Cristiano Angelini

Biological effects of a neurotoxic pesticide at low concentrations on sea urchin early development. a terathogenic assay

Dose-dependent terathogenic effects of an organophosphate insecticide were found during early sea urchin development. This biological assay is low cost, easy to measure, and allows to detect the effects of the exposure of organisms to the active principle at concentrations lower than the acceptable daily intake (ADI) for man. Effects were found independently from the stage of exposure, and were major as earlier exposure occurred. The stronger effects were exerted on the elongation of the skeletal rods, that was easily measured by following the migration of primary mesenchyme cells, labelled by WGA (wheat germ agglutinin).

CHOLINERGIC ACTIVATION OF SETTLEMENT IN CIONA INTESTINALIS METAMORPHOSING LARVAE

During metamorphosis, ascidian tadpoles settle on appropriate substrates, by secreting adhesive substances from the papillae. The papillae were shown by different authors to possess neural structures, probably for substrate recognition. In free-swimming tadpoles, AChE activity was localized by histochemical methods at the cell surface of the adhesive papillae, suggesting the possible presence of a cholinergic system in these structures. In vivo experiments showed that the treatment with AChCl caused dose-dependent enhancement of metamorphosis, shortening of the exploratory period of the larvae, and secretion within a few minutes of sConA-binding glycoconjugates in the tunica, beginning from the adhesive papillae.

Evidence for the presence of a mammalian-like cholinesterase in Paramecium Primaurelia (Protista, Ciliophora) developmental cycle

By histochemical and immunohistochemical methods, the presence of cholinergic-like molecules has previously been demonstrated in Paramecium primaurelia, and their functional role in mating-cell pairing was suggested. In this work, both true acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) activities were electrophoretically investigated, and the presence of molecules immunologically related to BuChE was checked by immunoblotting. The AChE activity, shown in the membrane protein fraction of mating-competent cells and in the cytoplasmic fraction of immature cells, is due to a 260-kDa molecular form, similar to the membrane-bound tetrameric form present in human erythrocytes. This AChE activity does not appear in either the cytoplasmic fraction of mating-competent cells or in the membrane protein fraction of immature cells. No evidence was found for the presence or the activity of BuChE-like molecules. The role of AChE in P. primaurelia developmental cycle is discussed.

Cell Signalling During Sea Urchin Development: A Model for Assessing Toxicity of Environmental Contaminants

The early development of sea urchins has been thoroughly studied since the beginning of the 20th century thanks to the particular features of the model involving cell signalling, making it easy to follow the complex cell-to-cell interactions that lead to development. In this chapter, the prominent role of cell-to-cell communication in developmental events is discussed, as well as the role of intracellular ion changes that are in turn regulated by signal molecules belonging to the cholinergic system. The results seem to indicate that the zygote stage is the most suitable to study the role of the cholinergic system, as at this stage, a calcium spike can be evoked by exposure to acetylcholine (ACh) or to muscarinic drugs, at any time before the nuclear breakdown. The described outcomes also open a path to a new way of considering biomarkers. In fact, most environmental factors have the capacity to interfere with the cholinergic system: stress, wounds, inflammation and pollution in general. In particular, this offers a way to investigate the presence in the environment and the degree of aggressiveness of neurotoxic contaminants, such as organophosphate and carbamate pesticides, largely used in European countries for many purposes, including agricultural pest control and medical treatment. These drugs exert their function by interfering with the regulation of the cholinergic system and the consequent electrical events. Thus, the sea urchin zygote could represent a reliable model to be used in biosensors with the capacity to translate the effect of neurotoxic pesticides, and generally of stress-inducing contaminants, in living cell responses, such as electrical responses.

Changes in the ultrastructure and glycoproteins of the cyst wall of Colpoda cucullus during resting encystment

Summary Our previous studies on long-term resting encystment of Colpoda inflata showed transcriptional activity of the macronucleus and the occurrence of protein synthesis. In this work we have investigated the structure, the glycoprotein composition, and the lectin-binding site localization of the cyst wall of C. cucullus at increasing cyst ages, by means of scanning and transmission electron microscopy, confocal microscopy, and cytochemical and lectin blotting techniques. During an 18-day resting encystment, the cyst wall undergoes structural modifications resulting in the compaction of its previously-formed layers and the production of new material that is placed on the inner side of the cyst wall. Changes in the localization of ConA-/WGA-binding sites have been checked by comparing 7-day-old cysts to 1-day-old ones. Moreover, differences in the electrophoretic and ConA-/WGA-positive patterns have been found in 18-day-old cysts when compared to 1-day-old ones. The results of in vivo ConA and WGA assays suggest that ConA promotes encystment and delays excystment.

Expression pattern of mUBPy in the brain and sensory organs of mouse during embryonic development

Mouse UBPy (mUBPy) belongs to the family of ubiquitin-specific processing proteases (UBPs). In this study we have investigated the expression of mUBPy in the brain and sensory organs of mouse at different embryonic stages (E9, E11, E13, E15, E17, E19) and during the postnatal stages P0, P1, P2, P4 and P5 using Western blot and immunohistochemistry. mUBPy-immunoreactive cell bodies first appeared at stage E11 in several brain regions, particularly in the walls surrounding the vesicles and the ventricles. Subsequently, at stage E13, new mUBPy-positive cells appeared in the corpus striatum, the caudate nucleus, the thalamus, the epithalamus, the hypothalamus and the pons. At E15 the mUBPy pattern was very similar to that observed at E13, whereas at stage E17 mUBPy-immunoreactivity significantly decreased and a high number of mUBPy-immunoreactive cells was found only to line the third ventricle and within the mantle layer of the fourth ventricle. At E19 and P0, no mUBPy-immunoreactive element was found in the brain. At the postnatal stages P2 and P5, mUBPy-positive cells were detected in all subdivisions of the brain, with high concentrations in several cortex regions. Double labeling with the mUBPy antiserum and antisera against specific cell markers showed that the enzyme is expressed both in neurons and astrocytes. Outside the brain, mUBPy was detected, from stage E11, in the eye, within the lens and the cornea, in the inner ear, at the level of the cochlear and vestibular systems and in the olfactory epithelium. The spatio-temporal expression of mUBPy suggests that the enzyme may be involved in neuroregulatory processes during embryogenesis.