A. C. H. Yu

Astrocytic response to injury

Publisher Summary The chapter discusses the role of the astrocyte in the central nervous system (CNS) injury and disease. Astrocytes comprise as much as 25% of the cells and 35% of the total mass of the CNS. Astrocytes form barriers around blood vessels and connections between nerve cells. Numerous functions have been assigned to the astrocyte depending on its stage of maturation, location in the CNS, and response to CNS insult. Some signals that regulate gene expression in development and response to astrocyte injury are: growth factors, prion protein from Scrapies, neural and immunological adhesion molecules, such as NCAM, LFA-1, gangliosides, low density lipoproteins, cytokines from T-cells, macrophages and other glia, neurotransmitters and neuropeptides, such as catecholamines, monoamines, glutamate, ATP, substance P, and antigen-antibody complexes. Astrocytic responses to these signals include: (1) proliferation, movement and differentiation; (2) changes in shape, cell volume, cytoskeletal organization, endocytic activity, lysosomal fragility, and enzyme content; (3) buffering capacity for K + , glutamate and GABA; (4) expression of nerve growth factor, tumor necrosis factor, interferon α and β , interleukin 1 and 6, colony stimulating factor-1, fibroblast growth factor, neurotropic factors, neurite promoting agents, MHC class I and I1 histocompatibility antigens, amyloid protein, GD3 ganglioside, ICAM- 1, Na + channel protein, GFAP, crystallin, vimentin and heat shock proteins.

Chapter 17: Regulatory role of astrocytes for neuronal biosynthesis and homeostasis of glutamate and GABA

Publisher Summary The chapter discusses an overview of the current status of the knowledge about transmembrane fluxes of glutamate, glutamine and GABA, and their apparent metabolic interconversions. The glutamine synthesizing enzyme glutamine synthetase (GS) is localized in astrocytes and not in neurons. As a consequence of this, there exists a cycling of glutamate, glutamine, and presumably GABA between neurons and astrocytes, and this cycling is defined as the glutamate/glutamine cycle. Regardless of whether the precursor for transmitter glutamate is glutamine or a tricarboxylic acid constituent, a communication between neurons and astrocytes must take place as the astrocytes control the availability of these metabolites. Transmitter GABA is synthesized from glutamate by the action of glutamate decarboxylase, which is found only in GABAergic neurons. However, it appears that in the brain in vivo in synaptosomes, as well as in cultured GABAergic neurons glutamine is a better precursor for GABA than glutamate.

Gene expression in astrocytes during and after ischemia.

Involvement of the IEGs in brain injury and ischemia is under intensive investigation (Gubits et al., 1993). There are several families of the IEGs. They include the fos, jun, and zinc finger genes that encode transcription factors. Products of the fos family (c-fos, fra-1, fra-2, and fos B) bind to members of the jun family (c-jun, jun B, jun D) via leucine zippers, and this dimer then binds to the AP-1 site (consensus sequence -TGACTCA-) in the promoter of target genes, which in turn regulate the expression of late response genes that produce long-term changes in cells. For example, c-fos may regulate the long-term expression of preproenkephalin, nerve growth factor, dynorphin, vasoactive intestinal polypeptide, tyrosine hydroxylase and other genes with AP-1 sites in their promoters (Curran and Morgan, 1987; Sheng and Greenberg, 1990). It is likely that the c-fos gene up-regulation observed in ischemic astrocytes leads to the changes observed in the expressions of hsp and cytoskeleton protein genes in this experimental model. This is supported by the findings of Sarid (1991) and Pennypacker et al. (1994) who have shown that AP-1 DNA binding activity in hippocampus recognized an AP-1 sequence from the promoter region of the GFAP which is a potential target gene. van de Klundert et al. (1992) also suggested the involvement of AP-1 in transcriptional regulation of vimentin. IEGs can be induced within minutes by extracellular stimuli including transmitters, peptides, and growth factors. In this study, we have shown that c-fos induction by ischemia was rapid and transient.(ABSTRACT TRUNCATED AT 250 WORDS)

Tumor Necrosis Factor‐α and Basic Fibroblast Growth Factor Decrease Glial Fibrillary Acidic Protein and Its Encoding mRNA in Astrocyte Cultures and Glioblastoma Cells

