Ultrastructural changes of receptor endings in prolonged action of local anesthetics

Ultrastructural changes of the terminal plates of the bushy receptors in the frog urinary bladder have been studied after two hours' exposition in 0.05% novocaine solution and one hour's exposition in 0.05% dicaine and trimecaine solution. During these periods a steady block of the receptor impulse activity develops. The local anesthetics essentially change ultramicroscopic structure of the terminals. The reaction to the anesthetics investigated has both some features in common and certain peculiarities. At each effect three types of changes can be determined, characterized with various degree of rearrangement in neurilemma, neuroplasm and organelles. Each type of the changes is supposed to reflect a certain phase of the plate reactive response. Specificities of the reaction to novocaine are minimal changes of mitochondria, accumulation of glycogen granules, deformity and decreasing amount of vesicles. Under dicaine effect mitochondria do not change, amount of vesicles increases, their form does not change; under trimecaine effect mitochondria undergo most noticeable alterations. The changes of the terminal plates observed are interpreted as adaptive. The effect of the local anesthetics on the receptors is not limited with the blockade of the sodium canals of the afferent fibers, in parallel, biochemical processes, occurring in cytosol of the terminals also change; their morphological manifestations are the ultrastructural changes observed.     

Ultrastructural changes in the receptor portion of the vestibular apparatus in response to noise

The experiments have been performed on 24 guinea pigs (48 labyrinthes), 6 of them--control and 18 have been subjected to noise (one octava) with the average geometrical in diaposone of 2,000 Hz, at intensity level 100 dB. Six animals are subjected to a single effect for 6 h and 12--to repeated effect during 6 days running, 4-6 h daily. Isolation of both the vestibular and the cochlear parts of the membranous labyrinth is performed simultaneously. This gives a possibility to study all the receptors of the internal ear as a whole. Certain ultrastructural changes in all the vestibular receptors both at a single and repeated effects are revealed. Dilatation of the granular endoplasmic reticulum is observed, situating mainly in the basal part of the cell; swelling of mitochondria is accompanied with a sharp clearance of their matrix. In some mitochondria there is a local destruction of their external and internal membranes. Moreover, in cytoplasm of the receptory cells sharply osmiophilic fibrillar structures are revealed, they resemble crists and are arranged in bundles. In some cells they are not numerous and localize mainly in mitochondria, in others--their number is greater, and they localize not only in mitochondria, but in the surrounding cytoplasm, too. Similar structures are observed in some preganglionic myelin fibers. These phenomena can be considered as development of calcification processes. The changes described, evidently, form the basis of the vestibular disorders under the noise effect.     

Silver carbonate staining reveals mitochondrial heterogeneity.

Silver staining methods, when selective, yield a high-contrast and high-resolution image in optical microscopy. A classical method for silver impregnation of mitochondria has been applied to murine tissues and reveals a marked heterogeneity among mitochondria in single cells. This heterogeneity can be detected in the optical microscope but is even more evident at the ultrastructural level. The differences in staining intensity may reflect different stages in the mitochondrial life cycle. The progressive accumulation of uranyl-argyrophilic material may be a marker of mitochondrial aging. This highly selective staining procedure may be of use in studies of mitochondrial changes under pathological conditions and during apoptosis.     

Effect of acetylcholine on the structuro-functional characteristics of bush-like interoceptors of the urinary bladder of the frog

In the preparation of the frog (Rana temporaria) isolated urinary bladder, it has been stated by morphological and physiological experiments that under an exogenic acetilcholine influence certain structural-functional changes take place in the bushy interoceptor. They are manifested as a sharp deterioration in coloration the receptor with methylene blue, the receptory plates decrease in their size, character of granuloformation changes in them. Simultaneously, an increased electrical activity is observed and, on the contrary, under the influence of atropine the impulse activity drops. It should be suggested that acetilcholine acts as an activator of metabolic and as a modulator of physiological processes in the receptor.     

