Benzodiazepine Interference with Fertility and Embryo Development: A Preliminary Survey in the Sea Urchin Paracentrotus lividus.

Psychotropic drugs and benzodiazepines are nowadays among the primary substances of abuse. This results in a large and constant release into aquatic environments where they have potentially harmful effects on non-target organisms and, eventually, human health. In the last decades, evidence has been collected on the possible interference of benzodiazepines with reproductive processes, but data are few and incomplete. In this study, the possible negative influence of delorazepam on fertilization and embryo development has been tested in Paracentrotus lividus, a key model organism in studies of reproduction and embryonic development. Sperm, eggs, or fertilized eggs have been exposed to delorazepam at three concentrations: 1 µg/L (environmentally realistic), 5 µg/L, and 10 µg/L. Results indicate that delorazepam reduces the fertilizing capacity of male and female gametes and interferes with fertilization and embryo development. Exposure causes anatomical anomalies in plutei, accelerates/delays development, and alters the presence and distribution of glycoconjugates such as N-Acetyl-glucosamine, α-linked fucose, and α-linked mannose in both morulae and plutei. These results should attract attention to the reproductive fitness of aquatic species exposed to benzodiazepines and pave the way for further investigation of the effects they may exert on human fertility. The presence of benzodiazepines in the aquatic environment raises concerns about the reproductive well-being of aquatic species. Additionally, it prompts worries regarding potential impacts on human fertility due to the excessive use of anxiolytics.

Histopathological effects of long-term exposure to realistic concentrations of cadmium in the hepatopancreas of Sparus aurata juveniles.

In recent decades, cadmium has emerged as an environmental stressor in aquatic ecosystems due to its persistence and toxicity. It can enter water bodies from various natural and anthropogenic sources and, once introduced into aquatic systems, can accumulate in sediments and biota, leading to bioaccumulation and biomagnification in the food chain. For this reason, the effects of cadmium on aquatic life remain an area of ongoing research and concern. In this paper, a multidisciplinary approach was used to assess the effects of long-term exposure to an environmental concentration on the hepatopancreas of farmed juveniles of sea bream, Sparus aurata. After determining metal uptake, metallothionein production was assessed to gain insight into the organism's defence response. The effects were also assessed by histological and ultrastructural analyses. The results indicate that cadmium accumulates in the hepatopancreas at significant concentrations, inducing structural and functional damage. Despite the parallel increase in metallothioneins, fibrosis, alterations in carbohydrate distribution and endocrine disruption were also observed. These effects would decrease animal fitness although it did not translate into high mortality or reduced growth. This could depend on the fact that the animals were farmed, protected from the pressure deriving from having to search for food or escape from predators. Not to be underestimated is the return to humans, as this species is edible. Understanding the behaviour of cadmium in aquatic systems, its effects at different trophic levels and the potential risks to human health from the consumption of contaminated seafood would therefore be essential for informed environmental management and policy decisions.

Benzodiazepine Delorazepam Induces Locomotory Hyperactivity and Alterations in Pedal Mucus Texture in the Freshwater Gastropod Planorbarius corneus.

Benzodiazepines, psychotropic drugs, are ubiquitous in the aquatic environment due to over-consumption and inefficient removal by sewage treatment plants. Bioaccumulation with consequent behavioral and physiological effects has been reported in many aquatic species. However, the responses are species-specific and still poorly understood. To improve the knowledge, we exposed the freshwater snail Planorbarius corneus to 1, 5, or 10 µg/L of delorazepam, the most widely consumed benzodiazepine in Italy. Conventional behavioral tests were used to assess the effects on locomotor and feeding behavior. Histological and biochemical analyses were also performed to detect possible changes in the structure and composition of the foot mucus and glands. The results show a paradoxical response with reduced feeding activity and locomotor hyperactivity. Pedal mucus was altered in texture but not in composition, becoming particularly rich in fibrous collagen-like material, and a significant change in the protein composition was highlighted in the foot. In conclusion, exposure to delorazepam induces disinhibited behavior in Planorbarius corneus, potentially increasing the risk of predation, and an increase in mucus protein production, which, together with reduced feeding activity, would severely compromise energy resources.

Adverse Effect of Metallic Gold and Silver Nanoparticles on Xenopus laevis Embryogenesis.

