José Manuel Cifuentes

Pharmacokinetic and toxicological data of spirolides after oral and intraperitoneal administration.

Spirolides are a kind of marine toxins included in the cyclic imine toxin group and produced by the dinoflagellate Alexandrium ostenfeldii. This study shows for the first time a complete and detailed description about the symptoms observed in mice when these toxins were intraperitoneal (i.p.) administered. It is also compared the i.p. toxicity of 13-desmethyl spirolide C (13-desMeC), 13,19-didesMeC (13,19-didesMeC) and 20-methyl spirolide G (20-Me-G) in experiments performed with highly purified toxins. The bioassay indicates that 13-desMeC and 13,19-didesMeC are extremely toxic compounds which have a LD(50) of 27.9μg/kg and 32.2μg/kg, respectively. However, when 20-MeG was i.p administrated with dose up 63.5μg/kg, no deaths were recorded. In order to evaluate the oral toxicity, spirolides were administered by gastric intubation into mice. Then, samples of blood, urine and faeces were collected and analyzed by liquid chromatography-mass spectrometry tandem (LC-MS/MS) technique. Spirolides appear in blood at 15min and in urine after 1h of being toxin administered. In summary, in this paper, it is provided new data about the toxicity, absorption, and excretion of spirolides in mouse. So far, little information is available on this item but necessary for spirolide regulation in the European Union (EU).

A descriptive and comparative lectin histochemical study of the vomeronasal system in pigs and sheep.

The accessory olfactory bulb (AOB) is the primary target of the sensory epithelium of the vomeronasal organ (VNO), and thus constitutes a fundamental component of the accessory olfactory system, which is involved in responses to behaviour‐related olfactory stimuli. In this study we investigated the characteristics of the AOB, VNO, vomeronasal nerves (VNNs) and caudal nasal nerve (CdNN) in pigs and sheep, species in which olfaction plays a key behavioural role both in the neonatal period and in adulthood. The patterns of staining of the AOB by the Bandeiraea simplicifolia and Lycopersicon esculentum lectins were the same in the 2 species, whereas the Ulex europeus and Dolichos biflorus lectins gave different patterns. In both species, lectin staining of the AOB was consistent with that of the VNNs, while the CdNN did not label any of the structures studied. The entire sensory epithelium of the pig was labelled by Ulex europeus and Lycopersicum esculentum lectins, and all 4 lectins used labelled the mucomicrovillar surface of the sensory epithelium in sheep.

Comparative anatomy of the vomeronasal cartilage in mammals: mink, cat, dog, pig, cow and horse

The vomeronasal cartilages of mink, cat, dog, pig, cow and horse were studied by dissection, microdissection and by means of series of transverse sections. In all the species studied the cartilage is of hyaline type and the medial sheet is well-defined and perfectly moulded to the adjacent bone. However, interspecies differences are apparent in the manner in which the medial sheet associates and eventually fuses with the cartilage of the incisive duct; the morphology of the horse vomeronasal cartilage is particularly distinctive in this respect. The lateral sheet of the vomeronasal cartilage, although always present, has a different arrangement in each species studied. Similarly, the gaps in the lateral sheet (corresponding to the opening of the vomeronasal organ) differ among the species studied in form, location and number.

Distribution of the arterial supply to the vomeronasal organ in the cat

The main goal of this work was to investigate the general distribution of arterial blood around and inside the vomeronasal organ (VNO) of the cat.

Oral Toxicity of Okadaic Acid in Mice: Study of Lethality, Organ Damage, Distribution and Effects on Detoxifying Gene Expression

In vivo, after administration by gavage to mice and rats, okadaic acid has been reported to produce lesions in liver, small intestine and forestomach. Because several reports differ in the damage detected in different organs, and on okadaic acid distribution after consumption, we determined the toxicity of this compound after oral administration to mice. After 24 hours, histopathological examination showed necrotic foci and lipid vacuoles in the livers of intoxicated animals. By immunohistochemical analysis, we detected this toxin in the liver and kidneys of intoxicated animals. Okadaic acid induces oxidative stress and can be activated in vitro into reactive compounds by the post-mitochondrial S9 fraction, so we studied the okadaic effect on the gene expression of antioxidant and phase II detoxifying enzymes in liver. We observed a downregulation in the expression of these enzymes and a reduction of protein expression of catalase and superoxide dismutase 1 in intoxicated animals.

Brain Pathology in Adult Rats Treated With Domoic Acid.

