Microglial Cell Population Research Articles

Enzyme histochemical identification of microglial cells in the retina of a fish ( Tinca tinca)

Histochemistry for nucleoside diphosphatase was used to study the microglial cells in the adult tench retina. An abundant population of microglial cells was located in the vascular membrane, nerve fibre layer, inner and outer plexiform layers and scattered cells were observed in the inner nuclear layer. Rounded and amoeboid cells could be seen close to the vessel in the vascular membrane, bipolar cells in the nerve fibre layer and ramified cells in the rest of the layers. Several microglial forms could correspond to developing cells. The pattern of distribution was similar to that described in other vertebrates, but with several differences, such as the presence of microglial cells in the vascular membrane and inner nuclear layer and the overlap of processes in the plexiform layers.

Enriched immune-environment of blood–brain barrier deficient areas of normal adult rats

The circumventricular organs (CVOs) in the brain are without a blood–brain barrier (BBB) and as such directly exposed to blood plasma constituents and blood-borne pathogens. In light of previous studies showing discrepancies regarding the immunocompetence of these organs, we initiated the present study to provide a comprehensive immunohistochemical analysis of the cellular expression of immune-associated antigens within the pineal gland, area postrema and the subfornical organ. In all CVOs, subpopulations of cells morphologically similar to complement receptor type 3 immunoreactive microglial/macrophage cells expressed major histocompatibility complex (MHC) class II antigen, leucocyte common antigen (LCA/CD45), as well as CD4 and ED1 antigen. Based on morphological criteria the MHC class II antigen expressing cells could be grouped into a major population of classical parenchymal and perivascular ramified microglial cells and a minor population presenting itself as scattered or small groups of rounded macrophage-like cells. CD4 and ED1 antigen were expressed by both cell types. CD45 was preferentially expressed by macrophage-like cells. MHC class I antigen was expressed by the vascular endothelium in both BBB-protected and BBB-deficient areas and was additionally present as a lattice-like network throughout the BBB-deficient parenchyma in all CVOs. The results suggest that the BBB-free areas of the brain besides being constantly surveyed by blood-borne macrophages, possess an intrinsic immune surveillance system based on resting and activated microglial cells, which may function as a non-endothelial, cellular barrier against blood-borne pathogens.

Exposure of Cultured Microglial Cells to Interferon-gamma

The reactions of a microglial cell population to exposure to interferon-gamma (IFN-γ) were studied in primary cultures of mixed glial cells from the brains of newborn rats. One week after seeding, the cells were exposed to 200U/ml of IFN-γ for 24 and 48 hours. After exposure to IFN-γ for 48 hours, a third of the cultures were exposed to phorbol myristate acetate (PMA) for 90 minutes. Transformation of the microglial cells in response to this treatment was observed by using a light microscope. On time-lapse film, an increase in the mobility of the cells and an increase in the number of amoeba-like microglia were among the most evident changes induced by IFN-γ. These changes became more pronounced when the exposure time was increased. The addition of PMA significantly enhanced this process. After exposure to IFN-γ for 48 hours, expression of the MHC class II antigen was observed. The interleukin-1β content of the cells was increased in the cultures after 24 hours, but declined after further exposure.

Transitory disappearance of microglia during the regeneration of the lizard medial cortex

In normal lizards, microglial cells populate the medial cortex (a zone homologous to the hippocampal fascia dentata), with a preferential distribution along the border between the granular cell layer and the plexiform layers. Intraperitoneal injection of the neurotoxin 3-acetylpyridine (3AP) induces a selective lesion in the medial cortex with a rapid degeneration of the granular layer and its zinc-enriched axonal projection. Within 6-8 weeks, the granular layer is, however, repopulated by a new set of neurons generated in the subjacent ependyma and the cell debris is removed. The aim of this study was to determine to what extent microglia were involved in the scavenging processes during the regeneration process. To this end we studied the brains of regenerating lizards at different times after 3AP lesion, visualising microglial cells by the nucleoside diphosphatase (NDPase) histochemical reaction. Surprisingly, we found that stained microglial cells disappeared 6-8 hours after 3AP injection and remained absent until 10-15 days after injection. One month postlesion an increased population of microglial cells was found scattered throughout all plexiform layers of the cortex. Thorough examination of semithin and ultrathin sections confirmed the absence of microglia in the medial cortex of recent lesioned animals but the presence of an exuberant population after 1 month postlesion. In the tissue, phagocytotic scavenging was carried out by radial ependymocytes, not by microglia.