Exposure to metal nanoparticles is potentially harmful, particularly when occurring during embryogenesis. In this study, we tested the effects of commercial AuNPs and AgNPs, widely used in many fields for their features, on the early development of Xenopus laevis, an anuran amphibian key model species in toxicity testing. Through the Frog Embryo Teratogenesis Assay-Xenopus test (FETAX), we ascertained that both nanoparticles did not influence the survival rate but induced morphological anomalies like modifications of head and branchial arch cartilages, depigmentation of the dorsal area, damage to the intestinal brush border, and heart rate alteration. The expression of genes involved in the early pathways of embryo development was also modified. This study suggests that both types of nanoparticles are toxic though nonlethal, thus indicating that their use requires attention and further study to better clarify their activity in animals and, more importantly, in humans.

Abnormal outer hair cell efferent innervation in Hoxb1-dependent sensorineural hearing loss.

Autosomal recessive mutation of HOXB1 and Hoxb1 causes sensorineural hearing loss in patients and mice, respectively, characterized by the presence of higher auditory thresholds; however, the origin of the defects along the auditory pathway is still unknown. In this study, we assessed whether the abnormal auditory threshold and malformation of the sensory auditory cells, the outer hair cells, described in Hoxb1null mutants depend on the absence of efferent motor innervation, or alternatively, is due to altered sensory auditory components. By using a whole series of conditional mutant mice, which inactivate Hoxb1 in either rhombomere 4-derived sensory cochlear neurons or efferent motor neurons, we found that the hearing phenotype is mainly reproduced when efferent motor neurons are specifically affected. Our data strongly suggest that the interactions between olivocochlear motor neurons and outer hair cells during a critical postnatal period are crucial for both hair cell survival and the establishment of the cochlear amplification of sound.

Hepatocyte Aquaporins AQP8 and AQP9 Are Engaged in the Hepatic Lipid and Glucose Metabolism Modulating the Inflammatory and Redox State in Milk-Supplemented Rats.

Milk is an important source of nutrients and energy, but there are still many uncertainties regarding the health effects of milk and dairy products consumption. Milk from different species varies in physicochemical and nutritional properties. We previously showed that dietary supplements with different milks in rats trigger significant differences in metabolic and inflammatory states, modulating mitochondrial functions in metabolically active organs such as the liver and skeletal muscle. Here, we have deepened the effects of isoenergetic supplementation of milk (82 kJ) from cow (CM), donkey (DM) or human (HM) on hepatic metabolism to understand the interlink between mitochondrial metabolic flexibility, lipid storage and redox state and to highlight the possible role of two hepatocyte aquaporins (AQPs) of metabolic relevance, AQP8 and AQP9, in this crosstalk. Compared with rats with no milk supplementation, DM- and HM-fed rats had reduced hepatic lipid content with enhanced mitochondrial function and decreased oxidative stress. A marked reduction in AQP8, a hydrogen peroxide channel, was seen in the liver mitochondria of DM-fed rats compared with HM-fed, CM-fed and control animals. DM-fed or HM-fed rats also showed reduced hepatic inflammatory markers and less collagen and Kupffer cells. CM-fed rats showed higher hepatic fat content and increased AQP9 and glycerol permeability. A role of liver AQP8 and AQP9 is suggested in the different metabolic profiles resulting from milk supplementation.

Structural and functional damage to the retina and skeletal muscle in Xenopus laevis embryos exposed to the commonly used psychotropic benzodiazepine delorazepam.

Benzodiazepines, psychotropic drugs, are among the most frequently found pharmaceuticals in aquatic matrices. An increasing number of studies are reporting their harmful effects on adults' behaviour and physiology, while little information is available regarding developing organisms exposed since early stages. Improper activation of GABA receptors during embryonic development is likely to induce relevant consequences on the morphogenesis and, at later stages, on behaviour. This study investigated the negative effects of three increasing concentrations of delorazepam on Xenopus laevis retinal and skeletal muscle morphogenesis. Morphological and ultrastructural investigations were correlated with gene expression, while Raman spectroscopy highlighted the main biochemical components affected. Conventional phototactic response and orientation in the magnetic field were assessed as indicators of proper interaction between sensory organs and the nervous system. Results confirm the profound impact of delorazepam on development and return an alarming picture of the amphibians' survival potentialities in a benzodiazepine-contaminated environment.

Water contamination by delorazepam induces epigenetic defects in the embryos of the clawed frog Xenopus laevis.

