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G Alonso

Publications and source records attributed to G Alonso.

At least 55 records · Page 3Linked to original sources
PubMed1999

Tangential migration of young neurons arising from the subventricular zone of adult rats is impaired by surgical lesions passing through their natural migratory pathway.

In the brain of adult rodents, young neurons arising from the subventricular zone (SVZ) of the lateral ventricle migrate tangentially along the rostral migratory stream (RMS) toward the olfactory bulb. The aim of this study was to determine whether surgical lesions placed through the RMS could affect the rostral migration of these newly formed neurons. Confocal and electron microscopy were used to characterize their anatomical organization within the intact and lesioned forebrains. As soon as 7 days and up to 45 days after placing a surgical lesion through the proximal portions of the RMS, numerous cells immunostained for polysialylated neural cell adhesion molecule (PSA-NCAM) were detected both (1) throughout the lesional cavity extending from the cortex to the anterior commissura, and (2) within the tissue located caudal to the lesion. In both regions, these PSA-NCAM-immunostained cells were labeled for neuronal markers but were negative for glial fibrillary acidic protein (GFAP). After administration of the proliferation marker bromodeoxyuridine (BrdU), nuclear labeling was associated with cells immunostained for PSA-NCAM but GFAP-negative, that accumulated within the lesional cavity and in the tissue caudal to the lesion. For the longest postlesional delays, a number of the PSA-NCAM-immunostained neurons located in various portions of the lesional cavity exhibited intense immunostaining for gamma-aminobutyric acid, whereas only a few of them exhibited faint immunostaining for tyrosine hydroxylase. These data indicate that surgical lesions placed through the RMS of adult rats impede the migration toward the olfactory bulb of the neuroblasts arising from the SVZ, inducing their accumulation and their partial differentiation in forebrain regions caudal to the lesion.

Animals↗
PubMed1999

Effects of stimulus preexposure on the generalization of conditioned taste aversions in infant rats.

Generalization of a conditioned taste aversion in infant rats and how this is affected by stimulus preexposure was investigated in a series of experiments. In Experiment 1 generalization of a conditioned aversion between two tastes (sweet and salty) was found, and the effect of tastes preexposure was a reduction in generalization (Experiment 2). However, when these tastes were combined with a common taste (acid) that was less (Experiment 3) or more intense (Experiment 3b), the effect of stimulus preexposure was a stronger generalization of the conditioned aversion. In this case, a reduction on generalization was again observed by increasing the number of preexposure trials to the taste compounds (Experiment 4). In all cases the generalization levels were directly related to the effect of stimulus preexposure on the acquisition rate of conditioning. It can be concluded that, with the appropriate parameters, a reduction of generalization of a conditioned taste aversion can be obtained after taste exposure in preweanling rats.

Animals↗
PubMed1999

Differential sensitivity of cultured tanycytes and astrocytes to hydrogen peroxide toxicity.

Tanycytes present in the mediobasal hypothalamus are able to support axonal regeneration and neuron survival. Pilot experiments of transplantation of these cells into various lesioned areas of the central nervous system (CNS) were thus performed to determine whether these cells could support the regeneration of the lesioned axons. These pilot experiments, however, demonstrated that the grafted tanycytes failed to survive in the lesioned sites. The present study was designed to determine which of the compounds released at the lesion would be toxic for tanycytes. Tanycyte cultures obtained from the median eminence of 10-day-old rats and astrocyte cultures obtained from the cortex of 10-day-old rats or E-14 embryos were incubated with two types of toxic molecules, including excitatory amino acids (EAA) and hydrogen peroxide (H2O2). The effect of these substances on cell death was estimated by measuring the lactate deshydrogenase (LDH) released and the surface occupied by immunostained glial structures after each treatment. The results indicated that the viability of both the tanycytes and the astrocytes was not affected by incubation for 24 h with 1 mM glutamate or 1 mM kainate. In contrast, increasing concentrations of H2O2 induced concentration-dependent cell death of tanycytes and immature astrocytes, without affecting the mature astrocytes. The use of antioxidant molecules such as catalase, tempol, or vitamin C effectively protected cultured tanycytes from H2O2 toxicity. These data indicate that (1) both mature astrocytes and tanycytes are resistant to EAA and (2) contrary to mature astrocytes, immature astrocytes and tanycytes are sensitive to the free radicals generated by H2O2. This suggest that oxidative stress is at least partly responsible for the death of tanycytes grafted into the lesioned CNS.