Structure and synaptic organization of the osphradium of the lamellibranch mollusk Unio pectorum

The osphradial organ has been studied in Lamellibranchia--Unio pectorum--by means of scanning and transmissive electron microscopy. On the surface of the distal part of this hemosensory organ there is a distinct division into zones. The central part of the osphradial torus is occupied by the receptory zone, formed predominantly by supporting cells with microvilli and by peripheral processes of the subepithelial receptory cells. The lateral surfaces are occupied with ciliar areas of the ciliar supporting cells. In the receptory zone two types of the peripheral processes of the receptory cells are identified; they differ by the number of kinocilia and by ultrastructural organization of the apical part. Axon-like processes of the receptory cells interact with axons and dendrites of the ganglionic cells, forming axo-axonal, axodendritic and axosomatic synapses. The facts revealed demonstrate a high level of specialization of the osphradial receptory surface, connected with polymodality of this organ.     

Ultrastructure of the surface of the osphradium of the anterior gilled mollusk Murex saxatilis L

The Murex saxatilis L. (Mollusca, Prosobranchia) osphradium is of the most complex "ctenidial" type and is situated immediately behind the siphon that brings water to the mantel cavity. By means of scanning and transmissive electron microscopical investigation, three zones have been revealed on the surface of the osphradium petals: secretory, intermediate and receptory. The secretory zone occupies the lateral part of the petal and is formed with 1-2 layers of cells. The intermediate zone, as a narrow stria, is situated between the secretory and the receptory zones. Here, together with the secretory cells, the ciliary cells are present. The receptory zone occupies nearly the whole surface of the petal. It is much thicker and is formed of pseudostratified epithelium where 5-7 raws of nuclei can be counted. Besides the cell types mentioned, there are defined bipolar receptory cells in it. The apical surface of the receptory cells has up to 200 cilia, and fine peripheral processes--1-2 cilia. Presence of the complex receptory zone makes it possible to suggest certain differentiation of the stimuli already at the peripheral level of the sensory system.     

The Freshwater Dinoflagellate Woloszynskia limnetica n. sp. Membrane and Protoplasmic Structures

SYNOPSIS. A new species of a freshwater reticulate dinoflagellate, Woloszynskia limnetica , is described. The membrane structure and the shape of the membrane plates change during cell development. The rod-shaped mitochondria found among the membrane plates disappear in the adult cells; they constitute a center from which starch grains develop. Mitochondria also occur free in the endoplasm. Different physiological activities appear to be performed by morphologically identical chondriosomes. The stigma changes in different phases of the life cycle. The physicochemical properties of the ectoplasm govern the formation of specific structures of the membrane plates, which in turn determine the taxonomic position.     

线粒体PT孔参与甘草诱导MGC-803细胞凋亡的调控

不久前我们从中药中首次筛选发现了甘草能显著诱导胃癌MGC-803细胞凋亡,本文进一步研究甘草诱导MGC-803细胞凋亡过程中凋亡百分率、线粒体膜电位、胞内游离钙、DNA电泳和细胞膜通透性以及染色质DNA凝聚的时相变化,并研究了线粒体PT孔专一抑制剂环孢菌素A(CsA)对凋亡过程的影响.我们观察到,细胞膜通透性增强、胞内游离钙升高和线粒体膜电位下降为细胞凋亡的早期事件,先于凋亡峰出现、染色质凝聚和DNA电泳梯状条带出现,CsA明显抑制线粒体膜电位下降,细胞膜通透性增强和胞内游离钙变化,并极大程度地延迟细胞凋亡过程.结果提示,钙和CsA敏感性的线粒体PT孔开放参与甘草提取物诱导MGC-803细胞凋亡的调控.     