Delorazepam, a derivative of diazepam, is a psychotropic drug belonging to the benzodiazepine class. Used as a nervous-system inhibitor, it treats anxiety, insomnia, and epilepsy, but is also associated with misuse and abuse. Nowadays benzodiazepines are considered emerging pollutants: conventional wastewater treatment plants indeed are unable to eliminate these compounds. Consequently, they persist in the environment and bioaccumulate in non-target aquatic organisms with consequences still not fully clear. To collect more information, we investigated the possible epigenetic activity of delorazepam, at three concentrations (1, 5 and 10 µg/L) using Xenopus laevis embryos as a model. Analyses demonstrated a significant increase in genomic DNA methylation and differential methylation of the promoters of some early developmental genes (otx2, sox3, sox9, pax6, rax1, foxf1, and myod1). Moreover, studies on gene expression highlighted an unbalancing in apoptosis/proliferation pathways and an aberrant expression of DNA-repair genes. Results are alarming considering the growing trend of benzodiazepine concentrations in superficial waters, especially after the peak occurred as a consequence of the pandemic COVID-19, and the fact that benzodiazepine GABA-A receptors are highly conserved and present in all aquatic organisms.

Behavioral alterations and gills damage in Mytilus galloprovincialis exposed to an environmental concentration of delorazepam.

Psychoactive compounds, and benzodiazepines (BZPs) in particular, represent an important class of emerging pollutants due to their large (ab)use and high resistance to degradation. Nowadays it is known that sewage treatment does not completely eliminate these substances and, therefore, BZPs and their metabolites reach concern levels in most aquatic environments all over Europe, ranging from µg/L to ng/L. In this study, we investigated the effects of delorazepam on Mytilus galloprovincialis, a model organism in toxicity testing and a key species in coastal marine ecosystems. Given its psychoactive activity, the study primarily addressed discovering the effects on behavior, by conventional valve opening and closure tests. Possible cytotoxic activity was also investigated by analyzing valve abductor muscles, gills histology, and correlated oxygen consumption. Results demonstrate negative effects on mussel behavior, interference with metabolism, and alteration of gill morphology and protein content. In conclusion, delorazepam confirms its toxicity to aquatic environments, highlighting the possibility that BZDs can ultimately affect the structure of the food web and the functions of the coastal ecosystems.

Salicylate attenuates gentamicin-induced ototoxicity and facilitates the recovery in the basilar papilla of the lizard Podarcis siculus.

It is known that ototoxicity is the main cause of toxicity induced by aminoglycoside antibiotics. Effects on cochlea and vestibule in vertebrates are variable, depending on the typology of the aminoglycoside and the animal model examined. Despite this, they are routinely used to prevent postoperative and urinary tract infections and in the treatment of tuberculosis and cystic fibrosis. Gentamicin causes hearing loss by damaging stereocilia and by causing degeneration of hair cells due to free radical formation and eventual activation of caspase-dependent pathways. Its toxicity increases with the frequency of administration, dose concentration, and duration of treatment. Turnover of new hair cells may occur spontaneously, throughout life, or may be triggered by an acoustic or ototoxic insult to replace dead cells. Turnover and repair of damage are common in fish and amphibians and in birds' vestibule. In contrast, in the papilla basilaris of birds, and in the vestibule of mammals, hair cell regeneration is activated only after damage. Sensory epithelium repair and hair cell regeneration also occur in the reptiles' vestibule, but no data is available on regeneration or repair in the basilar papilla, involved in sound perception. The purpose of this work is therefore to assess the damage induced by gentamicin on the papilla basilaris of a reptile model organism, the Lacertidae Podarcis siculus. Recovery was also evaluated 3, 8 and 18 days after the end of exposure, in absence of gentamicin and in presence of the otoprotective salicylate. Scanning electron microscopy (SEM) was carried out to check for morphological damage while the occurrence of cell proliferation events was evaluated by fluorescence microscopy, after administration of 5-Bromo-2'-deoxyuridine (BrdU). Results show that salicylate administration facilitates recovery and reduces damage to hair cells after gentamicin treatment. Following the incorporation of bromodeoxyuridine, we demonstrated that sensory epithelium repair and hair cell regeneration have occurred, and that the recovery is due to either proliferation of the supporting cells and/or self-repair of hair cell bundles in the weakly damaged sensory cells.

The Hepatic Mitochondrial Alterations Exacerbate Meta-Inflammation in Autism Spectrum Disorders.

