its solidity and instability in aqueous solutions, the silver chromate is also a source of inconvenience during the preparation of the ultrathin sections. This review summarizes methods that have been developed with the aim of replacing the Golgi precipitate in CNS neurons with a more convenient electron dense material--for example, heavy metal salts or metallic particles. Conversion of the precipitate into a stable electron dense marker is done before the material is embedded for electron microscopy. The methods include lead, gold, and bromide substitution, treatment with ammonia, direct chemical reduction into metallic silver, and photoreduction of the silver chromate into silver through irradiation with ultraviolet light.     

Synaptic connections of neurones identified by Golgi impregnation: Characterization by immunocytochemical,enzyme histochemical,and degeneration methods

For more than a century the Golgi method has been providing structural information about the organization of neuronal networks. Recent developments allow the extension of the method to the electron microscopic analysis of the afferent and efferent synaptic connections of identified, Golgi-impregnated neurones. The introduction of degeneration, autoradiographic, enzyme histochemical, and immunocytochemical methods for the characterization of Golgi-impregnated neurones and their pre-and postsynaptic partners makes it possible to establish the origin and also the chemical composition of pre-and postsynaptic elements. Furthermore, for a direct correlation of structure and function the synaptic interconnections between physiologically characterized, intracellularly HRP-filled neurones and Golgi-impregnated cells can be studied. It is thought that most of the neuronal communication takes place at the synaptic junction. In the enterprise of unravelling the circuits underlying the synaptic interactions, the Golgi technique continues to be a powerful tool of analysis.     

Application of the Golgi/electron microscopy technique for cell identification in immunocytochemical, retrograde labeling, and developmental studies of hippocampal neurons.

In this study the Golgi/electron microscopy (EM) technique has been used for an analysis of the fine structure, specific synaptic connections, and differentiation of neurons in the hippocampus and fascia dentata of rodents. In a first series of experiments the specific synaptic contacts formed between cholinergic terminals and identified hippocampal neurons were studied. By means of a variant of the section Golgi impregnation procedure, Vibratome sections immunostained for choline acetyltransferase, the acetylcholine-synthesizing enzyme, were Golgi-impregnated in order to identify the target neurons of cholinergic terminals in the hippocampus. It could be shown with this combined approach that cholinergic septohippocampal fibers form a variety of synapses with different target structures of the Golgi-impregnated and gold-toned hippocampal neurons. In this report cholinergic synapses on the heads of small spines, the necks of large complex spines, dendritic shafts, and cell bodies of identified dentate granule cells are described. The variety of cholinergic synapses suggests that cholinergic transmission in the fascia dentata is a complex event. Next, the Golgi/EM technique was applied to Vibratome sections that contained retrogradely labeled neurons in the hilar region of the fascia dentata following horseradish peroxidase (HRP) injection into the contralateral hippocampus. With this combined approach some of the hilar cells projecting to the contralateral side were identified as mossy cells by the presence of retrogradely transported HRP in thin sections through these Golgi-impregnated and gold-toned neurons. Our findings suggest that the mossy cells are part of the commissural/associational system terminating in the inner molecular layer of the fascia dentata. They are mainly driven by hilar collaterals of granule cell axons that form giant synapses on their dendrites. Finally, the Golgi/EM procedure was used to study the differentiation and developmental plasticity of hippocampal and dentate neurons in transplants and slice cultures of hippocampus. Under both experimental conditions, the differentiating neurons are deprived of their normal laminated afferent innervation but develop their major cell-specific characteristics including a large number of postsynaptic structures (spines). As revealed in thin sections of gold-toned identified cells, all these spines formed synapses with presynaptic boutons suggesting sprouting of the transplanted and cultured neurons, respectively. Altogether, the present report demonstrates the usefulness of the Golgi/EM technique, particularly of the section impregnation procedure, for a variety of studies requiring the identification of individual neurons at the ultrastructural level.     

