Optical Disector Research Articles

Alterations in expression of Cat‐315 epitope of perineuronal nets during normal ageing, and its modulation by an open‐channel NMDA receptor blocker, memantine

AbstractThe perineuronal net (PNN), a specialized aggregate of the extracellular matrix, is involved in neuroprotection against oxidative stress, which is now recognized as a major contributor to age‐related decline in brain functions. In this study, we investigated the age‐related molecular changes of PNNs using monoclonal antibody Cat‐315, which recognizes human natural killer‐1 (HNK‐1) glycan on aggrecan‐based PNNs. Western blot analysis showed that the expression levels of Cat‐315 epitope in the hippocampus were higher in middle‐aged (MA, 12‐month‐old) mice than in young adult (YA, 2‐month‐old) mice. Although there were no differences in the expression levels of Cat‐315 epitope between old age (OA, 20‐month‐old) and MA mice, Cat‐315 immunoreactivity was also detected in astrocytes of OA mice. To focus on Cat‐315 epitope in PNNs, we used YA and MA mice in the following experiments. Optical disector analysis showed that there were no differences in the numbers of Cat‐315‐positive (Cat‐315+) PNNs between YA and MA mice. Fluorescence intensity analysis indicated that Cat‐315 immunoreactivity in PNNs increased with age in the dorsal hippocampus, which is mainly involved in cognitive functions. Administration of an open‐channel blocker of NMDA receptor, memantine, reduced the expression levels of Cat‐315 epitope in the hippocampus. Furthermore, the numbers of glutamatergic and GABAergic terminals colocalized with Cat‐315 epitope around parvalbumin‐positive neurons were decreased by memantine. These findings provide novel insight into the involvement of PNNs in normal brain ageing, and suggest that memantine may counteract the age‐related alterations in expression levels of Cat‐315 epitope via regulation of its subcellular localization.

Alterations in expression of Cat-315 epitope of perineuronal nets during normal ageing, and its modulation by an open-channel NMDA receptor blocker, memantine.

The perineuronal net (PNN), a specialized aggregate of the extracellular matrix, is involved in neuroprotection against oxidative stress, which is now recognized as a major contributor to age-related decline in brain functions. In this study, we investigated the age-related molecular changes of PNNs using monoclonal antibody Cat-315, which recognizes human natural killer-1 (HNK-1) glycan on aggrecan-based PNNs. Western blot analysis showed that the expression levels of Cat-315 epitope in the hippocampus were higher in middle-aged (MA, 12-month-old) mice than in young adult (YA, 2-month-old) mice. Although there were no differences in the expression levels of Cat-315 epitope between old age (OA, 20-month-old) and MA mice, Cat-315 immunoreactivity was also detected in astrocytes of OA mice. To focus on Cat-315 epitope in PNNs, we used YA and MA mice in the following experiments. Optical disector analysis showed that there were no differences in the numbers of Cat-315-positive (Cat-315+ ) PNNs between YA and MA mice. Fluorescence intensity analysis indicated that Cat-315 immunoreactivity in PNNs increased with age in the dorsal hippocampus, which is mainly involved in cognitive functions. Administration of an open-channel blocker of NMDA receptor, memantine, reduced the expression levels of Cat-315 epitope in the hippocampus. Furthermore, the numbers of glutamatergic and GABAergic terminals colocalized with Cat-315 epitope around parvalbumin-positive neurons were decreased by memantine. These findings provide novel insight into the involvement of PNNs in normal brain ageing, and suggest that memantine may counteract the age-related alterations in expression levels of Cat-315 epitope via regulation of its subcellular localization.

Late postnatal shifts of parvalbumin and nitric oxide synthase expression within the GABAergic and glutamatergic phenotypes of inferior colliculus neurons.

