Publicaciones por año
Publicaciones del Departamento de Histología y Embriología
Sympathetic pharmacological denervation in ageing rats: effects on ovulatory response and follicular population
Reprod Fertil Dev 2007 19(8):954-60
Rebeca Chávez-Genaro 1 4, Paula Lombide 1 2, Roberto Domínguez 3, Patricia Rosas 3, Francisco Vázquez-Cuevas 3
1 Histology and Embryology Department, School of Medicine, General Flores 2125, CP 11800, Montevideo, Uruguay. 2 Cell Biology Section, Sciences Faculty, Iguá 4225, CP 11400, Montevideo, Uruguay. 3 Biology of Reproduction Research Unit, Laboratory of Reproductive Physiology, Facultad de Estudios Superiores Zaragoza, Universidad Nacional Autónoma de México, AP 9-020, CP 15000, México, D. F. México. 4 Corresponding author. Email: rchavez@fmed.edu.uy
DOI: 10.1071/rd07075
PMID: 18076827
Pubmed: https://pubmed.ncbi.nlm.nih.gov/18076827
Texto completo: https://www.publish.csiro.au/RD/RD07075
Abstract:
The present study analyses the participation of ovarian innervation during reproductive senescence. We use the model of acute peripheral pharmacological sympathetic denervation with guanethidine in young (3 months old), middle-aged (12 months old) or old (18 months old) rats with spontaneous or induced ovulation. Ovarian levels of norepinephrine (NE) were measured by HPLC and the oestrous cycle, the number of ovulating animals and the percentage of atretic follicles were also assessed. Aged animals showed a progressive reduction in ovulatory capacity and an increase in ovarian NE content. Acute denervation increased the percentage of healthy follicles in 12- and 18-month-old rats compared with control adult animals. Combined treatment of denervation plus stimulation with gonadotrophins doubled the number of ova shed in young adult rats and restablished a partial ovulation in 12-month-old rats. The results suggest that ovarian noradrenergic innervation plays a modulator role in ovarian physiology during the ageing ovary process. The action of ovarian noradrenergic innervation seems to be associated with folliculogenesis and the ovarian response to gonadotrophins.
A quantitative study of rat uterine sympathetic innervation during pregnancy and post partum
Reprod Fertil Dev 2006 2006;18(5):525-31
R Chávez-Genaro 1 4, P Lombide 1 2, G Anesetti 1 3
1 Departamento de Histología y Embriología, Facultad de Medicina, General Flores 2125, CP 11800, Montevideo, Uruguay. 2 Sección Biología Celular, Departamento de Biología Celular y Molecular, Instituto de Biología, Facultad de Ciencias, Iguá 4245, CP 11400, Montevideo, Uruguay. 3 Area de Histología y Embriología, Facultad de Veterinaria, Las Places 1550, CP 11600, Montevideo, Uruguay. 4 Corresponding author. Email: rchavez@fmed.edu.uy
DOI: 10.1071/rd05053
PMID: 16836959
Pubmed: https://pubmed.ncbi.nlm.nih.gov/16836959
Texto completo: https://www.publish.csiro.au/rd/RD05053
Abstract:
In mammals, pregnancy induces a transient and extensive degeneration of uterine sympathetic innervation. We used the models of unilateral oviduct ligation and in oculo myometrium transplant in pregnant rats to address the role of stretching forces and/or hormone milieu in the loss of sympathetic innervation. The sympathetic fibres of the uterine horn and in oculo myometrial transplants were quantified on tissue sections processed by the glyoxylic acid technique. In normal pregnant rats, the density of uterine horn innervation was significantly reduced at late pregnancy and recovery took place during post partum. The empty horn of pregnant rats showed no significant changes in density of myometrial innervation during pregnancy or post partum. In oculo myometrial transplants were organotypically reinnervated in virgin animals. When the transplants were exposed to gestational hormonal milieu, few or no fibres were observed to the end of pregnancy; however, a significant increase at post partum was observed. Results showed that both the effects of stretching and the hormone milieu derived from the fetus-placenta complex play a role as inductors of changes on sympathetic myometrial innervation during pregnancy and support the idea that immature muscular uterine fibres are more susceptible to the effects of pregnancy than those originating from adult animals.