The role of the liver in autism spectrum disorders (ASD), developmental disabilities characterized by impairments in social interactions and repetitive behavioral patterns, has been poorly investigated. In ASD, it has been shown a dysregulation of gut-brain crosstalk, a communication system able to influence metabolic homeostasis, as well as brain development, mood and cognitive functions. The liver, with its key role in inflammatory and metabolic states, represents the crucial metabolic organ in this crosstalk. Indeed, through the portal vein, the liver receives not only nutrients but also numerous factors derived from the gut and visceral adipose tissue, which modulate metabolism and hepatic mitochondrial functions. Here, we investigated, in an animal model of ASD (BTBR mice), the involvement of hepatic mitochondria in the regulation of inflammatory state and liver damage. We observed increased inflammation and oxidative stress linked to hepatic mitochondrial dysfunction, steatotic hepatocytes, and marked mitochondrial fission in BTBR mice. Our preliminary study provides a better understanding of the pathophysiology of ASD and could open the way to identifying hepatic mitochondria as targets for innovative therapeutic strategies for the disease.

Environmental concentrations of a delorazepam-based drug impact on embryonic development of non-target Xenopus laevis.

Benzodiazepines, psychotropics drugs used for treating sleep disorders, anxiety and epilepsy, represent a major class of emerging water pollutants. As occurs for other pharmaceutical residues, they are not efficiently degraded during sewage treatment and persist in effluent waters. Bioaccumulation is already reported in fish and small crustaceans, but the impact and consequences on other "non-target" aquatic species are still unclear and nowadays of great interest. In this study, we investigated the effects of a pharmaceutical preparation containing the benzodiazepine delorazepam on the embryogenesis of Xenopus laevis, amphibian model species, taxa at high risk of exposure to water contaminants. Environmental (1 µg/L) and two higher (5 and 10 µg/L) concentrations were tested on tadpoles up to stage 45/46. Results demonstrate that delorazepam interferes with embryo development and that the effects are prevalently dose-dependent. Delorazepam reduces vitality by decreasing heart rate and motility, induces marked cephalic and abdominal edema, as well as intestinal and retinal defects. At the molecular level, delorazepam increases ROS production, modifies the expression of some master developmental genes and pro-inflammatory cytokines. The resulting stress condition significantly affects embryos' development and threatens their survival. Similar effects should be expected as well in embryos belonging to other aquatic species that have not been yet considered targets for these pharmaceutical residues.

Effects of Cadmium Exposure on Gut Villi in Danio rerio.

In aquatic organisms, cadmium exposure occurs from ovum to death and the route of absorption is particularly wide, being represented by skin, gills and gastrointestinal tract, through which contaminated water and/or preys are ingested. It is known that cadmium interferes with the gut; however, less information is available on cadmium effects on an important component of the gut, namely goblet cells, specialized in mucus synthesis. In the present work, we studied the effects of two sublethal cadmium concentrations on the gut mucosa of Danio rerio. Particular attention was paid to changes in the distribution of glycan residues, and in metallothionein expression in intestinal cells. The results show that cadmium interferes with gut mucosa and goblet cells features. The effects are dose- and site-dependent, the anterior gut being more markedly affected than the midgut. Cadmium modifies the presence and/or distribution of glycans in the brush border and cytoplasm of enterocytes and in the goblet cells' cytoplasm and alters the metallothionein expression and localization. The results suggest a significant interference of cadmium with mucosal efficiency, representing a health risk for the organism in direct contact with contamination and indirectly for the trophic chain.

Ketogal Safety Profile in Human Primary Colonic Epithelial Cells and in Mice.

In our previous studies, a ketorolac-galactose conjugate (ketogal) showed prolonged anti-inflammatory and analgesic activity, causing less gastric ulcerogenic effect and renal toxicity than its parent drug ketorolac. In order to demonstrate the safer profile of ketogal compared to ketorolac, histopathological changes in the small intestine and liver using three staining techniques before and after repeated oral administration in mice with ketorolac or an equimolecular dose of its galactosylated prodrug ketogal were assessed. Cytotoxicity and oxidative stress parameters were evaluated and compared in ketorolac- and ketogal-treated Human Primary Colonic Epithelial cells at different concentrations and incubation times. Evidence of mitochondrial oxidative stress was found after ketorolac treatment; this was attributable to altered mitochondrial membrane depolarization and oxidative stress parameters. No mitochondrial damage was observed after ketogal treatment. In ketorolac-treated mice, severe subepithelial vacuolation and erosion with inflammatory infiltrates and edematous area in the intestinal tissues were noted, as well as alterations in sinusoidal spaces and hepatocytes with foamy cytoplasm. In contrast, treatment with ketogal provided a significant improvement in the morphology of both organs. The prodrug clearly demonstrated a safer profile than its parent drug both in vitro and ex vivo, confirming that ketogal is a strategic alternative to ketorolac.