The inferior colliculus (IC) is partitioned into three subdivisions: the dorsal and lateral cortices (DC and LC) and the central nucleus (ICC), and serves as an integration center of auditory information. Recent studies indicate that a certain population of IC neurons may represent the non-GABAergic phenotype, while they express well-established cortical/hippocampal GABAergic neuron markers. In this study we used the optical disector to investigate the phenotype of IC neurons expressing parvalbumin (PV) and/or nitric oxide synthase (NOS) in C57BL/6J mice during the late postnatal period. Four major types of IC neurons were defined by the presence (+) or absence (-) of PV, NOS, and glutamic acid decarboxylase 67 (GAD67): PV+ /NOS- /GAD67+ , PV+ /NOS+ /GAD67+ , PV+ /NOS- /GAD67- , and PV- /NOS+ /GAD67- . Fluorescent in situ hybridization for vesicular glutamate transporter 2 mRNA indicated that almost all GAD67- IC neurons represented the glutamatergic phenotype. The numerical densities (NDs) of total GAD67+ IC neurons remained unchanged in all subdivisions. The NDs of PV+ /NOS- /GAD67+ neurons and PV- /NOS+ /GAD67- neurons were reduced with age in the ICC, while they remained unchanged in the DC and LC. By contrast, the NDs of PV+ /NOS+ /GAD67+ neurons and PV+ /NOS- /GAD67- neurons were increased with age in the ICC, although there were no changes in the DC and LC. The cell body size of GAD67+ IC neurons did not vary according to the expression of PV with or without NOS. The present findings indicate that the expression of PV and NOS may shift with age within the GABAergic and glutamatergic phenotypes of IC neurons during the late postnatal period. J. Comp. Neurol. 525:868-884, 2017. © 2016 Wiley Periodicals, Inc.

Absolute number of parvicellular and magnocellular neurons in the red nucleus of the rat midbrain: a stereological study.

The absolute number of parvicellular and magnocellular neurons in the red nucleus was estimated using design-based stereological counting methods and systematic random sampling techniques. Six young adult male rats, and a complete set of serial 40-μm glycolmethacrylate sections for each rat, were used to quantify neuronal numbers. After a random start, a systematic subset (i.e. every third) of the serial sections was used to estimate the total volume of the red nucleus using Cavalieri's method. The same set of sampled sections was used to estimate the number of neurons in a known subvolume (i.e. the numerical density Nv ) by the optical disector method. Multiplication of the total volume by Nv yielded the absolute number of neurons. It was found that the right red nucleus consisted, on average, of 8400 parvicellular neurons (with a coefficient of variation of 0.16) and 7000 magnocellular neurons (0.12). These total neuronal numbers provide important data for the transfer of information through these nuclei and for species comparisons.

Neurostereology protocol for unbiased quantification of neuronal injury and neurodegeneration.

Neuronal injury and neurodegeneration are the hallmark pathologies in a variety of neurological conditions such as epilepsy, stroke, traumatic brain injury, Parkinson’s disease and Alzheimer’s disease. Quantification of absolute neuron and interneuron counts in various brain regions is essential to understand the impact of neurological insults or neurodegenerative disease progression in animal models. However, conventional qualitative scoring-based protocols are superficial and less reliable for use in studies of neuroprotection evaluations. Here, we describe an optimized stereology protocol for quantification of neuronal injury and neurodegeneration by unbiased counting of neurons and interneurons. Every 20th section in each series of 20 sections was processed for NeuN(+) total neuron and parvalbumin(+) interneuron immunostaining. The sections that contain the hippocampus were then delineated into five reliably predefined subregions. Each region was separately analyzed with a microscope driven by the stereology software. Regional tissue volume was determined by using the Cavalieri estimator, as well as cell density and cell number were determined by using the optical disector and optical fractionator. This protocol yielded an estimate of 1.5 million total neurons and 0.05 million PV(+) interneurons within the rat hippocampus. The protocol has greater predictive power for absolute counts as it is based on 3D features rather than 2D images. The total neuron counts were consistent with literature values from sophisticated systems, which are more expensive than our stereology system. This unbiased stereology protocol allows for sensitive, medium-throughput counting of total neurons in any brain region, and thus provides a quantitative tool for studies of neuronal injury and neurodegeneration in a variety of acute brain injury and chronic neurological models.

Cryostat Slice Irregularities May Introduce Bias in Tissue Thickness Estimation: Relevance for Cell Counting Methods.

