Barbara Cuoghi

Macroglial cells of the teleost central nervous system: a survey of the main types

Following our previous review of teleost microglia, we focus here on the morphological and histochemical features of the three principal macroglia types in the teleost central nervous system (ependymal cells, astrocyte-like cells/radial glia and oligodendrocytes). This review is concerned with recent literature and not only provides insights into the various individual aspects of the different types of macroglial cells plus a comparison with mammalian glia, but also indicates the several potentials that the neural tissue of teleosts exhibits in neurobiological research. Indeed, some areas of the teleost brain are particularly suitable in terms of the establishment of a “simple” but complete research model (i.e. the visual pathway complex and the supramedullary neuron cluster in puffer fish). The relationships between neurons and glial cells are considered in fish, with the aim of providing an integrated picture of the complex ways in which neurons and glia communicate and collaborate in normal and injured neural tissues. The recent setting up of successful protocols for fish glia and mixed neuron-glia cultures, together with the molecular facilities offered by the knowledge of some teleost genomes, should allow consistent input towards the achievement of this aim.

Cytogenetic analysis of the pufferfish Tetraodon fluviatilis (Osteichthyes).

Because of their compact genome, pufferfish (Tetraodontiformes) have been proposed as a model for the study of the vertebrate genome. The genome of pufferfish is peculiar as it has the structural complexity of the genomes of higher vertebrates, but has small introns and lacks large clusters of highly repetitive sequences. Despite such interest, information about the genetics of pufferfish is still scanty. To fill this gap, we have performed a cytogenetic analysis of the pufferfish, Tetraodon fluviatilis, which can be maintained in an aquarium for a long time and, unlike the pufferfish, Fugu rubripes, it is not difficult to obtain. Karyotype analysis shows that T. fluviatilis has 2n = 42 with two metacentric chromosomes, four submetacentrics, two subtelocentrics and 34 acrocentrics. C-banding, followed by DAPI staining, showed that heterochromatin is essentially AT-rich and is located at centromeres. Staining of the same metaphase plates with CMA3 showed the presence of four heterochromatic regions located on two pairs of submetacentric chromosomes. Silver staining and FISH with a 28S rDNA probe showed that these GC-rich regions are nucleolar organizing regions (NORs). Finally, regardless of the technique used, no difference in the chromosome complement was found between males and females.

Endoreplication: a molecular trick during animal neuron evolution.

The occurrence of endoreplication has been repeatedly reported in many organisms, including protists, plants, worms, arthropods, molluscs, fishes, and mammals. As a general rule, cells possessing endoreplicated genomes are largesized and highly metabolically active. Endoreplication has not been frequently reported in neuronal cells that are typically considered to be fully differentiated and nondividing, and which normally contain a diploid genome. Despite this general statement, various papers indicate that giant neurons in molluscs, as well as supramedullary and hypothalamic magnocellular neurons in fishes, contain DNA amounts larger than 2C. In order to study this issue in greater detail here, we review the available data about endoreplication in invertebrate and vertebrate neurons, and discuss its possible functional significance. As a whole, endoreplication seems to be a sort of molecular trick used by neurons in response to the high functional demands that they experience during evolution.

Microglia of teleosts: facing a challenge in neurobiology

This review is concerned with recent literature on teleost fish CNS microglia. It covers not only various aspects of these cells, notably comparing them with mammalian microglia, but also points out the several potentialities neural tissue of teleosts exhibits in neurobiological research. The relationships between neurons and glial cells are considered in fish, aiming at an integrated picture of the complex ways neurons and glia communicate and collaborate in normal and injured neural tissues. In addition, attention has been paid to different teleost models according to their availability, easy maintenance in experimental conditions, possibilities of embryos manipulation and sequenced genome. The recent setting up of successful protocols for fish glia and mixed neuron-glia cultures, together with the molecular facilities offered from genome knowledge, should provide a new boost to studies about microglia and neuron-microglia relationships.

