Neuroglia in the inferior olivary nucleus during normal aging and Alzheimer's disease

It is likely that neuronal loss occurs in certain brain regions in Alzheimer's Disease (AD) without any neurofibrillary pathology. In the human principle inferior olivary nucleus (PO), we have shown that neuronal loss is about 34% (Lasn et al. Journal of Alzheimer Disease, 2001; 3: 159-168), but the fate of the neuroglial cells is unknown. Since the unique network of neurons and neuroglial cells and their cohabitation are essential for normal functioning of CNS, we designed a study to estimate the total number of oligodendrocytes and astrocytes in normally aged and AD brains. The study is based on 10 control and 11 AD post-mortem human brains. An unbiased stereological fractionator method was used. We found significant oligodendroglial cell loss (46%) in AD as compared to control brains, while the total number of astrocytes showed a tendency to decrease. It is likely that the ratio of oligodendroglial cells to neurons remains unchanged even in degenerative states, indicating that oligodendroglial cells parallel neuronal loss. Astroglial cells did not increase in total number, but the ratio to neurons was significantly increased due to the neuronal loss. Using a novel unbiased quantitative method, we were able to describe significant oligodendroglial loss in the PO but the pathogenic mechanism behind remains unknown.     

Ultrastructure and Dimensions of Chloroplasts in Leaves of Three Maize (Zea mays L.) Inbred Lines and their F1 Hybrids Grown Under Moderate Chilling Stress

Influence of moderate chilling stress on vascular bundle sheath cell (BSC) and especially mesophyll cell (MC) chloroplasts of mature maize leaves was studied by electron microscopy and stereology. Plants of two inbred lines of maize, differing in their photosynthetic activity, and their F₁ hybrids were cultivated during autumn in heated or unheated glasshouse. Generally, chilling temperatures resulted mainly in the decrease in stereological volume density (VD) of both granal and intergranal thylakoids of MC chloroplasts, while the ratio of granal to all thylakoids (granality) was less affected. The VD of peripheral reticulum and plastoglobuli usually increased after cold treatment of plants. The volume of MC chloroplasts usually increased under chilling stress, the shape of the chloroplasts changed only slightly. The ultra-structure of chloroplasts differed between individual genotypes; chilling-stressed hybrid plants showed positive heterosis particularly in the granal thylakoids' VD of MC chloroplasts.     

Acute effects of pulsed microwaves and 3-nitropropionic acid on neuronal ultrastructure in the rat caudate-putamen

Ultrastructure of the medium sized "spiny" neuron in rat dorsal-lateral caudate-putamen was assessed after administration of 3-nitropropionic acid (3-NP) and exposure to pulsed microwaves. Sprague-Dawley male rats were given two daily intraperitoneal doses of 0 or 10 mg/kg 3-NP and 1.5 h after each dose were exposed to microwave radiation at a whole body averaged specific absorption rate (SAR) of 0 (sham exposure), 0.6, or 6 W/kg for 30 min. Microwave exposure consisted of 1.25 GHz radiation delivered as 5.9 micros pulses with repetition frequency 10 Hz. Tissue samples taken 2-3 h after the second sham or microwave exposure showed no injury with light microscope methods. Blinded qualitative assessment of ultrastructure of randomly selected neurons from the same samples did reveal differences. Subsequent detailed, quantitative measurements showed that, when followed by sham exposure, administration of 3-NP significantly increased endoplasmic reticulum (ER) intracisternal width, ER area density, and nuclear envelope thickness. Microwave exposure at 6 W/kg alone also significantly increased these measures. Exposure of 3-NP treated animals at 6 W/kg significantly increased effects of 3-NP on ultrastructure. Although exposure at 0.6 W/kg alone did not affect ultrastructure measures, exposure of 3-NP treated animals at 0.6 W/kg reduced the effects of 3-NP. We concluded that 3-NP changed neuronal ultrastructure and that the microwave exposures used here changed neuronal ultrastructure in ways that depended on microwave SAR and neuron metabolic status. The apparent cancellation of 3-NP induced changes by exposure to pulsed microwaves at 0.6 W/kg indicated the possibility that such exposure can protect against the effects of mitochondrial toxins on the nervous system.     