Stereological techniques using the optical disectors require estimation of final section thickness, but frozen tissue irregularities may interfere with this estimation. Cryostat slices from rodent nerve tissues (dorsal root ganglia, spinal cord, and brain), cut at 16, 40, and 50 μm, were digitized with a confocal microscope and visualized through 3D software. Geometric section thickness of tissue (T geom) was defined as tissue volume/area. Maximal section thicknesses (T max), from the top to the bottom of the section, were measured in a random sample of vertical ZX planes. Irregularities were mostly related to blood vessels traversing the tissue and neuronal somas protruding over the cut surfaces, with other neuron profiles showing a fragmented appearance. Irregularities contributed to increasing the distance between the tops and bottoms of slices sectioned in different laboratories. Significant differences were found between T max and T geom for all thickness studies and counting frames (p<0.01). The T geom/T max average rate was 68.4-85.7% in volumes around cell profiles (∼600-1,200 μm2) and 83.3-91.8% in subcellular samples (∼25-160 μm2). Confocal microscopy may help to assess tissue irregularities, which might lead to an overestimation of tissue volume if section thickness is estimated by focusing on the top and bottom of the sections.

Huntington's Disease (HD): Neurodegeneration of Brodmann's Primary Visual Area 17 (BA17).

Huntington's disease (HD), an autosomal dominantly inherited polyglutamine or CAG repeat disease along with somatomotor, oculomotor, psychiatric and cognitive symptoms, presents clinically with impairments of elementary and complex visual functions as well as altered visual-evoked potentials (VEPs). Previous volumetric and pathoanatomical post-mortem investigations pointed to an involvement of Brodmann's primary visual area 17 (BA17) in HD. Because the involvement of BA17 could be interpreted as an early onset brain neurodegeneration, we further characterized this potential primary cortical site of HD-related neurodegeneration neuropathologically and performed an unbiased estimation of the absolute nerve cell number in thick gallocyanin-stained frontoparallel tissue sections through the striate area of seven control individuals and seven HD patients using Cavalieri's principle for volume and the optical disector for nerve and glial cell density estimations. This investigation showed a reduction of the estimated absolute nerve cell number of BA17 in the HD patients (71,044,037 ± 12,740,515 nerve cells) of 32% in comparison with the control individuals (104,075,067 ± 9,424,491 nerve cells) (Mann-Whitney U-test; P < 0.001). Additional pathoanatomical studies showed that nerve cell loss was most prominent in the outer pyramidal layer III, the inner granular layers IVa and IVc as well as in the multiform layer VI of BA17 of the HD patients. Our neuropathological results in BA17 confirm and extend previous post-mortem, biochemical and in vivo neuroradiological HD findings and offer suitable explanations for the elementary and complex visual dysfunctions, as well as for the altered VEP observed in HD patients.

Subclass-specific formation of perineuronal nets around parvalbumin-expressing GABAergic neurons in Ammon's horn of the mouse hippocampus.

Perineuronal nets (PNNs) are closely associated with parvalbumin-positive (PV+) neurons, and play a major role in controlling developmental neural plasticity. Considering the recent advances in classification of PV+ neurons, here we aimed to clarify whether PNNs might be associated with specific subclasses of PV+ neurons in the hippocampus. In this study, we labeled PNNs by Wisteria floribunda agglutinin (WFA), and classified PV+ neurons based on the combination of cellular location, molecular expression (neuropeptide Y [NPY], somatostatin [SOM], special AT-rich sequence-binding protein-1 [SATB1]), and retrograde tracing through stereotaxic injection of Fluoro-Gold (FG) into the medial septum. The criteria of each subclass can be summarized as follows: axo-axonic cells, PV+/SATB1-/NPY- cells in the stratum pyramidale; basket cells, PV+/SATB1+/NPY- cells in the stratum pyramidale; bistratified cells, PV+/SATB1+/NPY+ cells in the stratum pyramidale; oriens-lacunosum-moleculare (O-LM) cells, PV+/SOM+/FG- cells in the stratum oriens; hippocampo-septal projection (H-S) cells, and PV+/SOM+/FG+ cells in the stratum oriens. The ratios of formation of WFA+ PNNs around each subclass of PV+ neurons were estimated according to the optical disector principle. The vast majority (over 90%) of putative PV+ basket cells were surrounded by PNNs, while only a minor population (less than 10%) of putative PV+ axo-axonic, O-LM, and H-S cells were enwrapped with PNNs. The ratios of formation of PNNs around putative PV+ bistratified cells were intermediate (25-50%). These findings indicate that PNNs may be specifically associated with PV+ basket cells, and also provide a key to understand the functional significance of PNNs and PV+ neurons in the hippocampus.

Perineuronal nets affect parvalbumin expression in GABAergic neurons of the mouse hippocampus.