Microglia and the developing olfactory bulb

The developmental appearance of microglia in the rat olfactory bulb was investigated through the use of selective staining with the B 4-isolectin from Griffonia simplicifolia. No changes in the density or distribution of either the spherical, macrophage “ameboid” form or the highly arborized “ramified” variety of microglia were observed in the superficial layers of the bulb between postnatal days 10 and 30. The subependymal zone exhibited the only substantial population of ameboid cells and the only developmental increases in ramified cell density during this time-period. External single naris closure, which enhances cell death in the ipsilateral bulb, did not affect microglia density, presumably due to the unusually high numbers of microglia normally present in the bulb. The olfactory bulb has a dense and relatively uniform population of microglial cells from very early stages of postnatal life, perhaps because of the constant turnover of cells and processes.

Heterogeneity of microglial and perivascular cell populations: Insights gained from the facial nucleus paradigm

We reflect here on the development of a neuroimmunological concept which has been formulated over the past 5 years through studying microglial cell responses in the facial nerve system. A simple axotomy of the adult rat facial nerve which causes regeneration of facial motor neurons and little, if any, cell death can activate microglial cells just as easily as a full-blown degeneration of the entire nucleus induced by toxic ricin. In both instances, the prompt microglial reaction is characterized by a series of structural and phenotypic changes which are in many ways similar to an immune response, e.g., there is cell proliferation and upregulation of MHC antigens. However, since white blood cells do not participate in the retrograde response of facial motor neurons, we have adopted a notion which views microglia as a CNS-wide network of immunocompetent cells whose morphological dissimilarities from leukocytes are a result of their unique adaptation to the CNS architecture. We have continued our in vivo investigations of the phagocytic and immunophenotypic properties of microglial and perivascular cells during the retrograde reaction of facial motor neurons by using intra-neural injections of fluorogold (FG) and ricin followed by lectin and immunostaining for microglia. Two new findings can be added to the microglial neuroimmune network: (1) Microglia take up FG only after motor neuron degeneration, whereas perivascular cells may take up FG under nondegenerating conditions. (2) Immunologically important molecules, such as MHC class II, CD4, and leukocyte common antigens, are expressed by different microglial subpopulations. Thus there is functional and phenotypic heterogeneity among immunocompetent cells of the CNS.

Identification and distribution of microglial cells in the cerebral cortex of the lizard: A histochemical study

The histochemical demonstration of nucleoside diphosphatase as a specific microglial marker was used to study the distribution of this glial cell type in the cerebral cortex of Podarcis muralis and Podarcis hispanica. Our results showed that in both species, NDPase staining was specific for the microglial cell population and that microglial cells displayed a specific localization pattern in the different cortical areas. In the medal cortex, microglial cells were principally found in the outer and inner plexiform layers in the strata adjacent to the granular layer. Moreover, some microglial cells were found near the ependymal layer, but no microglial cells were normally present near the brain surface and never in the deep inner plexiform layer. In the dorsomedial cortex, microglial cells were found near the brain surface in the outer plexiform layer, in the upper part of the granular layer, and near the ependymal layer. No microglial cells were found, however, in the outer and inner plexiform layers adjacent to the granular layer. Finally, in the dorsolateral cortex, microglial cells were located in the upper part of the outer plexiform layer, in and bordering the granular layer, and scattered in the inner plexiform layer. This layered-pattern distribution of microglial cell population in the cerebral cortex of the lizard differs from the apparently homogeneous distribution of microglia in the brain of mammals.

Microglia in the neurohypophysis associate with and endocytose terminal portions of neurosecretory neurons

The rat neurohypophysis contains a population of microglial cells, the majority of which occupy a pericapillary position in the resting gland. The microglia are immunocytochemically identifiable by the presence of macrophage-associated antigens and resemble microglia of the CNS. Morphometry at light and electron microscopic levels reveals that such cells constitute approximately 19% of the intrinsic cell population, excluding the endothelial cells. Two other populations of neurohypophysial glial cells, parenchymatous pituicytes and fibrous pituicytes, do not express macrophage-associated antigens. The microglia have long processes which surround and, in some cases, engulf apparently viable portions of the magnocellular neurosecretory nerve terminals. A sequence of stages of selective endocytosis and degradation of the engulfed nerve terminals can be visualized within pericapillary microglia. Some phagosomes and secondary lysosomes contain morphologically intact neurosecretory granules; others contain partially destroyed neurosecretory granules or amorphous material all of which are identifiable as originating from the magnocellular neurosecretory terminals by their immunoreactivity for oxytocinor vasopressin-neurophysin. This finding indicates a novel role for the microglial cells in remodelling terminal arborizations of neurosecretory neurons and in processing or degrading hormones and peptides they contain. Because of their close and selective associations with other cellular elements of the neurohypophysis, any substances produced by microglia also have the potential to influence hormone secretion, pituicyte proliferation and neurohypophysial vasculature.