Signs of aging are apparent in the testis interstitium of Sprague Dawley rats at 6 months of age

The present study investigated the effects of aging in the testis interstitium in Sprague Dawley rats. Rats of 3, 6 and 24 months of age were used. Testes of rats (n = 5) were fixed by whole body perfusion using a fixative containing 2.5% glutaraldehyde in cacodylate buffer, processed and embedded in eponaraldite. Using 1 μm sections stained with methylene blue, qualitative and quantitative morphological studies were performed. Purified Leydig cell preparations, obtained by collagenase digestion followed by elutriation and density gradient centrifugation, were used to determine luteinizing hormone (LH; 100 ng/ml) stimulated testosterone secretory capacity per Leydig cell in vitro. Testosterone levels in the incubation medium, and testosterone and luteinizing hormone levels in serum of these three groups of rats were determined via radioimmunoassay. Morphological studies revealed that Leydig cells were more abundant in the testis interstitium at 6 and 24 months when compared to 3 months. Moreover, collagen fiber bundles were more frequently observed in the testis interstitium at older ages. Blood vessels of the testis interstitium in 24-month-old rats frequently showed partial and complete occlusion of their lumen and thickening of vessel walls. This feature was also present at 6 months, but less frequently. The results of the sterological studies revealed that the volumes of seminiferous tubules, interstitium and Leydig cells per testis was significantly higher (P < 0.05), at 6 and 24 months of age than those at 3 months. Moreover, volume of macrophages per testis was observed to be significantly higher (P < 0.05) at 6 months when compared to 3 and 24 months, and volume of connective tissue cells per testis was observed to be significantly lower (P < 0.05) at 6 and 24 months when compared to 3 months of age. No significant difference (P > 0.05) was observed for the volume of lymphatic space per testis in the three age groups studied. Volume of interstitial blood vessels per testis was not significantly different at 3 and 6 months of age, but a significantly greater (P < 0.05) volume was observed at 24 months. However, at 6 and 24 months, only 71% and 31% of the total blood vessel volumes respectively had completely open lumen in them; the rest of the blood vessels were either partially (12.5% at 6 months and 17% at 24 months) or completely (16.5% at 6 months and 52% at 24 months) occluded. The number of Leydig cells per testis was doubled at 6 and 24 months of age compared to 3 months. The average volume of a Leydig cell was not significantly different between 3 and 6 months of age, however, at 24 months a significantly lower (P < 0.05) value was observed. LH stimulated testosterone secretory capacity per Leydig cell in vitro was reduced by 50% at 6 months of age compared to 3 months; a further significant (P < 0.05) reduction was observed at 24 months. Serum testosterone and LH levels were not significantly different between 3 and 6 months of age but at 24 months a significantly lower (P < 0.05) value was observed for both of these hormones.In summary, the present study demonstrated many changes in the components of the testis interstitium in the aged Sprague Dawley rat. Modifications in the blood vessels and the occurrence of abundant collagen fibers in the interstitial space could possibly contribute to the reduced testosterone secretory capacity per Leydig cell with advancing in age. The observed Leydig cell hyperplasia could be suggested as a compensatory effort to maintain the normal androgen status of the aged rat, which is rather successful at 6 months but unsuccessful at 24 months. This investigation further revealed that these characteristic changes in the aged testis interstitium at 24 months are also present to some extent at 6 months of age in Sprague Dawley rats, suggesting that aging of the testis in this strain of rats commences early in life.     

Studies of Elodea nuttallii grown under photorespiratory conditions. III. Quantitative cytological characteristics

Abstract. Elodea nuttallii was grown in primary effluent from domestic wastewater under conditions in which CO₂ may be limiting. However, high photosynthetic inorganic carbon uptake rates have been reported for Elodea under these conditions. In order to determine if leaf cell structure showed modifications which would support proposed models of a plasma membrane (PM) bicarbonate transport system and observed high photo-synthetic rates, leaf cell ultrastructure was analysed using quantitative techniques (stereology). A Fold Index (FI) calculated for the PM showed that infolding increased surface area to 2.15 times that of an idealized cell of the same shape and size. Association Indices (Sa) showed a significant association of the mitochondria with the PM. These observations support models for an ATPase-driven HCO₃-cation cotransport system in Elodea cells. High chloroplast thylakoid surface density values (Sv) were similar to C₄:monocots and indicated high light-gathering potential. The granal/stromal (g/s), granal/cristae (g/c), and stromal/cristae (s/c) membrane ratios were similar to those of C₃ plant cells. Thus, Elodea chloroplasts and cells exhibited some structural features similar to both C₃ and C₄ plants. Membrane-bound inclusion bodies, which occupied 1% of the cell volume, were also observed in the leaf cells. Cell walls and nucleoid regions were absent in these bodies and X-ray (EDAX) analysis failed to detect any element (above the resolution limit of atomic no. 12) in substantial quantities. Inclusion bodies were observed in both the cytoplasm and the periplast and remain unidentified. A model is proposed for Elodea using an ATPase proton pump in the plasma membrane which extrudes protons into the periplast space between the cell wall and plasma membrane. This proton gradient is coupled to a bicarbonate symport.     