线粒体PT孔参与甘草诱导MGC—803细胞凋亡的调控

In our previous studies, we have discovered that the extract of glycyrrhiza uralensis Fisch (EGUF) can induce obvious apoptosis in gastric cancer cell Line MGC-803. Here, further investigation was carried on about the time-lapse changes of mitochondria transmembrane potential, intracellular free calcium ions, DNA electrophoresis, plasma membrane permeability and chromatin condensation during the apoptotic process of MGC-803 induced by EGUF and the influences of MPT-specific inhibitor Cyclosporin A(CsA) on these changes. Enhancement of plasma membrane permeability with PI staining, increase of intracellular free calcium ion and decrease of mitochondria transmembrane potential are early events in apoptotic cascades, prior to the appearances of apoptotic peak, chromatin condensation and DNA ladder. CsA significantly inhibited enhancement of plasma membrane permeability, change of intracellular free calcium ions and decrease of mitochondria transmembrane potential, also greatly delayed the progress of apoptosis. Thus, our results suggest that calcium and CsA-sensitive MPT is involved in the apoptosis of MGC-803 induced by EGUF.     

Composition and histotopography of receptors in antagonist muscles of the radial area of the human forearm

The m. extensor carpi radialis longus and m. flexor carpi in newborns are richly saturated in terminal sensitive apparatuses and are presented as peculiar reflexogenic zones. Quantity and topography of receptors in these zones are similar. Nevertheless, functionally different muscle areas (both in the extensor and flexor) are not equally supplied with the receptory apparatuses.     

Development of the neural and tissue components of the excretory ducts of the liver, pancreas and Odd's sphincter in human embryogeny]

The development of pancreatic, hepatic, cystic, common bile ducts, the Oddi's sphincter and their nervous apparatus were studied during prenatal human ontogenesis of fetuses and newborns. The process of formation of the nervous apparatus corresponds to the development of tissue structures of the ducts and the sphincter. The distinctions in the organization of nervous elements which are noted in adult humans are laid in the process of embryogenesis. These distinctions are especially pronounced in the structure of nervous plexuses and receptory endings. The nervous apparatus of the Oddi's sphincter region has a complex arrangement. This is the site of concentration of nerve nodules and receptory endings as well as abundant nervous connections between plexuses of the pancreatic head, duodenum and orifice zones of the both ducts. The receptors in nerve nodules and pericellular apparatuses on the bodies of ganglionic neurons were revealed.     

Light Scattering at Various Angles: Theoretical Predictions of the Effects of Particle Volume Changes

The Mie theory of scattering is used to provide new information on how changes in particle volume, with no change in dry weight, should influence light scattering for various scattering angles and particle sizes. Many biological cells (e.g., algal cells, erythrocytes) and large subcellular structures (e.g., chloroplasts, mitochondria) in suspension undergo this type of reversible volume change, a change which is related to changes in the rates of cellular processes. A previous study examined the effects of such volume changes on total scattering. In this paper scattering at 10° is found to follow total scattering closely, but scattering at 45°, 90°, 135°, and 170° behaves differently. Small volume changes can cause very large observable changes in large angle scattering if the sample particles are uniform in size; however, the natural particle size heterogeneity of most samples would mask this effect. For heterogeneous samples of most particle size ranges, particle shrink-age is found to increase large angle scattering.     

A permeability test for the study of mitochondrial injury in in vitro cultured heart muscle and endothelioid cells.

A cytochemical permeability test for the detection of injury to in situ mitochondria of cultured heart cells is presented. The test is based on the increased rate at which injured mitochondria stain for succinate dehydrogenase activity. Whereas an intact inner mitochondrial membrane limits the rate at which Nitro Blue tetrazolium and phenazine methosulphate reach succinate dehydrogenase, injured mitochondria allow these reactants to reach the enzyme more rapidly to form microscopically-observable formazan granules. The extent of staining at fixed durations of incubation with the reactants was assessed on a blind basis with pseudo dark-field microscopy, using a standardized rating scale. Differences in the staining of control and treated cells were analysed statistically by a semi-quantitative method. Treatment of the cultures with either vitamin A or chlorpromazine, resulted in more rapid mitochondrial staining. Brief pre-fixation of the cells with cold acetone also labilized the mitochondria as did a delay in the change of culture medium.     

Morphological heterogeneity of HeLa cell mitochondria visualized by a modified diaminobenzidine staining technique.