Domoic acid (DA) is a neurotoxin reported to produce damage to the hippocampus, which plays an important role in memory. The authors inoculated rats intraperitoneally with an effective toxic dose of DA to study the distribution of the toxin in major internal organs by using immunohistochemistry, as well as to evaluate the induced pathology by means of histopathologic and immunohistochemical methods at different time points after toxin administration (6, 10, and 24 hours; 5 and 54 days). DA was detected by immunohistochemistry exclusively in pyramidal neurons of the hippocampus at 6 and 10 hours after dosing. Lesions induced by DA were prominent at 5 days following treatment in selected regions of the brain: hippocampus, amygdala, piriform and perirhinal cortices, olfactory tubercle, septal nuclei, and thalamus. The authors found 2 types of lesions: delayed death of selective neurons and large areas of necrosis, both accompanied by astrocytosis and microgliosis. At 54 days after DA exposure, the pathology was characterized by still-distinguishable dying neurons, calcified lesions in the thalamus, persistent astrocytosis, and pronounced microgliosis. The expression of nitric oxide synthases suggests a role for nitric oxide in the pathogenesis of neuronal degeneration and chronic inflammation induced by DA in the brain.

Morphological and Immunohistochemical Features of the Vomeronasal System in Dogs

Each of the structures integrating the sense of smell in mammals has a different degree of development, even in the so‐called macrosmatic animals, according to the capacity of the olfactory system to detect thousands of different chemical signals. Such morphological diversity implies analogous physiological variation. The study of the accessory olfactory system, also known as the vomeronasal system, is a useful way to analyze the heterogeneity of the sense of smell. Macrodissection and microdissection methods as well as conventional histology and immunohistochemistry protocols were used to study aspects of the vomeronasal organ and the accessory olfactory bulbs in dogs. Observations regarding the end of the anterior part of the vomeronasal duct have been emphasized. Both lectins, Ulex europaeus agglutinin I and Lycopersicum esculentum agglutinin, and one G protein, Gαi2, show a similar pattern of binding in the sensory epithelium of the vomeronasal organ and in the vomeronasal nerve and glomerular layers of the accessory olfactory bulb, whereas the expression of protein Gαo was not observed. Taken together, our results emphasize the contribution of comparative data to our understanding of the vomeronasal system function. Anat Rec, 2013.

Morphogenesis and growth of the soft tissue and cartilage of the vomeronasal organ in pigs

The morphology of the soft tissue and supporting cartilage of the vomeronasal organ of the fetal pig was studied from early stages to term. Specimens obtained from an abattoir were aged by crown‐to‐rump distance. Series of transverse sections show that some time before birth all structures – cartilage, connective tissue, blood vessels, nerves, glands and epithelia – are well developed and very similar in appearance to those of the adult. Furthermore, in transmission electron microscopy photomicrographs obtained at this stage the vomeronasal glands exhibit secretory activity.

Origin and regional distribution of the arterial vessels of the vomeronasal organ in the sheep. A methodological investigation with scanning electron microscopy and cutting-grinding technique

The origin and location of the arteries of the vomeronasal organ (VNO) in the sheep were studied by means of dissection, scanning electron microscopy of corrosion casts, and the cutting-grinding technique after injection with Araldite CY23-HY2967 via one of the carotid arteries. Dissection revealed that the most ventral of the three main branches of the sphenopalatine artery is responsible for the blood supply to the VNO. Scanning electron microscopy of corrosion casts revealed that the arterioles of the vomeronasal organ form a microvascular network. Cross sections of the region of the nasal cavity containing the VNO, obtained by the cutting-grinding technique, showed that the arterioles of the vomeronasal plexus are located medial and ventral to the vomeronasal duct. These results confirm the usefulness of the cutting-grinding technique as a complementary procedure in morphological studies of structures containing hard tissues.

Tetracyclic Truncated Analogue of the Marine Toxin Gambierol Modifies NMDA, Tau, and Amyloid β Expression in Mice Brains: Implications in AD Pathology

Gambierol and its two, tetra- and heptacyclic, analogues have been previously proved as promising molecules for the modulation of Alzheimers disease (AD) hallmarks in primary cortical neurons of 3xTg-AD fetuses. In this work, the effect of the tetracyclic analogue of gambierol was tested in vivo in 3xTg-AD mice (10 months old) after 1 month of weekly treatment with 50 μg/kg. Adverse effects were not reported throughout the whole treatment period and no pathological signs were observed for the analyzed organs. The compound was found in brain samples after intraperitoneal injection. The tetracyclic analogue of gambierol elicited a decrease of amyloid β1-42 levels and a dose-dependent inhibition of β-secretase enzyme-1 activity. Moreover, this compound also reduced the phosphorylation of tau at the 181 and 159/163 residues with an increase of the inactive isoform of the glycogen synthase kinase-3β. In accordance with our in vitro neuronal model, this compound produced a reduction in the N2A subunit of the N-methyl-d-aspartate (NMDA) receptor. The combined effect of this compound on amyloid β1-42 and tau phosphorylation represents a multitarget therapeutic approach for AD which might be more effective for this multifactorial and complex neurodegenerative disease than the current treatments.