Recent studies have suggested that the perineuronal net (PNN), a specialised extracellular matrix structure, and parvalbumin (PV), an EF-hand calcium-binding protein, are involved in the regulation of plasticity of neural circuits. Here, we aimed to quantitatively estimate the relationship between the two plasticity regulators, PV and PNNs, in the hippocampus of young adult mice. Dual fluorescence staining for PV and Wisteria floribunda agglutinin (a broad PNN marker) showed that a substantial population of PV-expressing (PV(+) ) GABAergic neurons lacked PNNs. Optical disector analysis demonstrated that there were fewer PNN(+) neurons than PV(+) neurons. The ratio of PNN expression in PV(+) neurons was generally lower in the dendritic layers than in the principal cell layers, whereas the ratio of PV expression in PNN(+) neurons was effectively 100%. The mean PV fluorescence was significantly higher in PNN(+) /PV(+) neurons than in PNN(-) /PV(+) neurons. Cumulative frequencies for single-cell PV fluorescence indicated that intensely stained PV(+) neurons tend to be enwrapped by PNNs, whereas weakly stained PV(+) neurons are likely to lack PNNs. We digested the PNNs by a unilateral injection of chondroitinase ABC (chABC) into the dorsal CA1 region. Although the densities of PV(+) neurons remained unchanged, the PV fluorescence declined 7days after chABC injection. Quantitative real-time polymerase chain reaction analysis demonstrated a reduction in PV mRNA expression following chABC injection. These findings indicate that the presence or absence of PNNs affects the relative PV expression in GABAergic neurons in the hippocampus.

Stereological estimation of ovarian volume and number of follicles in low dose of Vitex agnus castus treated mice

Vitex agnus castus (VAC) has been proven to have a wide range of biological activities. It is commonly used in the treatment of menstrual disorders resulting from corpus luteum deficiency, including premenstrual symptoms and spasmodic dysmenorrheal, for certain menopausal conditions, and for insufficient lactation. The aim of this study was to investigate the effects of low dose of VAC essential oil on ovarian volume and oocyte number in mice by stereological technique. In this study 10 young adult female BALB/c mice were randomly divided into two groups: group 1 as control group and group 2 which received 75 mg/kg VAC essential oil via gastric gavage for seven consecutive days. At the eighth day, animals were euthanized and the ovaries were quickly removed, weighted and fixed in buffered formalin. The samples were processed by routine and standard paraffin embedding and using the isotropic Cavalieri design were sectioned. Eight to twelve sections from each ovary were sampled through systematic random sampling and were stained by H&E. The volume of the ovary (cortex and medulla) was estimated by unbiased stereological methods and the numbers of the follicles were estimated using an optical disector. The results showed that volume of ovaries and ovarian cortex was significantly increased in treated mice compared to control but volume of ovarian medulla didn’t changed. Stereological result revealed that number of primordial follicle was decreased in treated group but this changes was not significant (P>0.05) and number of preantral follicles was significantly increased in treated mice (P 0.05). It can be concluded that VAC essential oil can affected structure and maturation of ovarian follicles even in low dose.

Early exposure to bisphenol A alters neuron and glia number in the rat prefrontal cortex of adult males, but not females

Previous work has shown that exposure to bisphenol A (BPA) during early development can alter sexual differentiation of the brain in rodents, although few studies have examined effects on areas of the brain associated with cognition. The current study examined if developmental BPA exposure alters the total number of neurons and glia in the medial prefrontal cortex (mPFC) in adulthood. Pregnant Long-Evans rats were orally exposed to 0, 4, 40, or 400-μg/kg BPA in corn oil throughout pregnancy. From postnatal days 1 to 9, pups were given daily oral doses of oil or BPA, at doses corresponding to those given during gestation. Brains were examined in adulthood, and the volume of layers 2/3 and layers 5/6 of the mPFC was parcellated. The density of neurons and glia in these layers was quantified stereologically with the optical disector, and density was multiplied by volume for each animal. Males exposed to 400-μg/kg BPA were found to have increased numbers of neurons and glia in layers 5/6. Although there were no significant effects of BPA in layers 2/3, the pattern of increased neuron number in males exposed to 400-μg/kg BPA was similar to that seen in layers 5/6. No effects of BPA were seen in females or in males exposed to the other doses of BPA. This study indicates that males are more susceptible to the long-lasting effects of BPA on anatomy of the mPFC, an area implicated in neurological disorders.