Changes in microglial cell numbers in the spinal cord dorsal horn following brachial plexus transection in the adult rat.

The effect of peripheral nerve transection on the size of the microglial cell population in cytoarchitecturally distinct regions of the spinal cord dorsal horn of rats was evaluated at selected intervals 2 through 35 days after unilateral brachial plexotomy. The identification of cells was verified by electron microscopic examination of a representative random sample of cells included in the counts. Microglial cell numbers were increased in laminae I, II as well as the arbitrarily defined deeper laminae 3.5 days after surgery. Although microglial cell numbers in laminae I were within normal range 35 days after axotomy, those of the more ventrally located laminae remained significantly greater than control values for the duration of the experimental period. These findings demonstrate that: 1) microglial cell proliferation in the dorsal horn is an early event in the central changes that are attendant to peripheral nerve injury 2) the time course of the response varies in cytoarchitecturally different regions.

Early postnatal development of glial cells in the canine cervical spinal cord.

To study qualitative and quantitative changes in the glial cell population of young postnatal dogs, the cervical spinal cords of 20 beagle pups, ranging in age from 1 to 28 days, were prepared for light and electron microscopy. Glial cells in the lateral corticospinal tract were classified and quantified directly on the electron microscope. Quantification was performed by means of a stereological method designed to correct for sampling bias, and glia were classified according to morphological criteria as immature glial cell precursors, light and dark oligodendrocytes, astrocytes, and microglia. Glial cell precursors, which include undifferentiated glioblasts, oligodendroblasts, and astroblasts, predominated in the first few days after birth, constituting 43% of the glial cell population, and then declined to less than 5% by 28 days. Light and dark oligodendrocytes differed morphologically in their electron density and the appearance of their organelles. Light oligodendrocytes increased slightly prior to myelination, and then declined, whereas dark oligodendrocytes continued to increase throughout the 4-week period and became the predominant cell type at 28 days (66%). In contrast to the oligodendroglial population, the sizes of the astroglial and microglial cell populations were relatively stable. This study shows that the population of immature glial cell precursors, abundant at birth in the lateral corticospinal tract, appear to be differentiating primarily into oligodendroglia, because this population exhibits a rapid increase in size, and relatively little change occurs in the astrocyte population. The trends in glial cell development in the dog are similar to those reported for rodents, although there may be some variation in the maturation and activity of oligodendrocytes.

Microglia in the hypendyma of the rat subcommissural organ following brain lesion with serotonin neurotoxin.

The population of microglial cells in the subependymal layer of the subcommissural organ is sparse in normal adult rats. The number of microglial cells was substantially increased in this area following intraventricular injection of the serotonin neurotoxin 5,6-dihydroxytryptamine (5,6-DHT). In sections of plastic embedded material, 1 micron thick, the majority of phagocytic cells scattered in the subependymal layer had an appearance similar to that described in classical studies of microglial cells. At the electron microscopic level microglial cells exhibited the characteristic elongate nucleus with peripheral chromatin condensation. The perikaryon was scanty, containing strands of rough endoplasmic reticulum. The abundant organelles in the processes included Golgi complexes, mitochondria, rough and smooth endoplasmic reticulum as well as dense and multivesicular bodies. In addition, the processes contained phagocytosed axon terminals originating from the dense serotoninergic input to the subcommissural organ, which had degenerated on accumulating the serotonin neurotoxin. A fraction of the phagocytosed material was contained in subependymal subcommissural organ cells, astrocytes and oligodendrocytes. At the light microscopic level the phagocytosed terminals were visualized histochemically with Schmorl's reaction, which resulted in Prussian Blue precipitates. This allowed screening of microglial cells in complete series of sections through the well-defined subependymal layer of the subcommissural organ.