Fibroblast growth factor-1 induces heme oxygenase-1 via nuclear factor erythroid 2-related factor 2 (Nrf2) in spinal cord astrocytes: consequences for motor neuron survival
J Biol Chem 2005 Jul 8;280(27):25571-9
Marcelo R Vargas 1, Mariana Pehar 1, Patricia Cassina 2, Laura Martínez-Palma 2, John A Thompson 3, Joseph S Beckman 4, Luis Barbeito 1
1 Departamento de NeurobiologíaCelular y Molecular, Instituto de Investigaciones Biológicas ClementeEstable, Montevideo 11600, Uruguay, the 2 Departamento de Histología, Facultad deMedicina, Universidad de la República, Montevideo 11800, Uruguay, the 3 Department of Surgery, University of Alabama atBirmingham, Birmingham, Alabama 35294, and the 4 Linus Pauling Institute, Environmental HealthSciences Center, Department of Biochemistry and Biophysics, Oregon StateUniversity, Corvallis, Oregon 97331
DOI: 10.1074/jbc.M501920200
PMID: 15870071
Pubmed: https://pubmed.ncbi.nlm.nih.gov/15870071
Texto completo: https://linkinghub.elsevier.com/retrieve/pii/S0021-9258(20)61369-2
Abstract:
Fibroblast growth factor-1 (FGF-1) is highly expressed in motor neurons and can be released in response to sublethal cell injury. Because FGF-1 potently activates astroglia and exerts a direct neuroprotection after spinal cord injury or axotomy, we examined whether it regulated the expression of inducible and cytoprotective heme oxygenase-1 (HO-1) enzyme in astrocytes. FGF-1 induced the expression of HO-1 in cultured rat spinal cord astrocytes, which was dependent on FGF receptor activation and prevented by cycloheximide. FGF-1 also induced Nrf2 mRNA and protein levels and prompted its nuclear translocation. HO-1 induction was abolished by transfection of astrocytes with a dominant-negative mutant Nrf2, indicating that FGF-1 regulates HO-1 expression through Nrf2. FGF-1 also modified the expression of other antioxidant genes regulated by Nrf2. Both Nrf2 and HO-1 levels were increased and co-localized with reactive astrocytes in the degenerating lumbar spinal cord of rats expressing the amyotrophic lateral sclerosis-linked SOD1 G93A mutation. Overexpression of Nrf2 in astrocytes increased survival of co-cultured embryonic motor neurons and prevented motor neuron apoptosis mediated by nerve growth factor through p75 neurotrophin receptor. Taken together, these results emphasize the key role of astrocytes in determining motor neuron fate in amyotrophic lateral sclerosis.
Astrocyte activation by fibroblast growth factor-1 and motor neuron apoptosis: implications for amyotrophic lateral sclerosis
J Neurochem 2005 Apr;93(1):38-46
Patricia Cassina 1, Mariana Pehar 1 2, Marcelo R Vargas 2, Raquel Castellanos 1, Ana G Barbeito 1, Alvaro G Estévez 3, John A Thompson 4, Joseph S Beckman 5, Luis Barbeito 2
1 Departamento de Histología, Facultad de Medicina, Universidad de la República Montevideo, Uruguay 2 Departamento de Neurobiología Celular, Instituto de Investigaciones Biológicas Clemente Estable, Montevideo, Uruguay 3 Department of Physiology and Biophysics and 4 Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama, USA 5 Linus Pauling Institute, Environmental Health Sciences Center, Department of Biochemistry and Biophysics, Oregon State University, Corvallis, Oregon, USA
DOI: 10.1111/j.1471-4159.2004.02984.x
PMID: 15773903
Pubmed: https://pubmed.ncbi.nlm.nih.gov/15773903
Texto completo: https://onlinelibrary.wiley.com/doi/10.1111/j.1471-4159.2004.02984.x
Abstract:
Fibroblast growth factor-1 (FGF1 or acidic FGF) is highly expressed in motor neurons. FGF-1 is released from cells by oxidative stress, which might occur from SOD-1 aberrant function in amyotrophic lateral sclerosis (ALS). Although FGF-1 is known to be neuroprotective after spinal cord injury or axotomy, we found that FGF-1 could activate spinal cord astrocytes in a manner that decreased motor neuron survival in co-cultures. FGF-1 induced accumulation of the FGF receptor 1 (FGFR1) in astrocyte nuclei and potently stimulated nerve growth factor (NGF) expression and secretion. The FGFR1 tyrosine kinase inhibitor PD166866 prevented these effects. Previously, we have shown that NGF secretion by reactive astrocytes induces motor neuron apoptosis through a p75(NTR)-dependent mechanism. Embryonic motor neurons co-cultured on the top of astrocytes exhibiting activated FGFR1 underwent apoptosis, which was prevented by PD166866 or by adding either anti-NGF or anti-p75(NTR) neutralizing antibodies. In the degenerating spinal cord of mice carrying the ALS mutation G93A of Cu, Zn superoxide dismutase, FGF-1 was no longer localized only in the cytosol of motor neurons, while FGFR1 accumulated in the nuclei of reactive astrocytes. These results suggest that FGF-1 released by oxidative stress from motor neurons might have a role in activating astrocytes, which could in turn initiate motor neuron apoptosis in ALS through a p75(NTR)-dependent mechanism.
