M. Rubio

Ependyma: phylogenetic evolution of glial fibrillary acidic protein (GFAP) and vimentin expression in vertebrate spinal cord

The phylogenetic evolution was studied of both glial fibrillary acidic protein (GFAP) and vimentin expression in the ependyma of the adult vertebrate spinal cord. Eleven species from different vertebrate groups were examined using different fixatives and fixation procedures to demonstrate any differences in immunoreactivity. GFAP expression in the ependymal cells showed a clear inverse relation with phylogenetic evolution because it was more elevated in lower than in higher vertebrates. GFAP positive cells can be ependymocytes and tanycytes, although depending on their structural characteristics and distribution, the scarce GFAP positive ependymal cells in higher vertebrates may be tanycytes. Ependymal vimentin expression showed a species-dependent pattern instead of a phylogenetic pattern of expression. Vimentin positive ependymal cells were only found in fish and rats; in fish, they were tanycytes and were quite scarce, with only one or two cells per section being immunostained. However, in the rat spinal cord, all the ependymocytes showed positive immunostaining for vimentin. The importance of the immunohistochemical procedure, the cellular nature of GFAP positive ependymal cells and the relationship between tanycytes and ependymocytes are discussed, as well as GFAP and vimentin expression.

Astroglial pattern in the spinal cord of the adult barbel (Barbus comiza)

The distribution and the structural, ultrastructural and immunohistochemical characteristics of the astroglial cells in the spinal cord of the adult barbel (Barbus comiza) have been studied by means of metallic impregnations (Golgi and gold-sublimate), immunohistochemical (GFAP and vimentin) and electron microscopic techniques. GFAP-positive cells were mainly distributed in the ependyma and in the periependymal region, but they have also been observed at subpial level in the anterior column. The ependymocytes were heterogeneous cells because they showed different immunohistochemical characteristics: GFAP-positive, vimentin-positive or non-immunoreactive cells. The radial astrocytes showed only GFAP immunoreactivity, and their processes ended at the subpial zone forming a continuous subpial glia limitans. Desmosomes and gap junctions between soniata and processes of radial astrocytes were numerous, and a relationship between radial astroglial processes and the nodes of Ranvier was also described. The perivascular glia limitans was poorly developed and it was not complete in the blood vessels of the periependymal zone; in this case, the basal lamina was highly developed. An important characteristic in the barbel spinal cord was the existence of a zone with an abundant extracellular space near the ependyma. The presence of radial astroglial somata at subpial level, the existence of vimentin-positive ependymocytes and the abundant extracellular space in the periependymal zone is discussed in relation to the regeneration capacity and the continuous growth showed by fish. Moreover, the abundance of gliofilaments and desmosomes leads us to suggest that mechanical support might be an important function for the astroglial cells in the barbel spinal cord.

Down-regulation of the AMPA glutamate receptor subunits GluR1 and GluR2/3 in the rat cerebellum following pre- and perinatal Δ9-tetrahydrocannabinol exposure

This paper reports the effects of pre- and perinatal exposure to Δ9-tetrahydrocannabinol (THC) on expression levels of specific AMPA glutamate receptor subunits (GluRl and GluR2/3) in the cerebellum of male and female rats. Pregnant rats were administered saline or THC from gestational day 5 (ED5) to postnatal day 20 (PD20). Expression of the GluRl and GluR2/3 subunits of AMPA glutamate receptors was analyzed by immunohistochemistry in THC-exposed rats at three postnatal ages: PD20 (still exposed to THC) to study the direct effect of drug exposure, and PD30 and PD70 (10 and 50 days following THC withdrawal) to analyze the long-term effects of prenatal exposure. Compared to controls, pre- and perinatal THC exposure decreased the immunoreactivity levels of the GluRl subunit in Bergmann glial cells, as well as levels of the GluR2/3 subunit in Purkinje neurons at PD20. These changes in AMPA receptor subunit levels may correlate with the decreased excitatory neurotransmission described in the cerebellum after cannabinoid treatment, which could play a significant role in the biochemical effects of THC. In addition, the reduced glutamate receptor expression observed at PD20 did not return to normal even after THC withdrawal (PD30 and PD70). The results support the idea that THC exposure during critical stages of cerebellar development may alter the glutamatergic system, not only during the drug exposure period itself but also in adults following THC withdrawal. The decreased expressions of glutamate receptors induced by developmental THC exposure could lead to functional alterations through the inhibition of glutamatergic neurotransmission, and clearly demonstrate an interaction between cannabinoids and the glutamatergic system.