The diaminobenzidine (DAB) technique for the ultrastructural localization of sites of cytochrome c oxidase activity in animal tissues has been adapted to the visualization of mitochondria in animal cells growing in culture. The modified technique allows the staining of mitochondria in all cells in coverslip preparatins for light microscopy. Electron microscopy of thin sections of material treated by this method has revealed that all mitochondrial profiles within a cell (and only these) are stained and they exhibit a well preserved size and internal structure. Coverslip cultures of synchronized and unsynchronized HeLa (F-315) cells stained with the DAB reaction were examined under oil immersion. In the majority of the cells, mitochondria were recognized as discrete bodies in the thinner peripheral portion of the cytoplasm. This observation indicates that in a large proportion of HeLa F-315 cells, at least under the growth conditions used here, the mitochondrial complement is dividied into distinct organelles. This examination also revealed a considerable morphological heterogeneity of mitochondria, which exhibited an ovoid or short rod-like or, less frequently, long filamentous shape, with some evidence of branching. The variability in mitochondrial morphology appeared to be far more prounced between different cells than within individual cells; this cellular heterogeneity was not related in any obvious way to cell-cycle-dependent changes.     

Mitochondrial transmembrane potential changes support the concept of mitochondrial heterogeneity during apoptosis.

Dissipation of mitochondrial membrane potential (DeltaPsi(m)) and release of cytochrome c from mitochondria appear to be key events during apoptosis. The precise relationship (cause or consequence) between both is currently unclear. We previously showed in a model of serum-free cultured granulosa explants that cytochrome c is retained in a subset of respiring mitochondria until late in the apoptotic process. In this study we further investigated the issue of heterogeneity by using the DeltaPsi(m)-sensitive probe CM-H2TMRos in combination with a DNA fluorochrome. Changes of DeltaPsi(m) were assessed qualitatively by epifluorescence microscopy and were quantified using digital imaging microscopy. This approach yielded the following results: (a) CM-H2TMRos staining is a reliable and specific procedure to detect DeltaPsi(m) changes in granulosa cells explants; (b) dissipation of transmembrane potential is an early event during apoptosis preceding nuclear changes but is confined to a subpopulation of mitochondria within an individual cell; (c) in frankly apoptotic cells a few polarized mitochondria can be detected. These findings support the hypothesis that ATP needed for completion of the apoptotic cascade can be generated during apoptosis in a subset of respiring mitochondria and is not necessarily derived from anaerobic glycolysis.     

High-voltage electron microscopy of human diploid fibroblasts during ageing in vitro. Morphometric analysis of mitochondria

Since recent studies have suggested a diminished mitochondrial functional capacity in late-passage ('old') compared to early-passage ('young') normal fibroblasts and fibroblasts from the Hutchinson-Gilford (progeria) syndrome of premature ageing, we analysed whole-cell preparations on the high voltage electron microscope to look for mitochondrial and related defects. All strains examined showed considerable heterogeneity in cell size and intracellular morphology. Mitochondria were readily seen in all cells, predominantly as long slender rods with frequent branching, but occasional circular and saccular forms were also evident. Various parameters of mitochondrial mass including mean number, weight, and total length of mitochondria per cell weight tended to increase in old and progeria cells, but only the former attained statistical significance due to the heterogeneity and consequent variance. A significant finding was the decreased width of mitochondria in old and progeria cells. Cystic blebs were evident in mitochondria of some cells with an apparent increase in old and progeria fibroblasts. These blebs appeared to be due to weakening of the inner membrane, allowing dilatation of the outer membrane which otherwise appeared intact. The number of osmiophilic inclusions per cell weight, particularly lipofuscin granules and autophagic vacuoles, was significantly increased in old and progeria cells. In conclusion, despite some morphological changes, mitochondria of old and progeria cells maintain a structurally and bioenergetically adequate mass compatible with continued cellular viability.     

