Franco Ciani

Identification of surface glycoconjugates in the olfactory system of turtle

Lectin binding histochemistry was performed on the olfactory system of Pseudemys scripta to investigate the distribution and density of defined carbohydrate terminals on the cell surface glycoproteins of the olfactory receptors and their terminals in the olfactory bulbs. The lectin staining patterns indicate that the receptor cells of the olfactory mucosa are characterized by glycoconjugates containing alpha-D-galactose and N-acetyl-D-glucosamine terminal residues. The vomeronasal receptor cells contain instead alpha-N-acetyl-D-galactosamine, N-acetyl-D-glucosamine and alpha-D-galactose residues. The results demonstrate that the vomeronasal receptor cells contain high density of alpha-N-acetyl-D-galactosamine sugar residues that are not expressed by receptor cells of the olfactory mucosa. The presence of specific glycoproteins, whose terminal sugars are detected by lectin binding, might be related to the chemoreception and transduction of the odorous message into a nervous signal or in the histogenesis of the olfactory system. In fact, the olfactory receptors are the only known neurons in the vertebrate nervous system that undergo a continual cycle of proliferation not only in developing animals but also in mature ones. Moreover the results show that BSA-I-B4, an alpha-D-galactosyl-specific isolectin, targets the terminal sugar residues in the ramified microglial cells.

Histochemical study by lectin binding of surface glycoconjugates in the developing olfactory system of rat

Lectin‐binding histochemistry was used to investigate the distribution and density of defined carbohydrate sequences on the cell surface glycoproteins of the olfactory receptors of rat during development. The olfactory and vomeronasal receptors showed a positive labelling after biotinylated Lycopersicum esculentum lectin binding on embryonic day 16 (E16), while horseradish peroxidase‐labelled Glycine max, Bandeiraea simplicifolia (BS A‐I) and its B4 isomer BSA‐I‐B4 agglutinins started to label from day 18 (E18). From this stage onward there was a progressive increase in the intensity and number of lectin‐binding olfactory receptors. The first lectin‐labelled bundles of axons penetrating the olfactory bulb were observed on E20; from E21 it was possible to identify the first labelled glomeruli that, on the first day (P1) of postnatal life, showed a feature very similar to that of the adult. The lectin staining patterns indicate that during development there are differences in the kind and distribution of saccharidic moieties on the surface of rat olfactory neurons. The possible role of carbohydrate‐containing glycoproteins in the reception and transduction of the odours and in the modulation of the cell‐cell interactions in the olfactory system is discussed.

Ultrastructural localization of alkaline phosphatase in cultures of nervous tissuein vitro

SummaryThe fine localization of alkaline phosphatase in culturesin vitro of chick embryo spinal cord, explanted at early stages of development, was studied. Enzyme localization was found to vary in the different areas of the culture. In the central region, the reaction product was present only on the membranes of non-nervous cells and fibres directly in contact with the culture medium, while in the migration area the reaction product was also localized on membranes of deeper cells and fibres and on all membranes at the periphery of the culture. However, the membranes of nerve cells and their processes showed no evidence of reaction product when they were in direct contact with the medium. These observations are compared with data on the distribution of alkaline phosphatase in nervous tissue during embryonic development and in adults, and in relation to the possible role of alkaline phosphatase at the level of the haematoencephalic barrier. The hypothesis is considered that selective trophic mechanisms characteristic of nervous tissuein vivo may also occur in cultures.

Enzymic activities in dissociated neurons differentiated in cultures in vitro.

Abstract— The appearance of the enzymes succinate dehydrogenase, lactate dehydrogenase, acid phosphatase, monoamine oxidase and acetylcholinesterase in cultures of dissociated spinal cord, dorsal root ganglia and cerebellum removed at different stages of development have been studied. The neurons differentiated in vitro have morphological and cytochemical characteristics similar to those of the normally developing neurons. On the basis of the results obtained, it is argued that in this experimental model the histogenetic determination of the neuroblasts at the time of the explant may start and bring to an end the morphological and biochemical differentiation of the various types of neurons even under conditions extremely different from those of normal development.

Lectin cytochemical localisation of glycoconjugates in the olfactory system of the lizards Lacerta viridis and Podarcis sicula

To investigate the presence of defined carbohydrate moieties on the cell surface of the olfactory and vomeronasal receptor cells and the projections of the latter into the olfactory bulbs, a lectin binding study was performed on the olfactory system of the lizards: Lacerta viridis and Podarcis sicula. Both lizards showed a high lectin binding for N-acetyl-glucosamine in the sensory neurons. The lectin binding patterns in Lacerta indicated that the main olfactory system possessed a moderate density of N-acetyl-galactosamine residues and detectable levels of galactose ones. The vomeronasal system on the other hand contained a high density of N-acetyl-galactosamine moieties and a moderate density of glucosamine ones. In Podarcis the main olfactory system and vomeronasal organ contained respectively detectable and moderate levels of galactose residues. The expression of specific glycoconjugates may be associated with outgrowth, guidance and fasciculation of olfactory and vomeronasal axons.