Curcumin prevents the structural changes induced in the rats' deep cerebellar nuclei by sodium metabisulfite, a preservative agent

ObjectiveTo evaluate the the possible neurotoxic effects of sulfite and the protective potential of curcumin on the deep cerebellar nuclei using stereological methods. MethodsThe rats were randomly divided into five experimental groups (n=6): Group I: distilled water, Group II: Olive oil, Group III: Curcumin (100 mg/kg/day), Group IV: Sodium metabisulfite (25 mg/kg/day), and Group V: Sodium metabisulfite+curcumin. At the end of 56 d, the right cerebellar hemispheres were removed and assigned to stereological studies. The total volume and total neuron number of deep cerebellar nuclei were assessed using Cavalieri and optical disector methods, respectively. ResultsThe data showed ∼20% and ∼16% decrease was respectively observed in the total volume and the total neuron number of the deep cerebellar nuclei of the sulfite-treated rats in comparison to the distilled water group (P<0.04). However, no significant change was observed in the total volume and neuronal number of the deep cerebellar nuclei in sulfite+curcumin-treated rats and curcumin played a protective role against sulfite. Curcumin or its vehicle (olive oil) did not induce any significant changes. ConclusionsCurcumin, the main part of the turmeric, could prevent the structural changes induced in the deep cerebellar nuclei by sodium metabisulfite, a preservative agent, in rats.

Evaluation of PCNA, Caspase 3 and E-cadherin on the Ventral Prostate of Soy Treated Rats

The incidence of certain cancers, including prostate cancer, is considerably higher in western countries than in Southeast Asia. Many studies have linked soy consumption to the lower incidence of prostate cancer in these countries. Twenty male Sprague-Dawley rats were randomized and divided into a control group and a group treated with soy. The ventral prostates of each animal were sectioned and stained with hematoxilin-eosin for morphological description and immunostained to detect PCNA, cleaved caspase 3 and E-cadherin immunoreactivities. Estimates of the number of total epithelial cells and the number of epithelial cells immunoreactive to PCNA and cleaved caspase 3 were calculated using the optical disector technique. Measurement of E-cadherin was carried out by calculating the volume fraction of epithe- lium immunostained by E-cadherin. Soy treated group showed atrophy in the epithelium and a diminished expression of PCNA, cleaved caspase 3 and E-cadherin, which means there is a reduced cell proliferation, apoptosis through caspases way and cell adherence. It can be concluded that soy treatment induces atrophy in the epithelium by reducing cell prolif- eration.

Protective role of curcumin against sulfite-induced structural changes in rats’ medial prefrontal cortex

ObjectivesSodium metabisulfite as a food preservative can affect the central nervous system. Curcumin, the main ingredient of turmeric has neuroprotective activity. This study was designed to evaluate the effects of sulfite and curcumin on the medial prefrontal cortex (mPFC) using stereological methods.MethodsThirty rats were randomly divided into five groups. The rats in groups I–V received distilled water, olive oil, curcumin (100 mg/kg/day), sodium metabisulfite (25 mg/kg/day), and sulfite + curcumin, respectively, for 8 weeks. The brains were subjected to the stereological methods. Cavalieri and optical disector techniques were used to estimate the total volume of mPFC and the number of neurons and glial cells. Intersections counting were applied on the thick vertical uniform random sections to estimate the dendrites length, and classify the spines. Non-parametric tests were used to analyze the data.ResultsThe mean mPFC volume, neurons number, glia number, dendritic length, and total spines per neuron were 3.7 mm3, 365 000, 180 000, 1820 µm, and 1700 in distilled water group, respectively. A reduction was observed in the volume of mPFC (∼8%), number of neurons (∼15%), and number of glia (∼14%) in mPFC of the sulfite group compared to the control groups (P < 0.005). Beside, dendritic length per neuron (∼10%) and the total spines per neuron (mainly mushroom spines) (∼25%) were reduced in the sulfite group (P < 0.005).DiscussionThe sulfite-induced structural changes in mPFC and curcumin had a protective role against the changes in the rats.

Unbiased estimation of chloroplast number in mesophyll cells: advantage of a genuine three-dimensional approach.