热休克对于高粱细胞质雄性不育系线粒体数目及其育性的影响(英文)

对高梁雄性不育系 ３ １９７Ａ（３Ａ）及其保持系 ３ １９７Ｂ（３Ｂ）均分别进行热激处理,以常温处理为对照,以处于花粉母细胞期的小穗为材料提取其线粒体,首次采用流式细胞仪测量线粒体的数量。首次观察到：经热激处理后,不育系 ３Ａ线粒体的数目增加约 ３．５倍;相反,保持系３Ｂ在常温时,线粒体数量为３Ａ的３．５５倍,热激后却无明显变化（Ｆｉｇ．１）。同时取热激处理及常温下３Ａ及３Ｂ的花药淀粉粒作碘化钾染色,结果表明：热激后３Ａ花药中出现饱满淀粉粒（Ｆｉｇ．２）;对其细胞色素氧化酶和过氧物酶同功酶的研究表明,热激后３Ａ电泳酶谱中有新条带出现,而且与３Ｂ的带型一致（Ｆｉｇ．３）。     

Aprotinin induces surface changes in malignant cells in culture: a possible mode of antitumour action.

Treatment of TRES cells with aprotinin (Trasylol) produced marked changes in their surface properties. There was a concentration-dependent increase in the surface charge density of the cells. The agglutinability of the cells was inhibited by the antiproteinase. The cells became less adhesive to concanavalin A-linked plates. The adhesion of treated TRES cells appeared to be mediated by lectin receptors with lower specificity for concanavalin A. These observations and an analysis of the kinetics of adhesion have suggested the possibility that a new class of concanavalin A receptors appears on the surface of aprotinin-treated TRES cells, which might increase immunogenicity of the cells.     

Modality of the bush-like receptors

In an isolated preparation of the Rana temporaria urinary bladder after a simultaneous morphological and physiological investigation a, structural-functional differentiation of free bushy receptors has been demonstrated. According to arborization character and to the appearance of deferent fibers, the receptors are divided into two types. The first type receptors have a simple structure, a long deferent poorly branching myelin fiber, terminating in a diffuse bush near blood vessels. The second type receptors are of a more complex structure. Their myelin fibers, when leaving the fasciculus, are 4-8 times shorter than the first type receptors, undergo dichotomic and trichotomic divisions several times, and in their distal part they form two initial myelin segments. Their receptory apparatus has a tree-like bush-shaped form and consists of several compact bushes. The impulse activity of the receptors is also characterized by two types of action potentials, differing in their amplitude. When responding to a mechanical stimulation, the high voltage impulse frequency changes, when sodium chloride concentration is increased, the low voltage impulse frequency changes. There is a certain localization of the zones in the preparation from which it is possible to obtain predominantly either low voltage or high voltage responses. The response to the low voltage reaction proves to be obtained from the first type receptors, and that to the high voltage reaction--from the second type receptors. Thus, in the frog, that is on a low phylogenetic stage, differentiation of the free bushy sensitive terminal into mechano- and chemoreceptors is already outlined.     

Flow cytometric analysis of the effects of high protein diet on isolated rat liver mitochondria

We have investigated changes that occur in mitochondria obtained from the livers of rats that had been maintained on a high protein diet (80% casein instead of 20%) for 6 months. Liver homogenates were separated by centrifugation into a mitochondrial fraction, a nuclear fraction and the supernatant fluid of the nuclear fraction (nuclear wash). Rhodamine-123 was used to selectively stain mitochondria depending upon their membrane potential. The stained organelles were processed through a flow cytometer where the fluorescent stains were excited by the 488 nm wavelength of a laser and the resultant fluorescence signals analysed. After 6 months on a high protein diet, mitochondria displayed an increase in the fluorescence associated with rhodamine-123 uptake in both mitochondrial and nuclear wash fractions, while mitochondrial fluorescence in the nuclear fraction showed a heterogeneous distribution. This was interpreted as an increase in membrane potential in most of the liver mitochondria under these nutritional conditions, with a certain degree of heterogeneity. These functional changes may be correlated with morphological alterations previously reported and show the usefulness of flow cytometry for biochemical analysis of isolated mitochondria.