Animals↗
PubMed1999

Brain-derived neurotrophic factor and neurotrophin-3 enhance somatostatin gene expression through a likely direct effect on hypothalamic somatostatin neurons.

Although neurotrophins (NTs) have been extensively studied as neuronal survival factors in some areas of the central nervous system, little is known about their function or cellular targets in the hypothalamus. To understand their functional significance and sites of action on hypothalamic neurons, we examined the effects of their cognate ligands on neuropeptide content and messenger RNA (mRNA) expression in somatostatin neurons present in fetal rat hypothalamic cultures. Treatments were performed in defined insulin-free medium between days 6 and 8 of culture, since the maximal effects of NTs on somatostatin content and mRNA expression were observed after 48-h incubations. Brain-derived neurotrophic factor and NT-3, but not nerve growth factor, induced a dose-dependent increase in somatostatin content, which was influenced by plating density. The same treatment increased somatostatin mRNA and immunostaining intensity of somatostatin neurons, but had no effect on the number of these labeled neurons. The increased levels of somatostatin (peptide and mRNA) induced by NTs were not blocked by tetrodotoxin or by glutamate receptor antagonists, suggesting that endogenous neurotransmitters (e.g. glutamate) were not involved in these effects. In contrast, the stimulatory effects were completely blocked by K-252a, an inhibitor of tyrosine kinase (Trk) receptors, whereas the less active analog K-252b was ineffective. Double-labeling studies demonstrated that both TrkB or TrkC receptors were located on somatostatin neurons. Our results show that, in rat hypothalamic cultures, brain-derived neurotrophic factor, and NT-3 have a potent stimulatory effect on peptide synthesis in somatostatinergic neurons, likely through direct activation of TrkB and TrkC receptors.

Animals↗
PubMed1998

Src-like adaptor protein (Slap) is a negative regulator of mitogenesis.

The Src-like adaptor protein (Slap) is a recently identified adaptor protein containing Src homology 3 (SH3) and SH2 domains. Slap is found in a wide range of cell types and was shown to interact with the Eck receptor tyrosine kinase in a yeast two-hybrid interaction screen [1]. Here, we found that Slap is expressed in NIH3T3 cells and could associate with the activated platelet-derived growth factor (PDGF) receptor. Using mutated versions of the PDGF receptor and phosphopeptide competition experiments, we determined that Slap has the highest affinity for the Src-binding site of the PDGF receptor. Our inability to produce cell lines that stably expressed Slap suggested that Slap inhibited cell growth. We further investigated this issue by transiently expressing Slap by microinjection. Overexpression of Slap by this method inhibited DNA synthesis induced by PDGF and serum, whereas overexpression of the adaptor proteins Grb2 and Shc did not. Finally, microinjection of a Slap antibody into NIH3T3 cells that had been stimulated with suboptimal doses of growth factors potentiated the effects of the growth factors. These data suggest that, unlike other adaptor proteins, Slap is a negative regulator of signalling initiated by growth factors.

3T3 Cells↗
PubMed1998

Pregnenolone reverses the age-dependent accumulation of glial fibrillary acidic protein within astrocytes of specific regions of the rat brain.