Chloroplast number per cell is a frequently examined quantitative anatomical parameter, often estimated by counting chloroplast profiles in two-dimensional (2D) sections of mesophyll cells. However, a mesophyll cell is a three-dimensional (3D) structure and this has to be taken into account when quantifying its internal structure. We compared 2D and 3D approaches to chloroplast counting from different points of view: (i) in practical measurements of mesophyll cells of Norway spruce needles, (ii) in a 3D model of a mesophyll cell with chloroplasts, and (iii) using a theoretical analysis. We applied, for the first time, the stereological method of an optical disector based on counting chloroplasts in stacks of spruce needle optical cross-sections acquired by confocal laser-scanning microscopy. This estimate was compared with counting chloroplast profiles in 2D sections from the same stacks of sections. Comparing practical measurements of mesophyll cells, calculations performed in a 3D model of a cell with chloroplasts as well as a theoretical analysis showed that the 2D approach yielded biased results, while the underestimation could be up to 10-fold. We proved that the frequently used method for counting chloroplasts in a mesophyll cell by counting their profiles in 2D sections did not give correct results. We concluded that the present disector method can be efficiently used for unbiased estimation of chloroplast number per mesophyll cell. This should be the method of choice, especially in coniferous needles and leaves with mesophyll cells with lignified cell walls where maceration methods are difficult or impossible to use.

Abnormalities in oligodendrocyte clusters in the inferior parietal cortex in schizophrenia are associated with insight

Background and Objectives: Deficits in oligodendrocytes have been consistently reported in the brains of patients with schizophrenia and include alterations in the clustering pattern of oligodendrocytes. Recently it has been shown that oligodendrocyte progenitors proliferate in the adult mammalian brain to form oligodendrocyte clusters (OlC). We previously found a deficit of oligodendrocytes in layer 3 of the inferior parietal lobule (IPL) in subjects with schizophrenia with poor insight into disorder. We hypothesized that the number of OlC might be reduced in schizophrenia subjects with poor insight. Methods: Nissl-stained sections from the Stanley �Parietal Collection� from male schizophrenia subjects (n = 24) that have poor, fair, or good insight into their disorder and normal matched controls (n = 24) were studied. The numerical density (Nv) of OlC was estimated in layer 3 of BA 39 and BA 40 by optical disector method. Results: The Nv of OlC was 23% lower in BA 39 and 30% lower in BA40 in the schizophrenia group compared to the control group (p<0.01). Normal hemispheric differences in the Nv of OlC in BA 39 were absent in the schizophrenia group. The Nv of OlC was significantly decreased in BA39 in the subgroup with poor insight and in BA40 in the subgroups with fair and good insight as compared to controls. In BA40 lower Nv of OlC (-40%, p<0.01) was found in the subgroup with adolescent onset of disease as compared to controls. Conclusions: The deficit of OlC may be associated with altered proliferation and/or maturation of oligodendrocyte progenitors in schizophrenia.

Chronic intrauterine exposure to endotoxin does not alter fetal nephron number or glomerular size

A reduced nephron endowment early in life adversely impacts on long-term functional reserve in the kidney. A recent study has shown that acute exposure to chorioamnionitis during late gestation can adversely impact on nephrogenesis. The present study aimed to examine the effects of chronic, low-dose endotoxin exposure in utero, during the period of nephrogenesis, on nephron number and glomerular size in preterm lambs. Ewes were administered either endotoxin (lipopolysaccharide; 1 mg/day) or saline at 110-133 days of gestation (term approximately 147 days) via surgically implanted osmotic minipumps within the amniotic cavity. The ewes were induced to deliver preterm at 133 days gestation and the kidneys of the lambs were analysed at 8 weeks after term-equivalent age. Nephron number per kidney was determined using a combined optical disector and fractionator stereological approach; renal corpuscle size was also measured stereologically. At 8 weeks after term-equivalent age there was no significant effect of in utero exposure to endotoxin on bodyweight or kidney weight and there were no significant differences in nephron number, nephron density or renal corpuscle volume between groups. We conclude that chronic intrauterine inflammation during the period of nephrogenesis may not adversely impact on the number of nephrons formed within the kidney or on the volume of the renal corpuscle.

