Glial cells of the lamprey nervous system contain keratin‐like proteins

Abstract

Lamprey axons regenerate following spinal cord transection despite the formation of a glial scar. As we were unable to detect a lamprey homologue of glial fibrillary acidic protein (GFAP), a major constituent of astrocytes, we studied the composition of intermediate filament (IF) proteins of lamprey glia. Monoclonal antibodies (mAbs) were raised to lamprey spinal cord cytoskeletal extracts and these mAbs were characterized by using Western blotting and immunocytochemistry. On two-dimensional (2-D) Western blots, five of the mAbs detected three major IF polypeptides in the molecular weight (MW) range of 45-56 kD. Further studies were conducted to determine the relationship between the lamprey glial-specific antigen and other mammalian IF proteins. Antikeratin 8 antibody recognized two of the three polypeptides. Several of the glial-specific mAbs reacted with human keratins 8 and 18 on Western blots. Keratin-like immunoreactivity was found in all parts of the central and peripheral nervous systems in both larval and adult lampreys. The immunocytochemical staining patterns of glial-specific mAbs were indistinguishable on lamprey spinal cord sections. However, on brain sections, two distinct patterns were observed. A subset of mAbs stained only a few glial fibers in the brain, whereas others stained many more brain glia, particularly the ependymal cells. The former group of mAbs recognized only the two lower MW polypeptides on 2-D Western blots, but the latter group of mAbs recognized all three major IF polypeptides. This correlation is supported by the observation that the highest MW IF polypeptide has an increased level of expression in the brain relative to the spinal cord. Thus, in the lamprey, the glial cells of both spinal cord and brain express molecules similar to simple epithelial cytokeratins, but their IFs may contain these keratins in different stoichiometric proportions. The widespread presence in the lamprey of primitive glial cells containing keratin-like intermediate filaments may have significance for the extraordinary ability of lamprey spinal axons to regenerate.

D‐serine is distributed in neurons in the brain of the sea lamprey

“…The ependyma was mostly free of D-serine immunoreactivity, and no positive cells of the same size and appearance of lamprey astrocytes (see Merrick et al, 1995) were observed. The distribution of D-serine-ir cells in the different brain regions of adult lamprey brain is described first, and, where appropriate, the colocalization of D-serine with GABA immunoreactivity in neurons is also indicated.…”

“…Moreover, expression of glycine and GABA immunoreactivity in lamprey brain perikarya is largely maintained from larvae to adulthood (Villar-Cerviño et al, 2009), indicating that lampreys are highly useful for investigating the neuronal distribution of amino acid neurotransmitters in the brain. With regard to the glia, lampreys lack oligodendrocytes, and glial cells of the brain are ependymal tanycytes or small astrocytes (Merrick et al, 1995), which may greatly facilitate the identification of cells containing D-serine.…”

Lamprey neurofilaments contain a previously unreported 50‐kDa protein

“…To identify candidates for this missing protein, we surveyed 43 mAbs designated as LCM series, which were generated previously against cytoskeletal proteins of the lamprey nervous system by Merrick et al (1995) in this laboratory. Seven LCMs were neuron specific, and five were glial specific, as demonstrated by immunohistochemistry (described later).…”

“…Therefore, its electrophoretic labeling pattern was compared with those of LCM3 and LCM39 on Western blots, and all of these were compared with the electrophoretic pattern of LCM1, a mAb previously determined to be specific for lamprey glial keratins (Merrick et al, 1995) (Fig. 2).…”

“…LCM1 recognized three bands in Western blots with sizes ranging between 40 and 55 kDa. These proteins were previously characterized as glial keratins (Merrick et al, 1995).…”