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PubMed1988

Earliest enamel deposits of the rat incisor examined by electron microscopy, electron diffraction, and electron probe microanalysis.

In order to describe initial events in enamel mineralization and to help characterize inorganic-organic interactions in this tissue, the earliest rod and interrod enamel in mandibular incisors from normal young adult (100 gm) rats, perfused with 100% ethylene glycol, has been studied by transmission electron microscopy, selected area electron diffraction, and high-spatial-resolution electron probe microanalysis. Diffraction and probe data were correlated precisely from the same extracellular regions of the tissue. Sites were examined progressively as a function of location a) from the most recently deposited enamel adjacent to ameloblasts toward the dentin-enamel junction and b) from the apical portion of the tooth longitudinally toward its incisal end. Electron diffraction patterns consistent with that of a poorly crystalline hydroxyapatite were generated at all locations. Diffraction characteristics changed only slightly toward that of more crystalline hydroxyapatite at different locations. Earliest apical enamel generated molar Ca/P ratios in a range of 0.99-1.46 (average 1.24 +/- 0.15). Molar Ca/P ratios of the first enamel interrod elements increased from approximately 1.24 at ameloblast-enamel boundaries to approximately 1.40 at the dentin-enamel junction, small changes corresponding to those observed in electron diffraction characteristics.

Animals↗
PubMed1983

Element analysis of Lewy and adrenal bodies in Parkinson's disease by electron probe microanalysis.

Two possible interpretations of the origin of Lewy bodies and adrenal bodies found in Parkinson's disease are now under discussion: a disorder of (1) catecholamine metabolism or (2) sphingomyelin lipidosis. From the electron probe microanalysis of Lewy bodies and adrenal bodies, we find that the Lewy body contains sulfur, calcium, and phosphorus, and the adrenal body also contains these three elements. Furthermore, a positive correlation was obtained between the X-ray intensity of the sulfur and the diameter of adrenal body. For Lewy bodies, this correlation was not obtained. The results suggest that a common mechanism may exist for the production of Lewy and adrenal bodies, although they differ somewhat in their accumulation of sulfur. It is considered that both structures may originate, in part, from degenerated protein containing sulfur.

Adrenal Glands↗
PubMed1989

Detoxification of cadmium. Ultrastructural study and electron-probe microanalysis of the midgut in a cadmium-resistant strain of Drosophila melanogaster.

The midgut of a cadmium-resistant strain of Drosophila melanogaster has been studied at the ultrastructural level and by electron-probe microanalysis (EPMA). Chronic exposure to cadmium leads to a concentration of the metal in a lysosomal system developed in both anterior and posterior segments of the midgut, where it coexists with copper and sulfur. This mechanism apparently ensures a permanent cadmium detoxification and prevents cellular injury. Wild-type flies fed on a cadmium-contaminated medium manifest the same detoxification process. As a result of contamination, copper is stored along the entire length of the midgut, including a part of the middle-midgut previously named 'copper-accumulating region'. Our data demonstrate that the midgut, particularly the posterior segment, is an accumulative organ for both cadmium and copper. The involvement of the metallothionein system in the detoxification process is discussed.

Animals↗
PubMed1977

Electron probe microanalysis of the chemical elemental content of human follicular fluid.

Follicular fluid samples were obtained by puncturing follicles of ovaries in situ from patients undergoing laparotomy. Sodium, potassium, chloride, magnesium, calcium, phosphorus, and sulfur concentrations measured by electron probe microanalysis were similar to those of blood, with minimal differences. This suggested that culture media in which these electrolytes are added in concentrations similar to those of serum are appropriate for culture of the human oocyte.

Calcium↗
PubMed1984

Localization of zinc in the thymic reticulum of mice by electron-probe microanalysis.

Thymulin, a thymic hormone, is a nonapeptide requiring zinc for biological activity. It has been shown that epithelial cells, forming part of the thymic reticulum, secrete this hormone and/or store it within cytoplasmic vacuoles. X-ray electron-probe microanalysis (EPMA) has been used to detect zinc in the thymus. Low concentrations of zinc have been demonstrated in the dense granules contained in clear vacuoles of some epithelial cells in normal and ZnCl2-injected mouse thymuses, thus suggesting that the metal may be coupled to the peptide before the secretion of the hormone from the cells.