Abstract: Tumor necrosis factor‐α is a pluripotent cytokine that is reportedly mitogenic to astrocytes. We examined expression of the astrocyte intermediate filament component glial fibrillary acidic protein in astrocyte cultures and the U373 glioblastoma cell line after treatment with tumor necrosis factor‐α. Treatment with tumor necrosis factor‐α for 72 h resulted in a decrease in content of glial fibrillary acidic protein and its encoding mRNA. At the same time, tumor necrosis factor‐α treatment increased the expression of the cytokine interleukin‐6 by astrocytes. The decrease in glial fibrillary acidic protein expression was greater when cells were subconfluent than when they were confluent. Thymidine uptake studies demonstrated that U373 cells proliferated in response to tumor necrosis factor‐α, but primary neonatal astrocytes did not. However, in both U373 cells and primary astrocytes tumor necrosis factor‐α induced an increase in total cellular protein content. Treatment of astrocytes and U373 cells for 72 h with the mitogenic cytokine basic fibroblast growth factor also induced a decrease in glial fibrillary acidic protein content and an increase in total protein level, demonstrating that this effect is not specific for tumor necrosis factor‐α. The decrease in content of glial fibrillary acidic protein detected after tumor necrosis factor‐α treatment is most likely due to dilution by other proteins that are synthesized rapidly in response to cytokine stimulation.

Chapter 20 A RT-PCR study of gene expression in a mechanical injury model

Publisher Summary This chapter discusses the reverse transcriptase polymerase chain reaction (RT-PCR) method to study cytokine expression in astrocytes in response to exogenous agents and mechanical injury. By quantitative RT-PCR, it has been recently shown that TNFα/ interleukin (IL)-1β induced IL-6 in mouse results in astrocyte cultures. IL-6 belongs to a family of neuroactive cytokines including leukemia inhibitor factor (LIF) and ciliary neurotrophic factor (CNTF). RT-PCR has been used to demonstrate that the cholinergic differentiation factor/leukemia inhibitory factor (CDFLIF) and CNTF induce mRNAs for choline acetyltransferase, somatostatin, substance P, vasoactive intestinal peptide, cholecystokinin, and enkephalin in cultured sympathetic neurons. These data suggest that CDFLIF and CNTF may share receptor subunits and signal transduction pathways. Lipopolysaccharide, PMA, tumor necrosis factor (TNFα), and IL-lβ have also been found to induce LIF in cultured mouse astrocytes but not in the immortalized microglial cell line (BV-2) provided by Bocchini.

Inhibition of GFAP Synthesis with Antisense Nucleic Acid Constructs

Glial fibrillary acidic protein (GFAP), the major component of the intermediate filament in differentiated astrocytes (Eng et al., 1971; Eng, 1985), is extensively synthesized within and adjacent to the site of injury (Eng, 1988a; Condorelli et al., 1990; Hozumi et al., 1990; Vijayan et al., 1990). Other than GFAP accumulation, astrogliosis is also characterized by astrocyte proliferation (hyperplasia) and extensive hypertrophy of the cell body, nucleus as well as cytoplasmic processes (Eng, 1988a). Astrogliosis may participate in the healing phase following CNS injury by actively monitoring and controlling the molecular and ionic contents of the extracellular space of the CNS. They can wall off areas of the CNS that are exposed to non-CNS tissue environments following trauma. On the other hand, such responses may interfere with the function of residual neuronal circuits, by preventing remyelination, or by inhibiting axonal regeneration (Eng et al., 1987; Stensaas et al., 1987; Reier and Houle, 1988). Although astrogliosis has received considerable attention in term of its proposed inhibitory effect on CNS repair, there is still very little specific information available concerning the properties of reactive astrocytes, what triggers glial reactivity, and many of the cellular dynamics associated with scar formation. Control of astrocyte proliferation, differentiation, and astrogliosis may be linked to GFAP synthesis. Our aim was to transfect astrocytes with exogenous synthetic oligo- or polynucleotides, which would allow the manipulation of a transient suppression of GFAP synthesis which might delay the gliotic reaction and the scar formation, thus allowing neurons and oligodendrocytes to re-establish a functional environment.