Histochemical and immunocytochemical localization of nitric oxide synthase in the supramedullary neurons of the pufferfish Tetraodon fluviatilis

The presence of the nitric oxide (NO) converting enzyme, constitutive neuronal NO synthase (nNOS), was investigated in the supramedullary neurons (SN) cluster of the pufferfish Tetraodon fluviatilis. The identification of NADPH diaphorase- (NADPHd-) positivity and the demonstration of nNOS with the BAS technique and with immunofluorescence together, strongly indicate the presence of a constitutive NO converting enzyme in SN cellular bodies and axons, and provides evidence that the SN cluster represents a distinct nitrergic neuronal system in the vertebrate CNS. The possible roles of NO in the cluster are discussed, including an involvement in communication among neurons and between neurons and the glial cells in the cluster.

The supramedullary neurons of fish: present status and goals for the future

In this paper, we report the recent findings on supramedullary neurons of fish, with special attention to the studies, which made the nature of this neuronal system clear. Indeed, immunohistochemical, physiological and neuroanatomical data, taken together, point out that this neuronal system is a component of the autonomic nervous system. New goals have been opened by the more recent research, especially in comparative neurobiology. Indeed, the supramedullary neurons, owing to some characteristics, like the DNA endoreplication, the large size, the accessible localization and the relationship with glial cells, may be utilised as a very suitable model in several fields of neurobiology of vertebrates, such as molecular genetic, electrophysiology, nervous system ageing, glial-neuron interactions.

Glial cells: basic components of clusters of supramedullary neurons in pufferfish.

In this paper a cytochemical and ultrastructural study of clustered supramedullary neurons (SN) of Tetraodon fluviatilis (Tetraodontiformes) is presented. SN are large-sized nerve cells that have a high metabolic rate and are intensely engaged in protein biosynthesis. The SN are completely surrounded by two types of glial cell, which have been ultrastructurally, histochemically and immunohistochemically identified as astrocyte-like cells and microglial cells. The glial cells are located very close to the SN and sometimes contact them, which suggests that they do not only provide mechanical support but are also trophic for the SN and serve their functioning. We consider these glial cells as a constitutive and functional part of the cluster of SN, and therefore propose that the SN cluster constitute a suitable model to study in detail the morphological and functional relationships between neurons and glial cells in Anamnia.

Localization of the (TTAGGG)n telomeric repeat in the chromosomes of the pufferfish Tetraodon fluviatilis (Hamilton Buchanan) (Osteichthyes)

Abstract In order to characterise the telomeric repeats of the pufferfish Tetraodon fluviatilis, fluorescent in situ hybridization (FISH) was carried out on metaphase chromosomes using a PCR generated probe (TTAGGG)n. Distinct signals have been observed on the tel-omeres of all the chromosomes and hybridization signals appears of uniform size and intensity. Moreover FISH experiment does not evidence any interstitial non-telomeric signal. As non-telomeric FISH signal has been used as a marker for karyotype evolution and for determining the evolutionary status of fish species, it could be supposed that the absence of non telomeric signals makes T. fluviatilis (Tetraodontiformes) an evolutionary ancient species in respect to Perciformes.

The supramedullary cells of the teleost Coris julis (L.): a noradrenergic neuronal system.

This study, carried out on Coris julis (Labridae), is a contribution to the immunocytochemical characterization of fish supramedullary neurons. The significance of these giant cells has been debated since the beginning of the twentieth century. Our research provides the first evidence for a noradrenergic feature of this neuronal system. The possible role of supramedullary neurons as components of the autonomic nervous system is discussed. Moreover, the present results, taken together with our previous studies, surmise that this the first known case of colocalization of a neuropeptide (gastrin/CCK-like) and noradrenaline in the nervous system of teleosts.

DNA Endoreplication in the Clustered Supramedullary Neurons of the Pufferfish Diodon Holacanthus L. (Osteichthyes)

DNA contents, ranging from 4C to more than 500C, were determined by a quantitative microfluorimetric assay in supramedullary neuron nuclei of the pufferfish Diodon holacanthus. The distribution of C values after ethidium bromide staining indicates an inter- and intra-individual variation in DNA contents which do not correspond to duplications of the total genome, suggesting that DNA replication is specific for particular genes (endoreplication). Moreover, the DNA content appears to be correlated with nuclear size. A relationship between the DNA amounts and the presence of AT- and GC-rich sequences has been shown.This work demonstrates, for the first time, DNA endoreplication in a specific neuronal type in vertebrates.