Glial fibrillary acidic protein and vimentin immunohistochemistry in the posterior rhombencephalon of the Iberian barb (Barbus comiza)

The gustatory centers (vagal lobes and facial lobe) and the tegment of the posterior rhombencephalon of the Iberian barb (Barbus comiza) have been studied using anti-glial fibrillary acidic protein (anti-GFAP) and anti-vimentin immunohistochemical techniques. GFAP immunoreactivity was found in the tegment and in a part of the vagal lobes while vimentin immunoreactivity was located in the tegment. Two immunopositive cell types were found: ependymocytes and radial astrocytes. Since the distribution of GFAP in the barb rhombencephalon corresponds with zones previously described as cholinergic, the GFAP-immunopositive radial astrocytes might be involved in acetylcholine metabolism.

Prenatal cannabinoid exposure down- regulates glutamate transporter expressions (GLAST and EAAC1) in the rat cerebellum.

Efficient reuptake of synaptically released glutamate is essential for preventing glutamate receptor overstimulation and neuronal death. Glutamate transporters play a vital role in removing extracellular glutamate from the synaptic cleft. This study analyzed the expression of the glial (GLAST) and neuronal (EAAC1) subtypes of glutamate transporter in the cerebellum of male and female offspring exposed pre- and postnatally to Δ9-tetrahydrocannabinol (THC, the main component of marijuana). Pregnant rats were administered saline or THC from gestational day 5 to postnatal day 20 (PD20). The expression of glutamate transporters was examined at PD20, PD30 and PD70 (10 and 50 days after THC withdrawal) to analyze the short- and long-term effects of prenatal THC exposure. The expression of the glutamate transporter GLAST in astroglial cells and EAAC1 in Purkinje neurons decreased in THC-exposed offspring compared to controls. This reduction was observed at all ages but mainly in males. Moreover, the glial glutamate transporter level in THC-exposed rats (quantified by Western blot) was lower than in control rats. These results suggest that THC exposure during cerebellar development may alter the glutamatergic system not only during the period of drug exposure but in the postnatal stage following withdrawal. The down-regulation reported here might reflect an abnormal maturation of the glutamatergic neuron-glia circuitry.

Distribution and characteristics of the different astroglial cell types in the adult lizard (Lacerta lepida) spinal cord

SummaryThe astroglial cells have been studied in the lizard spinal cord by means of metallic impregnations, immunohistochemical (glial fibrillary acidic protein) and ultrastructural methods. Three astroglial cell types have been immunohistochemically identified: ependymocytes, radial astrocytes and astrocytes. Transitional forms have also been observed. Scarce immunopositive ependymocytes were located in the dorsal and ventral regions of the ependyma. The radial astrocytic somata were located around the ependymal layer and their processes reached the subpial glia limitans. Typical astrocytes were the most abundant astroglial cell type; astrocytes located in the ventral horn showed a greater development than those of the dorsal horn. In the white matter, the astrocytes were large and their processes formed part of the subpial glia limitans; on some occasions, astrocytic cell bodies also formed part of this subpial limitans. Transitional elements between astrocytes and radial astrocytes were observed in both grey and white matter. The perivascular and subpial glia limitans were continuous and showed a strong immunoreactivity. The comparative analysis of our results in the lizard spinal cord with those in other vertebrate groups leads us to conclude that reptiles could represent the key group in the phylogenetic evolution of the astroglial cells in vertebrates.

