Characterization of glial fibrillary acidic protein and astroglial architecture in the brain of a continuously growing fish, the rainbow trout.

Unlike mammals, some fish, including carp and trout, have a continuously growing brain. The glial architecture of teleost brain has been intensively studied in the carp and few data exist on trout brain. In this study, using immunoblotting we characterized the topographic distribution of glial fibrillary acidic protein (GFAP) in larval and adult rainbow trout brain and studied by immunohistochemistry the distribution and morphology of GFAP-immunoreactive cell systems in the rainbow trout hindbrain and spinal cord. Immunoblotting yielded a double band with an apparent molecular weight of 50-52 kDa in the spinal cord homogenate in the trout larval and adult stages. In the adult hindbrain and forebrain, our antibody cross reacted also with a second band at a higher molecular weight (90 kDa). Because the forebrain contained this band alone the two brain regions might contain two distinct isoforms. Conversely, the larval total brain homogenate contained the heavy 90 kDa band alone. Hence the heavy band might be a GFAP protein dimer or vimentin/GFAP copolymer reflecting nerve fiber growth and elongation, or the two isoforms might indicate two distinct astroglial cell types as recently proposed in the zebrafish. In sections from trout hindbrain and spinal cord the antibody detected a GFAP-immunoreactive glial fiber system observed in the raphe and in the glial septa separating the nerve tracts. These radial glia fibers thickened toward the pial surface, where they formed glial end feet. The antibody also labeled perivascular glia around blood vessels in the white matter, and the ependymoglial plexus surrounding the ventricular surface in the grey matter. Last, it labeled round astrocytes. The GFAP-immunoreactive glial systems had similar distribution patterns in the adult and larval spinal cord suggesting early differentiation.

ATP regulates oligodendrocyte progenitor migration, proliferation, and differentiation: involvement of metabotropic P2 receptors.

Extracellular nucleotides act as potent signaling molecules in the neuron-glia and glia-glia communication, via the activation of specific ligand-gated P2X and G-protein-coupled metabotropic P2Y receptors. Most of the data available about the effects of P2 receptor activation in the CNS concern astrocytes, microglia, and neurons. To gain insights into the role of purinergic receptors in oligodendrocyte development, we characterized the expression and functional activity of P2 receptors in rat oligodendrocyte progenitors (OPs) and investigated the effects of ATP and its breakdown products on their functions. We describe here that rat OPs express different types of P2 receptors and that nucleotide-induced Ca(2+) raises in these progenitor cells are mainly due to the activation of P2X(7) ionotropic and ADP-sensitive P2Y(1) metabotropic receptors. We also show that ATP and ADP stimulate OP migration, inhibit the mitogenic response of OPs to PDGF and promote oligodendrocyte differentiation. The pharmacological profile of the nucleotide-induced effects demonstrates the important regulatory role of P2Y(1) receptor signaling in OP functions. These findings suggest that ATP, which is released in high amounts under inflammatory conditions and following cell death, might regulate remyelination processes in inflammatory demyelinating diseases of the CNS, like multiple sclerosis.

Metabotropic P2 receptor activation regulates oligodendrocyte progenitor migration and development.

To gain insights into the role of purinergic receptors in oligodendrocyte development, we characterized the expression and functional activity of P2 receptors in cultured rat oligodendrocyte progenitors and investigated the effects of ATP and its breakdown products on the migration and proliferation of this immature glial cell population. Using Western blot analysis, we show that oligodendrocyte progenitors express several P2X (P2X(1,2,3,4,7)) and P2Y (P2Y(1,2,4)) receptors. Intracellular Ca(2+) recording by Fura-2 video imaging allowed to determine the rank potency order of the P2 agonists tested: ADPbetaS = ADP = Benzoyl ATP > ATP > ATPgammaS > UTP, alpha,beta-meATP ineffective. Based on the above findings, on pharmacological inhibition by the antagonists oxATP and MRS2179, and on the absence of alpha,betameATP-induced inward current in whole-cell recording, P2X(7) and P2Y(1) were identified as the main ionotropic and metabotropic P2 receptors active in OPs. As a functional correlate of these findings, we show that ATP and, among metabotropic agonists, ADP and the P2Y(1)-specific agonist ADPbetaS, but not UTP, induce oligodendrocyte progenitor migration. Moreover, ATP and ADP inhibited the proliferation of oligodendrocyte progenitors induced by platelet-derived growth factor, both in purified cultures and in cerebellar tissue slices. The effects of ATP and ADP on cell migration and proliferation were prevented by the P2Y(1) antagonist MRS2179. By confocal laser scanning microscopy, P2Y(1) receptors were localized in NG2-labeled oligodendrocyte progenitors in the developing rat brain. These data indicate that ATP and ADP may regulate oligodendrocyte progenitor functions by a mechanism that involves mainly activation of P2Y(1) receptors.