A role for astrocytes in motor neuron loss in amyotrophic lateral sclerosis
Brain Res Brain Res Rev 2004 Dec;47(1-3):263-74
Luis H Barbeito 1, Mariana Pehar 1 2, Patricia Cassina 2, Marcelo R Vargas 1, Hugo Peluffo 2, Liliana Viera 3, Alvaro G Estévez 3, Joseph S Beckman 4
1 Departamento de Neurobiologı́a Celular y Molecular, Instituto de Investigaciones Biológicas Clemente Estable, Avenida Italia 3318-CP 11600, Montevideo, Uruguay 2 Departamento de Histologı́a y Embriologı́a, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay 3 Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, AL, USA 4 Linus Pauling Institute, Environmental Health Sciences Center, Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR, USA
DOI: 10.1016/j.brainresrev.2004.05.003
PMID: 15572176
Pubmed: https://pubmed.ncbi.nlm.nih.gov/15572176
Texto completo: https://linkinghub.elsevier.com/retrieve/pii/S0165017304000700
Abstract:
A strong glial reaction typically surrounds the affected upper and lower motor neurons and degenerating descending tracts of ALS patients. Reactive astrocytes in ALS contain protein inclusions, express inflammatory makers such as the inducible forms of nitric oxide synthase (iNOS) and cyclooxygenase (COX-2), display nitrotyrosine immunoreactivity and downregulate the glutamate transporter EAAT2. In this review, we discuss the evidence sustaining an active role for astrocytes in the induction and propagation of motor neuron loss in ALS. Available evidence supports the view that glial activation could be initiated by proinflammatory mediators secreted by motor neurons in response to injury, axotomy or muscular pathology. In turn, reactive astrocytes produce nitric oxide and peroxynitrite, which cause mitochondrial damage in cultured neurons and trigger apoptosis in motor neurons. Astrocytes may also contribute to the excitotoxic damage of motor neurons by decreasing glutamate transport or actively releasing the excitotoxic amino acid. In addition, reactive astrocytes secrete pro-apoptotic mediators, such as nerve growth factor (NGF) or Fas-ligand, a mechanism that may serve to eliminate vulnerable motor neurons. The comprehensive understanding of the interactions between motor neurons and glia in ALS may lead to a more accurate theory of the pathogenesis of the disease.
A rapid method for fibronectin purification on nitrocellulose membranes suitable for tissue culture
J Biochem Biophys Methods 2004 May 31;59(2):139-43
Silvia Chifflet 1, Carmen Bolatto 2, Susana Tolosa 1
1 Departamento de Bioquı́mica, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800 Montevideo, Uruguay 2 Departamento de Histologı́a y Embriologı́a, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800 Montevideo, Uruguay
DOI: 10.1016/j.jbbm.2003.12.004
PMID: 15163525
Pubmed: https://pubmed.ncbi.nlm.nih.gov/15163525
Texto completo: https://linkinghub.elsevier.com/retrieve/pii/S0165022X03002161
Abstract:
We have developed a simple method for plasma fibronectin purification based on the well-known gelatin binding property of fibronectin. In this procedure we immobilize the melted gelatin to nitrocellulose membranes; these are then used to affinity-purify the fibronectin from the plasma sample. The fibronectin is eluted from the membrane by treatment with 8 M urea. The procedure described here gives a yield of up to 60% (from presumed fibronectin concentration) and the fibronectin obtained is homogeneous in SDS-PAGE and biologically active, as assessed by a cell migration assay. The method is rapid, simple, inexpensive, does not require the use of chromatographic equipment and is suitable for tissue culture applications.