Effects of postnatal hyperoxia exposure on the rat dentate gyrus and subventricular zone

Premature newborns may be exposed to hyperoxia in the first postnatal period, but clinical and experimental works have raised the question of oxygen toxicity for the developing brain. However, specific analysis of hyperoxia exposure on neurogenesis is still lacking. Thus, the aim of the present study was to evaluate possible changes in the morphometric parameters of the main neurogenic sites in newborn rats exposed to 60 or 95 % oxygen for the first 14 postnatal days. The optical disector, a morphometric method based upon unbiased sampling principles of stereology, was applied to analyse cell densities, total volumes, and total cell numbers of the dentate gyrus (DG) and subventricular zone (SVZ). Apoptosis and proliferation were also studied by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling method and anti-ki67 immunohistochemistry, respectively. Severe hyperoxia increased the percentage of apoptotic cells in the DG. Moderate and severe hyperoxia induced a proliferative response both in the DG and SVZ, but the two neurogenic sites showed different changes in their morphometric parameters. The DG of both the hyperoxic groups showed lower volume and total cell number than that of the normoxic one. Conversely, the SVZ of newborn rats exposed to 95 % hyperoxia showed statistically significant higher volume and total cell number than SVZ of rats raised in normoxia. Our findings indicate that hyperoxia exposure in the first postnatal period affects both the neurogenic areas, although in different ways, i.e. reduction of DG and expansion of SVZ.

S.12.08 Lithium-regulated genes in peripheral cell models of bipolar-affective patients

Previous studies have suggested that affective disorders are characterized by glial pathology. In this context, it has been hypothesized that elevated S100B serum and cerebrospinal fluid levels may represent a suitable surrogate marker. However, brain studies on the cellular distribution pattern of S100B in depressed patients are lacking so far. Such analyses are crucial, since S100B has been detected in various other cell types, even outside the central nervous system.Therefore, we performed a first postmortem analysis on this topic in the hippocampus – which is of major importance for emotional and cognitive aspects of affective disorders. S100B-immunopositive astrocytes and oligodendrocytes were evaluated in the alveus and the CA1 pyramidal layer of patients with major depressive disorder (MDD) or bipolar I disorder (BD) compared to controls.As revealed by the optical disector cell-counting method, the numerical density of S100B-immunopositive astrocytes was bilaterally decreased in the CA1 pyramidal layer of MDD and BD patients compared to controls, whereas only the bipolar group showed a decreased density of S100B-immunopositive oligodendrocytes in the left alveus. These results were not confounded by gender, age, duration of disease, medication dosage, or autolysis time.Confirming the idea of previous S100B serum and cerebrospinal fluid studies, our data suggest that S100B-immunopositive glia is dysregulated in the brains of depressed patients. These findings are in accordance with animal experiments in rodents showing a reduced astrocytic S100B-immunoreactivity in the hippocampus after pharmacological serotonin depletion (modeling depression).

S100B-immunopositive astrocytes and oligodendrocytes in the hippocampus are differentially afflicted in unipolar and bipolar depression: A postmortem study

BackgroundPrevious studies have suggested that affective disorders are characterized by glial pathology. In this context, it has been hypothesized that elevated S100B serum and cerebrospinal fluid levels may represent a suitable surrogate marker. However, brain studies on the cellular distribution pattern of S100B in depressed patients are lacking so far. Such analyses are crucial, since S100B has been detected in various other cell types, even outside the central nervous system. MethodsTherefore, we performed a first postmortem analysis on this topic in the hippocampus – which is of major importance for emotional and cognitive aspects of affective disorders. S100B-immunopositive astrocytes and oligodendrocytes were evaluated in the alveus and the CA1 pyramidal layer of patients with major depressive disorder (MDD) or bipolar I disorder (BD) compared to controls. ResultsAs revealed by the optical disector cell-counting method, the numerical density of S100B-immunopositive astrocytes was bilaterally decreased in the CA1 pyramidal layer of MDD and BD patients compared to controls, whereas only the bipolar group showed a decreased density of S100B-immunopositive oligodendrocytes in the left alveus. These results were not confounded by gender, age, duration of disease, medication dosage, or autolysis time. ConclusionsConfirming the idea of previous S100B serum and cerebrospinal fluid studies, our data suggest that S100B-immunopositive glia is dysregulated in the brains of depressed patients. These findings are in accordance with animal experiments in rodents showing a reduced astrocytic S100B-immunoreactivity in the hippocampus after pharmacological serotonin depletion (modeling depression).