Although aged-related modifications of astrocytes have been frequently described, little is known so far about the signals responsible for these modifications. Since it is well demonstrated that astrocytes are highly responsive to a variety of steroids, we hypothesized that modifications of cerebral astrocytes may result from the age-related decrease of circulating steroids. In the present study, we investigated the effects of the chronic administration of pregnenolone (PREG), the precursor of all steroid hormones, on the age-related extension of astrocytic processes in various brain regions. In adult (2-3 month-old) and aged (22-24 month-old) rats, quantitative image analysis was used to estimate, within each region, the number of astrocyte cell bodies immunostained (IS) for S100, and the surface occupied by astrocytic cell bodies and processes IS for glial fibrillary acidic protein (GFAP). In all regions, the surface occupied by GFAP-IS structures was increased in the aged vs. the adult rats, whereas no significant modifications were observed in the number of S100-IS cell bodies. Chronic administration of PREG to aged rats induced a marked decrease in the surface occupied by GFAP-IS structures in the cortex, amygdala and thalamus, without any significant effect on the number of S100-IS cell bodies present in these regions. By contrast, PREG had no significant effect when administered to adult animals. These data suggest that decreased levels of circulating steroid hormones may be responsible for the age-dependent modifications of the astrocytes present in various brain regions, and that these modifications can be at least partly corrected by the administration of PREG.

Aging↗
PubMed1998

Tanycytes present in the adult rat mediobasal hypothalamus support the regeneration of monoaminergic axons.

We have recently shown that tanycytes present in the median eminence (ME) constitute a preferential support for the regeneration of lesioned neurohypophysial oxytocinergic and vasopressinergic axons. However, although tanycytes are particularly abundant in the ME, they are also present along the third ventricle wall. This study was thus undertaken to determine whether tanycytes present in the mediobasal hypothalamus overlying the ME were also able to support the regeneration of the numerous monoaminergic axons innervating this region. Using confocal laser scanning microscopy combined with double or triple fluorescence immunostaining, we have compared the relationships occurring between glial cells and lesioned catecholaminergic and serotonergic axons at the levels of surgical cuts placed in the dorsomedial hypothalamus devoid of tanycytes or in the ventromedial hypothalamus containing numerous tanycyte processes. In dorsal lesions, catecholaminergic and serotonergic transected fibers were found to abut onto the scar formed along the surgical cut and composed of closely inderdigitating astrocyte processes strongly immunoreactive for both glial fibrillary acidic protein (GFAP) and vimentin (VIM). In ventral lesions, the lesional scar was composed of GFAP-immunoreactive (IR) and VIM-IR astrocyte processes and of VIM-IR but GFAP-negative processes that were identified as tanycytic processes. In all the ventral lesions examined, numerous catecholaminergic and serotonergic fibers were found to regenerate into the surgical cut in association with the VIM-IR, GFAP-negative tanycyte processes. On the other hand, such regenerating fibers were never found in scar portions containing only GFAP-IR astrocytic structures. These data indicate that, like in the ME, tanycytes present in the mediobasal hypothalamus of adult rat provide a substrate that favors the regeneration of lesioned axons.

Age Factors↗
PubMed1998

Glucocorticoids provoke a shift from alpha2- to alpha1-adrenoreceptor activities in cultured hypothalamic slices leading to opposite noradrenaline effect on corticotropin-releasing hormone release.

We have shown previously that noradrenaline (NA) stimulated or inhibited the release of corticotropin-releasing hormone (CRH) according to the availability of adrenal steroids. The aim of the present work was to examine whether the changes in the NA modulation of CRH release from hypothalamic neurons result from a steroid-induced plasticity of the adrenergic transduction pathways. From anterior hypothalamic slices cultured in standard medium (i.e., containing adrenal steroids at a final dilution of 61 +/- 9 ng/ml), (a) the stimulatory effect of NA on CRH release was reversed in a dose-dependent manner by increasing concentrations of the alpha1-adrenoreceptor antagonist prazosin, (b) activation of protein kinase C by acute treatment with phorbol 12-myristate 13-acetate (0.5 microM, 1 h) mimicked NA stimulation of CRH secretion, and (c) the activation of L-type Ca2+ channels by Bay K 8644 also produce an increased CRH secretion. In contrast, the inhibitory effect of NA on CRH secretion from slices cultured in steroid-free medium was markedly reversed by the alpha2-adrenoreceptor antagonist yohimbine, by pretreatment with pertussin toxin, or by the addition of 4-aminopyridine, a K+-channel blocker. Acute treatment with phorbol 12-myristate 13-acetate did not change the inhibitory NA effect. Moreover, all these effects were reversed by daily corticosterone supplementation, for as long as they were tested. These results are consistent with a steroid-dependent change in the nature of adrenergic receptors and its associated transduction pathways involved in the regulation of CRH secretion in the hypothalamus.