Animals↗
PubMed1993

Calcium displacement by lanthanum in subcellular compartments of rat ventricular myocytes: characterisation by electron probe microanalysis.

OBJECTIVE: Much of the calcium in cardiac myocytes resides in a kinetic compartment defined by rapid exchange and rapid displacement by La3+. The aim of this study was to ascertain the subcellular location of this calcium pool. METHODS: Electron probe microanalysis (EPMA) was used to measure redistribution of total calcium in rat cardiac myocytes after 30 s in 1 mM lanthanum. "Cells", mitochondria, and myofibrils were separately analysed. The data permitted calculating a third (difference) compartment, containing primarily sarcoplasmic reticulum, sarcolemma, and T tubule lumen. Total calcium levels in junctional sarcoplasmic reticulum and T tubule lumen were also measured directly. RESULTS: Lanthanum decreased total "cell" calcium from 1.4 (SEM 0.3) to 0.5(0.4) mmol.kg-1 dry weight (p < 0.05); loss from the third (difference) compartment was primarily responsible. Simultaneously, junctional sarcoplasmic reticulum increased from 5.1(0.6) to 8.2(0.9) mmol.kg-1 dry weight (p < 0.05), while the T tubule lumen was unchanged. CONCLUSIONS: These results suggest that lanthanum displaced calcium primarily from sarcolemmal sites and that calcium redistributed to sarcoplasmic reticulum from an intracellular source.

Animals↗
PubMed1988

Localization of cations by pyroantimonate. II. Electron probe microanalysis of calcium and sodium in skeletal muscle of mouse.

A new formulation of the pyroantimonate (PA) method for localization of calcium and sodium is proposed and evaluated in mouse skeletal muscle. This study, performed at the ultrastructural level by means of transmission electron microscopy (TEM) and electron probe microanalysis (EPMA), completes a previous work done at the optical level with analytical ion microscopy (AIM), which enabled us to define the appropriate composition of fixatives. In our present experiments, calcium and sodium were shown localized in various cell structures, e.g., T-tubules, glycogen, granules, nuclei. For AIM, the best fixatives were characterized by PA supersaturation, which resulted in smaller crystals and a high rate of penetration in the presence of paraformaldehyde and either phenol or collidine. Contrary to the findings at the optical level, collidine did not give satisfactory results at the ultrastructural level. The method of floating sections on the microtome trough was an important cause of cation displacement. We found that alkalinization of the floating medium significantly decreased ion loss. The technique also provided an indication of the form of these elements: free or easily liberated cations were precipitated into coarse PA deposits; electron-positive chelates were "stained" by PA; neutral chelates were not stained, but some of them could be detected by EPMA. This PA method should make possible more precise localization of cellular calcium, such as in glycogen metabolism, and perhaps detection of movements of cytoplasmic calcium and sodium.

Animals↗
PubMed1978

Electron probe microanalysis of red blood cells. II. Cation changes during maturation.

To understand the sequence of maturation of membrane transport and hemoglobin production during erythropoiesis, we have measured the K, Na, and Fe content in single mature red blood cells and bone marrow cells of dog using electron probe microanalysis (EPMA). Mature red blood cells of dog are low in potassium (LK) and high in sodium. These cells are derived from erythroblastic stem cells, which are high in potassium (HK) and low in sodium. This change from HK stem cells to LK red cells occurs in the marrow. The ratio of K/Na was found to be less than 0.2 independent of Fe/(K + Na) in circulating red cells. However, a significant number of marrow cells had both low K/Na and low Fe/(K + Na). We conclude that the changes in cation transport properties responsible for the conversion of HK to LK cells occur before the synthesis of hemoglobin in at least some marrow cells.

Animals↗
PubMed1991

Large and rapid changes of myofibrillar total calcium during the cardiac cycle. Electron probe microanalysis of voltage-clamped guinea-pig ventricular myocytes.