Reduced Glial Fibrillary Acidic Protein and Glutamine Synthetase Expression in Astrocytes and Bergmann Glial Cells in the Rat Cerebellum Caused by Δ9-Tetrahydrocannabinol Administration during Development

In this study we analyzed the responses of cerebellar astroglial cells to pre- and perinatal Δ9-tetrahydrocannabinol (THC) exposure in three postnatal ages and both sexes. To determine whether THC during development directly modifies astroglial growth, this study investigated the effects of THC on astroglial morphological changes and on the expression of specific astroglial markers (glial fibrillary acidic protein: GFAP and glutamine synthetase: GS). Our results demonstrated that the administration of THC during development has deleterious effects on astroglial maturation in the cerebellum. These results also indicate that THC might interfere with astroglial differentiation in a way dependent on sex. The effect of cannabinoids on the development of cerebellar astroglial cells (astrocytes and Bergmann glial cells) is to reduce protein synthesis, since both GFAP and GS decreased in astroglial cells, not only during THC exposure but also in adult ages. Our data suggest that pre- and perinatal THC exposure directly interferes with astroglial maturation by disrupting normal cytoskeletal formation, as indicated by the irregular disposition of GFAP and the lower GFAP expression observed at all the ages studied. THC exposure during development may also modulate glutamatergic nervous activity since GS expression is reduced in THC-exposed brains. GS expression increased progressively after THC withdrawal, but GS expression had still not reached control values two months after THC withdrawal. This indicates that glutamate uptake is lower in glial cells exposed to THC, since GS expression is lower than in older controls. Consequently, glutamatergic neurotransmission may be affected by cannabinoid exposure during gestation. Therefore, cannabinoids exert developmental toxicity, at least on astroglial cells, which could contribute to fetal brain growth retardation.

Neuronal and inducible nitric oxide synthase expression in the rat cerebellum following portacaval anastomosis.

In order to determine the role of neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS) in the pathogenesis of experimental hepatic encephalopathy (HE), the expression of both was analyzed in the cerebellum of rats 1 month and 6 months after performing portacaval anastomosis (PCA). In control cerebella, nNOS immunoreactivity was mainly observed in the molecular layer (ML), whereas the Purkinje cells did not express nNOS. However, nNOS expression was detected in the Purkinje cells at 1 month after PCA, correlating with a decrease in nNOS expression in the ML--part of an overall reduction in cerebellar nNOS concentrations (as determined by Western blotting). At 6 months post-PCA, a significant increase in nNOS expression was observed in the ML, as well as increased nNOS immunoreactivity in the Purkinje cells. nNOS immunoreactivity was also observed in the Bergmann glial cells of PCA-treated rats. While no immunoreactivity for iNOS was seen in the cerebella of control rats, iNOS immunoreactivity was significantly induced in the cerebellum 1 month after PCA. In addition, the expression of iNOS was greater at 6 months than at 1 month post-PCA. Immunohistochemical analysis revealed this iNOS to be localized in the Purkinje cells and Bergmann glial cells. The induction of iNOS in astroglial cells has been associated with pathological conditions. Therefore, the iNOS expression observed in the Bergmann glial cells might play a role in the pathogenesis of HE, the harmful effects of PCA being caused by them via the production of excess nitric oxide. These results show that nNOS and iNOS are produced in the Purkinje cells and Bergmann glial cells following PCA, implicating nitric oxide in the pathology of HE.

Very Late Thrombosis of a Transcatheter Aortic Valve-in-Valve.

An 85-year-old man presented with dyspnea New York Heart Association functional class IV 4 years after implantation of a 26-mm CoreValve transcatheter aortic valve (TAV) (Medtronic, Minneapolis, Minnesota). Angiography revealed severe aortic regurgitation due to the deep position of the prosthesis

Software project effort estimation based on multiple parametric models generated through data clustering

Parametric software effort estimation models usually consists of only a single mathematical relationship. With the advent of software repositories containing data from heterogeneous projects, these types of models suffer from poor adjustment and predictive accuracy. One possible way to alleviate this problem is the use of a set of mathematical equations obtained through dividing of the historical project datasets according to different parameters into subdatasets called partitions. In turn, partitions are divided into clusters that serve as a tool for more accurate models. In this paper, we describe the process, tool and results of such approach through a case study using a publicly available repository, ISBSG. Results suggest the adequacy of the technique as an extension of existing single-expression models without making the estimation process much more complex that uses a single estimation model. A tool to support the process is also presented.