Quantification of organelle changes in plant suspension cultures during growth

Stereological techniques were used to quantify ultrastructural changes which occurred during maturation of cultured Paul's Scarlet rose cells. The volume and ultrastructural composition of young, dividing, unsynchronized 5-day-old cells were compared to that of mature, nondividing 14-day-old cells. The volume of the 14-day-old cells was 4-fold greater than that of the 5-day-old cells, primarily due to vacuole expansion. Numerous quantitative changes occurred in the organelle composition during cell maturation, but distinctive differences were observed in the magnitude and direction of change among the different types of organelles. There was an overall decline in the plastid population as measured by both percent of cell volume and numbers of plastids per cell. The percent of cell volume and numbers of lipid bodies increased, whereas the percent volume of the mitochondria remained relatively constant while the number per cell declined.     

Histomorphometric changes in the placenta and umbilical cord during complications of pregnancy

Pregnancy complications may cause morphological changes and circulation defects in the placenta, which may lead to morbidity and mortality in fetuses and newborns. We investigated structural changes in the placenta and umbilical cord under various abnormal maternal conditions. Placenta and umbilical cord specimens were obtained from pregnant women during labor at 37 ? 42 weeks gestation. Volumetric measurements were made for each placenta and umbilical cord using the Cavalieri method. Significant differences were observed for volumetric densities of total villi, syncytial knots, intervillous vessels and perivillous fibrin deposition. We observed particular increases in the volumetric parameters of the pre-eclampsia group compared to the other groups. The tunica media of the umbilical arteries was increased significantly with intrahepatic cholestasis.     

Occludin is overexpressed in Alzheimer's disease and vascular dementia

The tight junctions (TJs) are key players in the control of blood-brain barrier (BBB) properties, the most complex TJs in the vascular system being found in the endothelial cells of brain capillaries. One of the main TJs proteins is occludin, which anchors plasma membranes of neighbour cells and is present in large amounts in the brain endothelia. Previous studies demonstrated that disruption of BBB in various pathological situations associates with changes in occludin expression, and this change could be responsible for malfunction of BBB. Therefore in this study, applying an immunohistochemical approach, we decided to explore the occludin expression in frontal cortex (FC) and basal ganglia in ageing control, Alzheimer's disease (AD), and vascular dementia (VD) brains, as far as all these pathologies associate microangiopathy and disruption of BBB. Strikingly, we found selected neurons, astrocytes and oligodendrocytes expressing occludin, in all cases studied. To estimate the number of occludin-expressing neurons, we applied a stereological approach with random systematic sampling and the unbiased optical fractionator method. We report here a significant increase in ratio of occludin-expressing neurons in FC and basal ganglia regions in both AD and VD as compared to ageing controls. Within the cerebral cortex, occludin was selectively expressed by pyramidal neurons, which are the ones responsible for cognitive processes and affected by AD pathology. Our findings could be important in unravelling new pathogenic pathways in dementia disorders and new functions of occludin and TJs.     

Surface density and volume density measurements of chloroplast thylakoids in maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) under chilling conditions

Measurements of ultrastructural characteristics of chloroplast thylakoids are important for studies of ontogenic or ecological limitations of leaf photosynthetic functions. Most frequently, volumetric proportion of thylakoids in the chloroplast is measured; however, such measurement does not provide a direct information about the surface area of thylakoids which is most important from the functional point of view. Therefore, we adapted the stereological method using “local vertical windows” for estimating thylakoid surface area in the chloroplast volume and compared thus obtained surface density results with results of conventional volume density measurements. The methods were tested in the study of chloroplast ultrastructure in the leaves of plants of two maize (Zea mays L.) hybrid combinations, 2013×CE810 and CE704×CE810, developing in control and chilling conditions. Correlation analysis revealed a tight relationship between the granal/intergranal thylakoid surface density and volume density results, both indicating that under chilling conditions the development of the system of thylakoids in maize leaves is suppressed, while the difference is more pronounced in CE704 than in CE810 genotype, known to have a better photosynthetic performance.