At 36 degrees C and 2 mM [Ca2+]o, single guinea-pig ventricular myocytes were voltage clamped with patch electrodes. When paired pulsing had potentiated the contraction to the maximum, the cells were shock-frozen for electron probe microanalysis (EPMA). Shock-freezing was timed at the end of diastole (-80 mV) or at different times during systole (+5 mV). The same paired-pulse protocol was applied to another group of myocytes from which contraction was recorded and [Ca2+]i was estimated by microfluospectroscopy (50 microMNa-Indo-1). In potentiated cells, during the first pulse, contraction peaked within 128 +/- 25 ms after start of depolarization. [Ca2+]i peaked within 25 ms to 890 /+- 220 nM (mean +/- SEM) and fell within 100 ms to about 450 nM. sigma Camyo, the total calcium concentration in the overlapping myofilaments (A-band), was measured by EPMA in 17 potentiated myocytes. During diastole, sigma Camyo was 2.6 +/- 0.4 mmol/kg dry weight (dw), which can be converted to 0.65 mM (mmoles per liter myofibrillar space). Since [Ca2+]i was 180 nM, we estimate that 99.97% of total calcium is bound. A time-course for systolic sigma Camyo was determined by shock-freezing 13 cells at different times after start of depolarization to +5 mV. sigma Camyo was 5.5 +/- 0.3 mmol/kg dw (1.4 mM) after 15-25 ms, 4.6 +/- 0.5 mmol/kg dw (1.1 mM) after 30-45 ms, and 3.1 mmol/kg dw (0.8 mM) after 60-120 ms. The fast time-course of sigma Camyo suggests that calcium binds to and unbinds from troponin C at a fast rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1997

Assay of acetylcholinesterase activity and elemental composition in brain compartments by electron probe microanalysis.

Using histochemical techniques the distribution of acetylcholinesterase (AChE) activity in rat brain has been well established. These assays are based on the precipitation of a specific deposit containing copper-ferricyanide. The amount of locally deposited product is proportional to the AChE activity in the examined area of the section. Thus, the amounts of copper or (and) iron as the deposited histochemical labels mark the distribution and magnitude of the enzyme activity. The present protocol describes an approach to obtain by electron probe microanalysis (EPMA) a map of copper deposited in different brain regions via AChE histochemistry. EPMA allows a wide range of chemical elements to be analyzed. Thus, together with the determination of the copper concentration, elemental contents (Na, Cl, K and Ca) are measured as the specific environment for local AChE activity.

Acetylcholinesterase↗
PubMed1997

Electron probe microanalysis and transverse microradiography studies of artificial lesions in enamel and dentin: a comparative study.

In cardiology and research involving the interactions between restorative materials and dental hard tissues, it is important for small changes in tooth mineral content to be measurable. Currently, transverse microradiography (TMR) is the most accepted tool for the above purpose. Electron Probe Microanalysis (EPMA) can yield both qualitative identification of elements and quantitative compositional information. The purpose of this study was to compare the mineral distribution in well-defined artificial lesions, in dentin and enamel, by the use of both TMR and EPMA on the same sample. The good correlation between the two sets of data validates EPMA as a technique and helps in the interpretation of its results. The data from TMR analysis are expressed as vol% of mineral, while EPMA gives the levels of Ca and phosphate in wt%. The conversion between the two sets of data is complicated by the fact that local density is, as yet, unknown.

Analysis of Variance↗
PubMed1995

Scanning electron microscopy-electron probe microanalysis study of the interface between apatite and wollastonite-containing glass-ceramic and rabbit tibia under load-bearing conditions after long-term implantation.