4-Aminopyridine↗
PubMed1998

Functional studies of twelve mutant V2 vasopressin receptors related to nephrogenic diabetes insipidus: molecular basis of a mild clinical phenotype.

X-linked nephrogenic diabetes insipidus (NDI) is a rare disease with defective renal and extrarenal arginine vasopressin V2 receptor responses due to mutations in the AVPR2 gene in Xq28. To study the cause of loss of function of mutant V2 receptors, we expressed 12 mutations (N55H, L59P, L83Q, V88M, 497CC-->GG, deltaR202, I209F, 700delC, 908insT, A294P, P322H, P322S) in COS-7 cells. Eleven of these, including P322H, were characterized by a complete loss of function, but the mutation P322S demonstrated a mild clinical and in vitro phenotype. This was characterized by a late diagnosis without any growth or developmental delay and a significant increase in urine osmolality after intravenous 1-deamino[D-Arg8]AVP administration. In vitro, the P322S mutant was able to partially activate the Gs/adenylyl cyclase system in contrast to the other V2R mutants including P322H, which were completely inactive in this regard. This showed not only that Pro 322 is important for proper V2R coupling, but also that the degree of impairment is strongly dependent on the identity of the substituting amino acid. Three-dimensional modeling of the P322H and P322S mutant receptors suggested that the complete loss of function of the P322H receptor could be due, in part, to hydrogen bond formation between the His 322 side chain and the carboxyl group of Asp 85, which does not occur in the P322S receptor.

Blotting, Western↗
PubMed1998

[Endovascular treatment of dural arteriovenous fistulas with medullary venous drainage. Experience with 18 patients].

INTRODUCTION AND OBJECTIVE: The authors report a series of 18 patients with myelopathy who were diagnosed of dural arteriovenous fistulas with venous medullary drainage (DFVMd). Purpose was to assess the effectiveness, initial and long term, of embolization, as the initial treatment, using polyvinyl alcohol particles (PVA) and liquid adhesives. N-butyl-cyanoacrylate (NBCA). MATERIAL AND METHODS: Magnetic resonance images were obtained in all patients showing spinal cord tissue changes consistent with an isquemic process secondary to venous hypertension. All 18 patients showed initially an improvement in clinical symptoms, demonstrating previous MR images resolution. RESULTS: The neurological status of 8 patients subsequently deteriorated with angiographically proven recurrences of their DFVMd. These patients underwent a second successful embolization procedure using NBVA. PVA embolization is long term ineffective and is not without risk. Endovascular treatment is less invasive than surgery, its morbidity is less, and it ensures earlier recovery for the patients. If embolization has failed, surgery can still be done. CONCLUSIONS: We recommend that NBVA embolization be the initial treatment of choice for DFVMd if referring the patient to an experienced interventional Neuroradiology unit is available. Careful clinical and neurologic examination is necessary to establish the diagnosis of DFVMd. Finally, we strongly recommend that patients be followed closely and aggressively. Periodic clinical and radiologic assessments, including MR and spinal angiography, are essential to achieve complete cure.

Aged↗
PubMed1998

The gluconate high affinity transport of GntI in Escherichia coli involves a multicomponent complex system.

Within the main system for gluconate utilization in E. coli, the gntT gene (located at the minute 76.4) that encodes a permease, is currently the only element involved in the high affinity transport. In this paper, the nucleotide sequence of the upstream region of this locus was determined. Two open reading frames of 729 bp (gntX) and 573 bp (gntY) were identified as additional gnt genes by complementation studies. Our observations suggest that these loci might conform an operon distinct of gntT under the control of the gntR gene product. Such operon encodes a gluconate periplasmic binding protein (GntX) and a putative membrane-bound protein (GntY). These products and the permease encoded by the gntT gene seem to conform a high-affinity complex transport system for gluconate. We suggest that this novel system could belong to the TRAP transporters.