Immunohistochemical and Histochemical Characteristics of the Olfactory System of the Guppy, Poecilia reticulata (Teleostei, Poecilidae)

Olfaction in fish has been studied using preferentially macrosmatic species as models. In the present research, the labelling patterns of different neuronal markers and lectins were analyzed in the olfactory neurons and in their bulbar axonal endings in the guppy Poecilia reticulata, belonging to the group of microsmatic fish. We observed that calretinin immunostaining was confined to a population of olfactory receptor cells localized in the upper layers of the sensory mucosa, probably microvillous neurons innervating the lateral glomerular layer. Immunoreactivity for S100 proteins was mainly evident in crypt cells, but also in other olfactory cells belonging to subtypes projecting in distinct regions of the bulbs. Protein gene product 9.5 (PGP 9.5) was not detected in the olfactory system of the guppy. Lectin binding revealed the presence of N‐acetylglucosamine and α‐N‐acetylgalactosamine residues in the glycoconjugates of numerous olfactory neurons ubiquitously distributed in the mucosa. The low number of sugar types detected suggested a reduced glycosidic variability that could be an index of restricted odorant discrimination, in concordance with guppy visual‐based behaviors. Finally, we counted few crypt cells which were immunoreactive for S100 and calretinin. Crypt cells were more abundant in guppy females. This difference is in accordance with guppy gender‐specific responses to pheromones. Cells immunoreactive to calretinin showed no evidence of ventral projections in the bulbs. We assumed the hypothesis that their odorant sensitivity is not strictly limited to pheromones or sexual signals in general. Anat Rec, 2009.

Lectin-Binding Patterns in the Olfactory System of the Lizard, Physignathus lesueurii

Lectin binding histochemistry was performed on the olfactory system of Physignathus lesueurii to investigate the distribution and density of defined carbohydrate terminals on the cell‐surface glycoproteins of the olfactory and vomeronasal receptor cells and their terminals in the olfactory bulbs. The lectin staining patterns indicate that the vomeronasal and olfactory receptor cells are characterized by glycoconjugates containing α‐D‐galactose and N‐acetyl‐D‐glucosamine terminal residues. The presence of specific glycoproteins, whose terminal sugars are detected by lectin binding, might be related to the chemoreception and transduction of the odorous message into a nervous signal or to the histogenesis and development of the olfactory system. The olfactory and vomeronasal receptor cells are vertebrate neurons that undergo a continual cycle of proliferation not only during development but also in mature animals. J. Morphol. 247:34–38, 2001.

Ultrastructural analysis on acetylcholinesterase localization in the cerebellar cortex of teleosts

SummaryThe histochemical localization of acetylcholinesterase (AChE) was studied by electron microscopy in the cerebellar cortex of the goldfish and the catfish. The patterns of enzyme distribution show noticeable differences in the two teleost species at the level of the corresponding cerebellar structures. Among the most distinctive features in the prevailing intracellular localization of enzyme activity in the goldfish and the prevailing extracellular localization in the catfish in the molecular layer and, to a lesser extent, the granular layer. Only quantitative differences in the ability to synthesize AChE can be recorded among the different cerebellar neurons in the two species, since all these neurons exhibit different amounts of enzyme activity linked to their cytoplasmic structures.Comparing the results obtained with those of previous histochemical, experimental and developmental researches, the hypothesis seems well founded that the embryonic pool of cerebellar neurons is made up to AChE-synthesizing neuroblasts which, during development, lose or maintain to a different degree the mechanisms for AChE synthesis. In addition the light and electron microscope histochemistry reveals at different levels of resolution that the final pattern of AChE distribution in the cerebellar cortex is the sum of different degrees of AChE synthesis by cerebellar neurons and different degrees of enzyme release in extracellular spaces.

Ultrastructural pattern of acetylcholinesterase distribution in the cerebellar cortex of the quail

SummaryThe ultrastructural localization of acetylcholinesterase (AChE) was studied in the cerebellar cortex of the quail by means of histochemical methods. The greater amount of AChE was detected at level of the molecular layer in the intercellular spaces between parallel fibers and between parallel fibers and dendritic terminals. Many neurons showed intracellular localization of enzyme activity: the AChE positive neurons were all Golgi cells, most stellate and basket cells and different aliquots of Purkinje and granule cells. The enzymatic activity was usually localized in the cisternae of endoplasmic reticulum, in the nuclear envelope (but this last localization was not present in Purkinje cells) and sometimes in the Golgi apparatus; reaction granules were usually scarce in the different dendritic branches ramifying in the molecular layer. On the basis of the ultrastructural pattern of AChE distribution, some considerations are developed on the methodological aspects concerning the reliability of histochemical methods, the differences recorded at light and electron microscope level, the problems related to extracellular localization of enzyme, the difficulty of establishing a precise correlation between AChE localization in a cerebellar neuron and its possible cholinergic and/or cholonoceptive nature.

Acetylcholinesterase activity in the normal and retino-deprived optic tectum of the quail

SummaryAcetylcholinesterase localization has been studied by electron microscopic histochemistry in the quail optic tectum. Ultrastructural analysis reveals that the different neuronal types in the tectum possess the metabolic pathways for AChE synthesis to different degrees. From the site of synthesis in cell bodies the enzyme spreads towards areas of neuropil. In the neuropil of AChE-rich areas a balance seems to exist between enzyme stored in dendrites (and sometimes axon terminals) and enzyme released into the extracellular spaces. Precise identification of cholinergic synapses by means of AChE localization is in most cases impossible, due to extensive spread of the enzyme through the extracellular compartments of the neuropil.Unilateral ocular ablation causes disappearance of the stratum opticum and decrease in thickness of the superficial tectal layers in the contralateral optic tectum, but only minor modifications in AChE localization. This finding is in agreement with biochemical results which show equivalence of the relative concentration of AChE in the right and left optic tectum 1 or 2 months after ablation of the right eye. The experimental evidence suggests that cholinergic mechanisms are not related to the discharge of retinal afferents on receptive tectal neurons, but more likely to intrinsic neural circuits which might be involved in the modulation of tectal activity.