Glass-ceramic implants containing oxy- and fluoroapatite [Ca10(PO4)6(O, F2)] and beta-wollastonite (CaSiO3) were studied under load-bearing conditions in a segmental replacement model in the tibia of the rabbit. A 16-mm segment of the middle of the tibial shaft was resected at a point distal to the junction of the tibia and the fibula. The defect was replaced by a 15 mm-long hollow, cylindrical implant that was fixed by intramedullary nailing using Kirschner wire. The implants were 9 mm in diameter and 15 mm long bearing a central hole 3.05 mm in dianeter. The rabbits used were killed 6 months, 1 year, 18 months, and 2 years after implantation. The interface between the bone and the glass-ceramic was investigated by scanning electron microscopy-electron-probe microanalysis (SEM-EPMA). None of the glass-ceramic implants broke, and the glass-ceramic had bonded directly to the bone tissue without any intervening soft tissue. A calcium-phosphorus layer (Ca-P layer) was observed at the glass-ceramic/bone interface. This layer was 30-100 microns thick at 6 months after implantation, 60-110 microns thick at 1 year after implantation, 80-200 microns thick at 18 months, and 120-350 microns thick at 2 years. At the lateral surface of the glass-ceramic uncovered by the bone, the calcium-phosphorus layer was 50-80 microns thick at 6 months after implantation, 250-450 microns thick at 1 year, 300 approximately 400 microns thick at 18 months, and 300 microns thick at 2 years. The thickness of the calcium-phosphorus layer increased moderately after long-term implantation. However, it was difficult to estimate the rate of increase in the thickness of calcium-phosphorus layer.

Animals↗
PubMed1993

Microheterogeneity of subsarcolemmal sodium gradients. Electron probe microanalysis in guinea-pig ventricular myocytes.

1. The effect of stimulation on possible subsarcolemmal sodium accumulation was studied in ventricular myocytes (2 mM [Ca2+]o, 36 degrees C). By trains of eighteen paired voltage-clamp pulses (180 ms to 0 mV, 20 ms to -45 mV, 180 ms to +50 mV, 620 ms to -45 mV) unloaded contractions were potentiated to an optimum. 2. Potentiation reversibly enlarged and prolonged the diastolic tail currents due to Na(+)-Ca2+ exchange. Eighteen pulse pairs were estimated to provide a sodium influx that could increment the total intracellular sodium concentration (sigma Na(i)) by no more than 0.5 mM. 3. Potentiation reversibly increased the current at +50 mV and made it more noisy. Cell-attached recordings with a second electrode attributed this noise to the activation of K+ (Na) channels. In inside-out patches, a comparable channel activity was obtained with 40 mM sodium. Hence, the cell-attached recordings suggest that potentiation can increase intracellular sodium concentration to 40 mM. 4. Electron probe microanalysis (EPMA) measured sigma Na in a volume within 20 nm of the inner side of the sarcolemma. Potentiation reversibly increased sigma Na20nm to 40 +/- 7 mM. When stimulation was terminated, sigma Na20nm fell within 8 s to 37 +/- 8 mM and within 3 min to 19 +/- 6 mM. In unstimulated cells sigma Na20nm was 17 +/- 5 mM. 5. In potentiated cells, shock-frozen at early systole, sigma Na fell with a space constant of 28 nm from the sarcolemma to the centre; at 1 microns distance sigma Na was 12 +/- 3 mM. The steep gradient suggests that sodium does not freely diffuse and sigma Na20nm is controlled by transmembrane fluxes rather than by cell dialysis. 6. sigma Na20nm data were distributed with peaks at 5, 30 and 60 mM. Quantitative elemental digital imaging demonstrated patches with 60-80 mM sigma Na20nm alternating with others of 0-15 mM sigma Na20nm. This 'sodium microheterogeneity' suggests that Ca2+ efflux at low sigma Na20nm and K+(Na) channel activation at high sigma Na20nm can operate simultaneously.

Animals↗
PubMed1977

Electron probe microanalysis in the study of gallstones.

Detailed information on the structure and composition of gallstones was obtained using an electron probe microanalyser in conjunction with the other methods. Gallstones were studied layer by layer without greatly disturbing the arrangement of the materials present. Elements, including trace elements such as copper, iron, and manganese, were identified and their distributions mapped. The range of the method was extended to determine the character and distribution of certain chemical groups present by treating sections of gallstone with reagents which contained easily detected elements. The nature of the bonding of the sulphur in the stones was studied by examining the sulphur-X-ray spectrum. Pigmented sulphur-containing deposits were found to contain sulphur in a low valence state but taurine conjugates and the sulphate groups of mucosubstances were not detected. Microcrystalline apatite present in the stones contained some manganese and seemed to be implicated in the absorption of the low valence sulphur compound and in the nucleation of some stones.

Calcium↗