Bacterial Proteins↗
PubMed1997

PDGF-induced phosphorylation of Tyr28 in the N-terminus of Fyn affects Fyn activation.

Binding of platelet-derived growth factor (PDGF) to its receptors leads to the activation of members of the Src family of protein tyrosine kinases. We show here that Fyn, a member of the Src family, is phosphorylated on Tyr28 in the unique N-terminal part of the molecule after interaction with the intracellular domain of the PDGF beta-receptor. Activated Fyn furthermore undergoes autophosphorylation on Tyr30, Tyr39 and Tyr420. When Fyn mutants with Tyr28, Tyr30 or Tyr39 replaced with phenylalanine residues were transfected into NIH3T3 cells a decreased activation after PDGF stimulation was seen, suggesting a functional importance of the N-terminal tyrosine phosphorylation of Fyn.

3T3 Cells↗
PubMed1997

PSA-NCAM and B-50/GAP-43 are coexpressed by specific neuronal systems of the adult rat mediobasal hypothalamus that exhibit remarkable capacities for morphological plasticity.

The present study was designed to determine whether the mediobasal hypothalamus of adult rats contains neurons that continue to coexpress the highly polysialylated neural cell adhesion molecule (PSA-NCAM) and B-50/GAP-43, two proteins coexpressed by virtually all of the neurons of the fetal and neonatal rat central nervous system. Confocal laser scanning microscopy combined with double- or triple-fluorescence immunostaining was used to identify the hypothalamic neurons that express high levels of both PSA-NCAM and B-50/GAP-43 and to study the possible modifications of their morphological organization following a surgical lesion through the mediobasal hypothalamus. In intact animals, PSA-NCAM and B-50/GAP-43 were found to be colocalized within numerous fibers projecting throughout the external layer of the median eminence that were immunoreactive for either gamma-aminobutyric acid (GABA) or tyrosine hydroxylase (TH). Three to 30 days after a lesion through this region, numerous regenerating axonal sprouts, triple-immunostained for PSA-NCAM, B-50/GAP-43, and either GABA or TH, were detected along the ventricular surface of, and throughout the perivascular layer of, the median eminence. Surprisingly, high levels of PSA-NCAM and B-50/GAP-43 were also associated with numerous supraependymal neurons that exhibited long ramified processes and were immunoreactive for GABA but TH-negative. The use of the proliferation marker, 3H-thymidine, further indicated that the emergence of such supraependymal neurons after median eminence lesion was not related to the proliferation of preexisting quiescent cells. These data indicate that the mediobasal hypothalamus of the adult rat contains two neuronal systems, in which the continued coexpression of PSA-NCAM and B-50/GAP-43 is related to remarkable capacities for postlesional, morphological plasticity.

Animals↗
PubMed1997

Mab22C11 antibody to amyloid precursor protein recognizes a protein associated with specific astroglial cells of the rat central nervous system characterized by their capacity to support axonal outgrowth.