A morphological study of glial cells in the hypoglossal nucleus of the cat during nerve regeneration

The cat hypoglossal nerve and nucleus have been used as a model for the study of the occurrence and time course of modifications in the size and composition of the perineuronal glial cell population as they relate to cytological changes in the nerve cell body and the initiation and progress of axon regeneration. Animals were killed at 2, 5, 10, 20, 35, 65, and 115 days after crush injury to the hypoglossal nerve. At 5 days after surgery, growth cones and regenerating unmyelinated axons were present at the lesion site, but no conspicuous changes were apparent in the nerve cell bodies. At 10 days after surgery, the granular endoplasmic reticulum was disaggregated and depleted. The elongation phase appeared to be completed at 20 days, as judged by the bilateral retrograde labeling of the hypoglossal nuclei with horseradish peroxidase. By 35 days, the cytoarchitecture of the nerve cell bodies and maturation of axons, as determined by a comparison of the relative frequency distribution of cross sectional areas proximal and distal to the lesion, were completely restored. Comparative quantitative light microscopic examination of the hypoglossal nuclei of intact and experimental animals failed to reveal any statistically significant differences in the total number of glial cells, number of glial cells/unit area of neuropil, or relative proportions of glial cell types at any of the postoperative time intervals. Moreover, electron microscopic quantitation of the microglial cell population did not reveal any significant alterations in the number, density, location, or morphology of these cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Endocytic activity of subependymal microglial cells in the toad brain: a cytochemical study of peroxidase uptake.

A population of microglial cells that rapidly incorporate extracellular material introduced into the ventricular system has been identified just beneath the ependyma of all four cerebral ventricles in the toad (Bufo marinus). In untreated tissue these cells appear to be scattered, possess few processes and have an elongate shape with their long axes lying parallel to the ventricular surface. Their most distinctive ultrastructural features are nuclei containing clumps of chromatin, cytoplasmic dense bodies and single strands of granular endoplasmic reticulum. When horseradish peroxidase (HRP) is perfused through the ventricular system and the tissue processed using the DAB cytochemical method, the cells change shape and incorporate HRP into cytoplasmic structures. Even after very short perfusion periods (2-5 minutes) cells become rounded, the surface is ruffled and pseudopodia develop that contain characteristic flocculent material. Reaction product for HRP is contained in plain and coated vesicles, tubules, vacuoles and long structures composed of two closely apposed membranes. At these early times, relatively few multivesicular bodies and dense bodies contain reaction product, but when the cells are viewed at longer time periods after the ventricular perfusion of HRP an increasing proportion of the multivesicular bodies and dense bodies contain reaction product. By 320 minutes reaction product is found almost exclusively in these two organelles. In addition, many pseudopodia containing dense bodies with peroxidase activity are found in the neurophile; some, but not all, can be traced from the subependymal microglial cells. The cell bodies have resumed their flattened shape. When compared to the subependymal microglial cells, other brain cells--oligodendrocytes, astrocytes, ependymal cells and neurons--contain relatively little reaction product at short time intervals; only by 320 minutes are moderate amounts of HRP present. Because of the position of the microglial cells and their ingestive capacity, it is suggested that they function to protect the brain from foreign substances entering from the CSF.

Neuroglial cells in the cerebral cortex of rats from young adulthood to old age: an electron microscope study.

Neuroglial cells were examined in the rat auditory cerebral cortex to determine the effects of aging on astrocytes, oligodendrocytes and microglia. The animals ranged in age from 3 months to 29.5 months. Over this age range of 26.5 months none of the neuroglial cells exhibit striking abnormalities in the morphology or the number of their organelles other than each of these cell types accumulates a distinct membrane-bound inclusion material. Astrocytes accumulate considerably more of this material than oligodendrocytes, and demonstrate earlier and more frequent signs of it. Microglia show the most striking alterations in regard to the inclusion material and in animals of increasing age microglial cells engorged with the heterogeneous material become increasingly common. Concurrently there is a slow transformation of the microglial population from an array of multipolar forms to larger, elongated or spherical forms which contain variable amounts of the dense membrane-bound material. These alterations in the microglial cell population do not occur at a chronologically uniform rate, and in animals 18 months and older, examples of the multipolar, the elongated, and the engorged spherical forms of microglia may be encountered. The total number of neuroglial cells increases with age. However, the numbers of the various types do not increase by similar proportions. While there is little change in the populations of astrocytes and oligodendrocytes in rats between the ages of 3 and 27 months, in the same animals the number of microglia increases by about 65%.