Tri-dimensional structure of the forming face of the Golgi apparatus as seen in the high voltage electron microscope after osmium impregnation of the small nerve cells in the semilunar ganglion of the trigeminal nerve

Hydrogen-ion microscopy has been used to study first and third stage tempered ferrous martensite. Both ε-carbide and cementite particles resulting from the tempering process appeared dark in the images. The image characteristics of the matrix phases in these specimens were intermediate between those of pure iron and as-quenched ferrous martensite. The sharpness of the (111) zone lines and the ‘butterfly’ pattern in {110} regions, the development of high index planes and the regularity of low index net plane edges decreased continuously with increasing carbon content of the matrix phase. The effects of carbon content and lattice strain on specimen geometry, field evaporation, hydrogen etching and field ionization are discussed. It is suggested that carbon atoms in random solid solution lead to an irregular evaporation, similar to that previously proposed for other solid solution alloys, and that the irregular evaporation is enhanced by the associated lattice strain and hydrogen etching process. The irregular end form thus produced lacks the sharp image features found in the smoother end form of the pure metal.     

Nondestructive testing of the quality of winding impregnation in electric machines

The topicality of this work is accounted for by a need to upgrade the evaluation of the quality of winding impregnation in electric machines and to raise their safety margin and reliability. Our goal is to substantiate the possibility of using the measured capacitance of the winding of an electric machine (with respect to its magnetic core) for testing the impregnation of cavities in the major insulation and assessing metrological characteristics of the proposed testing technique. Methods of electric measurements for studying the capacitance characteristics of electric-machine windings, formulae for calculating the dielectric constant of binary mixtures, and techniques for assessing the errors of indirect measurements have been used.     

A modification of the chromic tartrate silver impregnation technique for block impregnation of the central nervous system and paraffin wax embedding

This paper describes a block silver impregnation technique for the CNS. The procedure, which is quite simple, yields highly consistent and reproducible results. After fixation during 6–10 days in 10% saline formaldehyde, 4 mm thick blocks of brain are treated with chromic anhydride and sodium potassium tartrate solution for 4 days. After this period the specimens are rinsed in 0.75% silver nitrate solution to which 8–10 drops of pyridine per 100 ml of solution have been added. This is followed by impregnation for 4 days at 37°C in silver nitrate-pyridine solution identical to that used in the previous rinsing step. The impregnated blocks are reduced during 20–26 h in 1% pyrogallol to which 6 ml commercial formaldehyde per 100 ml of solution have been added, followed by dehydration in dioxan and paraffin embedding. Sections no thicker than 30 μm are then cut for histological study. This fundamentally neurofibrillar method reveals: (a) neuronal somata and their processes; (b) synaptic structures; (c) fibre bundles; and (d) cell nuclei and nucleoli.     

Plasticity in the electrophysiological properties of oxytocin neurons.

In mammals, the neurohypophysial hormone oxytocin (OT) is released into the bloodstream during labor and lactation to promote uterine contraction and milk ejection, respectively. Electrophysiological studies have established that OT neurons fire in brief, synchronized bursts during this release. During pregnancy and lactation, the intrinsic membrane and synaptic properties of OT, and to a lesser extent vasopressin (VP) neurons, are altered as a part of the adaptation to these specialized states. During lactation OT neurons specifically exhibit an enhanced rebound depolarization which could assist in instigating bursts and an increased gating of firing frequency which is correlated with an enhanced Ca(2+)-dependent after hyperpolarization. Spike broadening occurs in both VP and OT neurons, but in OT neurons this and other changes are present during late pregnancy, suggesting involvement of steroidal hormones in programming neuronal adaptations. Excitatory and inhibitory synaptic activity also are altered by reproductive state. There is a doubling of glutamatergic activity specific to OT neurons which is consistent with an increase in terminal numbers, but this is accompanied by an increase in paired-pulse facilitation, suggesting an increase in the probability of glutamate release during lactation as well. Together with profound changes in both pre- and postsynaptic GABAergic synaptic activity, these data suggest that neurosecretory, and particularly OT neuronal, properties are state-dependent. These modifications may adjust the responsiveness of these neurons to afferent stimulation during periods of increased hormone demand and thereby enhance stimulus-secretion coupling.     

A modification of microglia impregnation (author's transl)]

A modification of the Weil and Davenport (1933) silver carbonate method for microglia impregnation is described. Formalin-dextran-CaCl2 solution was used as a fixation solution. The technique is simple, reproducible and improved. The staining method includes the demonstration of as well resting as progressive microglia.     