Amyloid precursor protein (APP) is a transmembrane glycoprotein which is believed to promote neural cell adhesion, neural survival, and neuritogenesis. The present study was undertaken to determine whether APP could be detected within different types of astroglial cells present in the central nervous system (CNS) of neonatal or adult rats. The localization of this protein within glial cells was studied by using a monoclonal antibody (Mab22C11) that recognizes all APP isoforms and in addition cross-reacts with APP-like proteins. In the brain of neonatal rats, Mab22C11 immunostaining was associated with numerous elongated radial glia-like structures. In the intact brain and spinal cord of adult rats, Mab22C11 immunostaining was associated with (i) numerous neuron-like structures and (ii) glial structures immunostained for glial fibrillary acidic protein (GFAP) and/or vimentin, including tanycytes mostly located in the mediobasal hypothalamus, fibrous astrocytes located in the white matter and ependymocytes bordering the ventricles. On the other hand, all the GFAP-immunostained astrocytes located in the grey matter were Mab22C11 negative. In the lesioned brain and spinal cord of adult rats, Mab22C11 immunostaining was associated with intensely GFAP-immunostained reactive astrocytes located close to a surgical lesion, but not with those induced by Wallerian degeneration that appear at a distance from a lesion. Electron microscopic observations further indicated that in all these labeled astroglial cells, Mab22C11 immunostaining was mainly localized to the limiting plasma membrane and the membrane of intracytoplasmic cisternae and vesicles. These data indicate that Mab22C11 antibody induces strong immunostaining of specific astroglial cells of the neonatal and adult rat CNS that support axonal outgrowth, therefore suggesting that an APP-like protein associated with these cells participates in their axonal outgrowth promoting properties.

Amyloid beta-Protein Precursor↗
PubMed1997

Monoclonal antibodies against glutaraldehyde-conjugated histamine: application to immunocytochemistry.

We have developed mouse monoclonal antibodies (AHA-1-5, all IgG1 sub-isotype mAbs) against histamine (HA) conjugated to bovine serum albumin using glutaraldehyde-NaBH4. Among these, AHA-1 mAb was found to be the most useful for HA immunocytochemistry (ICC) in terms of specificity and sensitivity without non-specific immunobinding. AHA-1 was demonstrated to be specific to HA with an enzyme-linked immunosorbent assay (ELISA) binding test, simulating the ICC of tissue sections, and not reactive to any of the other amino acids and peptides with N-terminal histidine tested. By use of this antibody, indirect immunoperoxidase staining was observed in rat stomach fixed with glutaraldehyde (GA) in combination with NaBH4 reduction. In contrast, no immunoreactivity was seen in tissue fixed only with GA. Absorption controls indicated that the immunostaining could be completely inhibited by GA-conjugated HA, which was consistent with the results of an ELISA inhibition test. No cross-reactivity occurred with other GA-conjugated amino acids. ICC staining was dense in the cytoplasm of gastric enterochromaffin-like cells and very weak in mast cells. A new finding was that staining was noticed in some cell bands of the intermediate layer between the stratum lucidum and the stratum corneum of the stratified squamous epithelium of the gastric cardia, esophagus, tongue, and skin in rats. The results strongly suggest that the monoclonal antibody allowed highly specific detection of HA in animal tissues.

Animals↗
PubMed1997

Immunolocalization of Bcl-xL/S in the central nervous system of neonatal and adult rats.

A polyclonal antibody raised against a peptide corresponding to the (2-19) amino-terminal sequence of the Bcl-xL/S protein was used to localize Bcl-x immunostaining in the central nervous system of rats at various postnatal ages. Whereas Bcl-x immunostaining was present in virtually all neurons of young animals (4 days postnatal), this staining became progressively restricted during the course of postnatal development. In adults, Bcl-x immunostaining was particularly strong in certain neurons present in a few hypothalamic nuclei, such as the supraoptic or the arcuate nuclei. Moderate staining was observed in some discrete brain regions, such as the olfactory bulb, the hippocampus, some catecholaminergic nuclei of the brainstem, and the cerebellum. Strong Bcl-x immunostaining was also exhibited in axon-like fibers located in the pyriform cortex, the median eminence, the dorsal medulla oblongata, and spinal cord. Bcl-x immunostaining was also present in astrocytes scattered throughout the white matter in the brain and the spinal cord, but was absent from those located in gray matter. Staining was particularly strongly expressed in reactive astrocytes densely packed along the borders of a central lesion or surrounding them, and in a large number of reactive astrocytes detected at a distance from the lesion. Our data suggest that, in addition to the possible stimulating effects on cell survival generally ascribed to Bcl-x, its maintained expression throughout adulthood or its re-expression following injury characterizes those neuronal or non-neuronal cells of the adult central nervous system that synthesize a range of molecules enabling them to adapt rapidly and successfully to a changing environment.

Animals↗