Dietary Micronutrient Management to Treat Mitochondrial Dysfunction in Diet-Induced Obese Mice.

Obesity and associated metabolic disturbances, which have been increasing worldwide in recent years, are the consequences of unhealthy diets and physical inactivity and are the main factors underlying non-communicable diseases (NCD). These diseases are now responsible for about three out of five deaths worldwide, and it has been shown that they depend on mitochondrial dysfunction, systemic inflammation and oxidative stress. It was also demonstrated that several nutritional components modulating these processes are able to influence metabolic homeostasis and, consequently, to prevent or delay the onset of NCD. An interesting combination of nutraceutical substances, named DMG-gold, has been shown to promote metabolic and physical wellness. The aim of this research was to investigate the metabolic, inflammatory and oxidative pathways modulated by DMG-gold in an animal model with diet-induced obesity. Our data indicate that DMG-gold decreases the metabolic efficiency and inflammatory state and acts as an antioxidant and detoxifying agent, modulating mitochondrial functions. Therefore, DMG-gold is a promising candidate in the prevention/treatment of NCD.

Transient Anomalous Diffusion MRI in Excised Mouse Spinal Cord: Comparison Among Different Diffusion Metrics and Validation With Histology.

Neural tissue is a hierarchical multiscale system with intracellular and extracellular diffusion compartments at different length scales. The normal diffusion of bulk water in tissues is not able to detect the specific features of a complex system, providing nonlocal, diffusion measurement averaged on a 10-20 µm length scale. Being able to probe tissues with sub-micrometric diffusion length and quantify new local parameters, transient anomalous diffusion (tAD) would dramatically increase the diagnostic potential of diffusion MRI (DMRI) in detecting collective and sub-micro architectural changes of human tissues due to pathological damage. In DMRI, the use of tAD parameters quantified using specific DMRI acquisition protocols and their interpretation has often aroused skepticism. Although the derived formulas may accurately fit experimental diffusion-weighted data, the relationships between the postulated dynamical feature and the underlying geometrical structure remains elusive, or at most only suggestive. This work aimed to elucidate and validate the image contrast and information that can be obtained using the tAD model in white matter (WM) through a direct comparison between different diffusion metrics and histology. Towards this goal, we compared tAD metrics extracted from pure subdiffusion (α-imaging) and super-pseudodiffusion (γ-imaging) in excised mouse spinal cord WM, together with T2 and T2* relaxometry, conventional (normal diffusion-based) diffusion tensor imaging (DTI) and q-space imaging (QSI), with morphologic measures obtained by optical microscopy, to determine which structural and topological characteristics of myelinated axons influenced tAD contrast. Axon diameter (AxDiam), the standard deviation of diameters (SD ax.diam ), axonal density (AxDens) and effective local density (ELD) were extracted from optical images in several WM tracts. Among all the diffusion parameters obtained at 9.4 T, γ-metrics confirmed a strong dependence on magnetic in-homogeneities quantified by R2* = 1/T2* and showed the strongest associations with AxDiam and ELD. On the other hand, α-metrics showed strong associations with SD ax.diam and was significantly related to AxDens, suggesting its ability to quantify local heterogeneity degree in neural tissue. These results elucidate the biophysical mechanism underpinning tAD parameters and show the clinical potential of tAD-imaging, considering that both physiologic and pathologic neurodegeneration translate into alterations of WM morphometry and topology.

Effects of four food dyes on development of three model species, Cucumis sativus, Artemia salina and Danio rerio: Assessment of potential risk for the environment.