Evaluation of fluorescent dyes for epoxy impregnation of porous ceramics

Eight different commercially available fluorescent dyes (fluorochromes) were tested for suitability for use in low-viscosity epoxy resin. Dyes were compared based on solubility in different solvents and epoxy resin and a numerical criterion for each dye's fluorochromicity. The two best dyes, based upon the brightness of each dye after illumination by a UV source, were Hostasol Red GG and Hostasol Yellow 3G. These two dyes in epoxy resin were used to visualize impregnation and remnant porosity in porous superconductor ceramic pellets. The impregnant was either cured epoxy or a low melting point alloy.     

Analysis of glass fabric impregnation using a resin drop method

Wettability of a glass fabric was studied by use of a resin drop method. The mixing ratios of epoxy resin and anhydride hardener were adopted as 1:0.5, 1:1 and 1:1.2. A catalyst of 2-ethyl-4-methylimidazole added as much as 0.1wt% of the mixed resin. A curing analysis by differential scanning calorimeter (DSC) showed that the mixed resin could be infiltrative at room temperature. An effective contact angle and the height of the resin drop onto the glass fabric preset on a flat glass plate were measured as a function of time. The wet area of the resin drop was also measured. Behaviors of the contact angle, the drop height, the net wet area and the coefficient of wettability were analyzed in the glass fabric impregnation. The resin drop method was shown to be quite effective in evaluating the capillary-mode resin penetration into the fabric.     

铝电解电容器含浸锅的设计研究

针对国内铝电解电容器生产设备存在的问题,依据新的含浸方法以及工艺要求,设计了一种含浸锅结构。简述了铝电解电容器的含浸方法和工艺流程,重点分析了该含浸锅的结构特点和工作原理。同时运用有限元分析软件,仿真得到了含浸锅的应力和应变分布图。最后,对含浸锅结构进行了优化。研究结果表明,该含浸锅结构能够实现连线生产,同时也能够可靠含浸,不污染导针和设备,提高了电容器的品质。     

Quantitative aspects of synapses on Golgi-impregnated neurons.

With the classical Golgi techniques, numerous types of neurons can be distinguished in the cerebral cortex, each with a specific dendritic geometry and pattern of axonal ramifications. In the present review we describe two techniques which allow quantification of synapses on identified neurons: (1) Golgi-rapid impregnation-gold toning-electron microscopy, and (2) Golgi-Kopsch impregnation-gold toning-electron microscopy in combination with staining of the tissue with ethanolic phosphotungstic acid (E-PTA). Both techniques were applied on neurons in the visual cortex of young and adult rabbits. By means of rotating and tilting specimens in the electron microscope, the nondistinctive ultrastructure of obliquely sectioned synapses can be circumvented, leading to precise estimates of asymmetrical vs. symmetrical synapses without complete reconstruction of the neuron.     

Iron-polyethylene glycol impregnation: An attempt to reduce shrinkage of specimens in SEM preparation

Glutaraldehyde-fixed HeLa cells were soaked in a mixture of fine cationic iron colloid and polyethylene glycol, immersed in tannic acid solution containing guanidine hydrochloride, and stained with osmic acid. The treated cells showed little shrinkage in the scanning electron microscope even after ethanol dehydration and CO₂ critical point drying. On the assumption that every HeLa cell maintained contact with each other, preservation rate was computed as 0.975 × 0.0033 in linear dimension. Microvilli on the cell surface were well preserved, and few undersirable deposits were noted on the specimen surface. This treatment was also applicable to bulk staining of tissue blocks, such as rat kidneys. The podocyte foot processes and endothelial micropores of the glomerulus were well preserved; the epithelial cells of the Bowman's urinary capsule were not collapsed; the microvilli of the brush border of the proximal convoluted urinary tubule kept their ordinary length (2 μm).     

Mathematical modeling for impregnation of reinforcing filler of fiberglasses during vacuum infusion

Composite products under vacuum infusion are formed by impregnating a reinforcing filler with resin by means of a vacuum. The task of designing the infusion process involves the development of a system of infusion. A mathematical model of the impregnation process which allows carrying out virtual technological experiments in order to find the optimal method of product impregnation is described. Examples of numerical simulation are given.