Food dyes, or color additives, are chemicals added to industrial food products and in domestic cooking to improve the perceived flavor and attractiveness. Of natural and synthetic origin, their safety has been long discussed, and concern for human safety is now clearly manifested by warnings added on products labels. Limited attention, however, has been dedicated to the effects of these compounds on aquatic flora and fauna. For this reason, the toxicity of four different commercially available food dyes (cochineal red E120, Ponceau red E124, tartrazine yellow E102 and blue Patent E131) was assessed on three different model organisms, namely Cucumis sativus, Artemia salina and Danio rerio that occupy diverse positions in the trophic pyramid. The evidence collected indicates that food dyes may target several organs and functions, depending on the species. C. sativus rate of germination was increased by E102, while root/shoot ratio was ∼20% reduced by E102, E120 and E124, seed total chlorophylls and carotenoids were 15-20% increased by E120 and 131, and total antioxidant activity was ∼25% reduced by all dyes. Mortality and low mobility of A. salina nauplii were increased by up to 50% in presence of E124, E102 and E131, while the nauplii phototactic response was significantly altered by E102, E120 and E124. Two to four-fold increases in the hatching percentages at 48 h were induced by E124, E102 and E131 on D. rerio, associated with the occurrence of 20% of embryos showing developmental defects. These results demonstrated that the food dyes examined are far from being safe for the aquatic organisms as well as land organisms exposed during watering with contaminated water. The overall information obtained gives a realistic snapshot of the potential pollution risk exerted by food dyes and of the different organism' ability to overcome the stress induced by contamination.

Gross anatomy and ultrastructure of Moorish Gecko, Tarentola mauritanica skin.

The epidermis of Tarentola mauritanica in the skin regions of back, flank and belly has been described using light and electron microscopy. This animal model was useful to give an insight of the functional pattern involved in pigmentation, cryptism and photosensitivity. Skin from back and flanks, in electron microscopy, shows a high concentration of chromatophores, among those melanophores, xanthophores and iridophores have been reported. Interestingly, in the flank-back transition region electron microscopy reveals the presence of nerve endings. Our contribution adds new knowledge about the skin of this species, and it could be useful to study in deep the mechanism of cryptic colour change in reptiles.

Ketogal: A Derivative Ketorolac Molecule with Minor Ulcerogenic and Renal Toxicity.

Ketorolac is a powerful non-steroidal anti-inflammatory drug (NSAID), with a great analgesic activity, present on the Italian market since 1991. Despite the excellent therapeutic activity, the chronic use of ketorolac has long been limited owing to the high incidence of gastrointestinal and kidney side events. In our previous study, we demonstrated that ketorolac-galactose conjugate (ketogal), synthesized and tested in a single-dose study, was able to reduce ulcerogenicity, while preserving the high pharmacological efficacy of its parent drug. In this paper, in order to verify the suitability of this compound, for repeated administration, ex vivo experiments on naïve mice were performed. Mice were treated for 5 or 7 days with the highest doses of two drugs (ketorolac 10 mg/kg and ketogal 16.3 mg/kg), and the expression of both gastric COX-1 and PGsyn was evaluated. Results showed that oral ketorolac treatment significantly reduced both enzymes; surprisingly, oral treatment with ketogal did not produce significant variation in the expression of the two constitutive enzymes. Moreover, histological experiments on stomach and kidneys clearly indicated that repeated administration of ketogal induced lower toxicity than ketorolac. At same time, in vivo results clearly showed that both ketorolac and ketogal had a similar therapeutic activity in a model of inflammation and in pain perception. These effects were accompanied by the reduction of enzyme expression such as COX-2 and iNOS, and by the modulation of levels of nuclear NF-κB and cytosolic IκB-α in the inflamed paws. These very encouraging results demonstrate for the first time that ketogal could represent a valid and novel therapeutic alternative to the ketorolac and might pave the way for clinical studies.

Long term exposure to cadmium: Pathological effects on kidney tubules cells in Sparus aurata juveniles.

The effects of an exposure to cadmium chloride 0.47µM for 150days were studied in kidneys of juveniles Sparus aurata by a multidisciplinary approach so to correlate uptake and detoxification potential to changes in brush border and glycocalyx sugar composition. Results demonstrated that cadmium concentration in kidney significantly increased from day 30 reaching a plateau on day 120 while metallothioneins reached a peak on day 90 and by day 120 were already decreasing to control values. Cytological damage was extensive on day 90, clearly detectable at both structural and ultrastructural levels, in tubular cells and brush-border. Staining with a panel of four lectins revealed a significant increase in N-Ac-Gal and a decrease in mannose in the glycocalyx and the tubular basal membranes. From day 120, when cadmium concentration was high and metallothionein concentration decreasing, a clear recovery was observed in tubular cells morphology and sugar composition. Possible significance of these apparently contrasting data are discussed.