Induction of motor neuron apoptosis by free 3-nitro-L-tyrosine
J Neurochem 2004 May;89(3):602-12
Hugo Peluffo 1, John J Shacka 7, Karina Ricart 7, C Gaston Bisig 3, Laura Martìnez-Palma 1, Otto Pritsch 2, Andrès Kamaid 1, Jason P Eiserich 4, John P Crow 5, Luis Barbeito 6, Alvaro G Estèvez 7
Departamentos de 1 Histologia y Embriología and 2 Bioquimica, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay 3 Departamento de Química Biologica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba , Córdoba, Argentina 4 Department of Internal Medicine and Human Physiology, University of California at Davis, Davis, California, USA 5 Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, LittleRock, Arkansas, USA 6 División Neurobiología Celular y Molecular, Instituto de Investigaciones Biológicas Clemente Estable, Montevideo, Uruguay 7 Department of Physiology and Biophysics, and 8 Department of Neurobiology and Center for Free Radical Biology, The University of Alabama at Birmingham, Birmingham, Alabama, USA
DOI: 10.1046/j.1471-4159.2004.02363.x
PMID: 15086517
Pubmed: https://pubmed.ncbi.nlm.nih.gov/15086517
Texto completo: https://onlinelibrary.wiley.com/doi/10.1046/j.1471-4159.2004.02363.x
Abstract:
Peroxynitrite-dependent tyrosine nitration has been postulated to be involved in motor neuron degeneration in amyotrophic lateral sclerosis (ALS). Evidence supporting this supposition includes the appearance of both free and protein-linked 3-nitro-l-tyrosine (nitrotyrosine) in both sporadic and familial ALS, as well as of increased free nitrotyrosine levels in the spinal cord of transgenic mice expressing ALS-linked superoxide dismutase mutants at symptom onset. Here we demonstrate that incubation with clinically relevant concentrations of nitrotyrosine induced apoptosis in motor neurons cultured with trophic factors. Nitrotyrosine was bound to proteins, but it was not incorporated into alpha-tubulin, as previously demonstrated for other cell types. Neither inhibition of nitric oxide production nor scavenging of superoxide and peroxynitrite prevented increases in cell nitrotyrosine immunoreactivity or motor neuron death, suggesting that these effects are not due to the endogenous formation of reactive nitrogen species. In contrast, some populations of astrocytes incorporated nitrotyrosine into alpha-tubulin, but free nitrotyrosine had no effect on the viability and phenotype of astrocytes in culture, as evaluated by glial fibrillary acidic protein immunoreactivity, cell growth and morphology. Co-culture of motor neurons on astrocyte monolayers delayed, but did not prevent, nitrotyrosine-induced motor neuron death. These results suggest that free nitrotyrosine may play a role in the induction of motor neuron apoptosis in ALS.
Environment temperature affects cell proliferation in the spinal cord and brain of juvenile turtles
J Exp Biol 2003 Sep;206(Pt 17):3085-93
Milka Radmilovich 1, Anabel Fernández 2 3, Omar Trujillo-Cenóz 2 3
1 Departamento de Histología y Embriología, Facultad de Medicina, Montevideo, Uruguay ZC 11800 2 Laboratorio de Neuroanatomía Comparada, Instituto de Investigaciones Biológicas Clemente Estable, Avda. Italia 3318,Montevideo, Uruguay ZC 11600 3 Unidad Asociada a la Facultad de Ciencias, Montevideo, Uruguay ZC 11600
DOI: 10.1242/jeb.00515
PMID: 12878675
Pubmed: https://pubmed.ncbi.nlm.nih.gov/12878675
Texto completo: https://journals.biologists.com/jeb/article-lookup/doi/10.1242/jeb.00515
Abstract:
The spinal cords and brains--comprising dorsal cortex (DC), medial cortex (MC) and diencephalon (Dien)--of juvenile turtles acclimated to warm temperature [27-30 degrees C; warm-acclimated turtles (WATs)] revealed higher density values of bromodeoxyuridine-labeled cells (BrdU-LCs) than those acclimated to a cooler environment [5-14 degrees C; cold-acclimated turtles (CATs)]. Both populations were under the influence of the seasonal daily light-dark rhythms. Pronounced differences between WATs and CATs (independent t-test; confidence level, P<0.01) were found in the central area of the spinal gray matter and in the ependymal epithelium lining the brain ventricles. Forebrain regions (DC, MC and Dien) also revealed significant differences between WATs and CATs (independent t-test; confidence level, P<0.01-0.05). Unexplored biological clocks that may be affecting cell proliferation were equalized by performing paired experiments involving one WAT and one CAT. Both animals were injected on the same day at the same time and both were sacrificed 24 h later. These experiments confirmed that a warm environment increased cell proliferation in the CNS of turtles. Double- and triple-labeling experiments involving anti-BrdU antibody together with anti-glial protein antibodies revealed that temperature modulates not only cell populations expressing glial markers but also other cells that do not express them. As expected, in the case of short post-injection (BrdU) surviving time points, no cells were found colabeling for BrdU and NeuN (neuronal marker). The probable direct effect of temperature on the cell division rate should be analyzed together with potential indirect effects involving increased motor activity and increased food intake. The fate of the increased BrdU-LCs (death, permanence as progenitor cells or differentiation following neuronal or glial lines) remains a matter for further investigation. Results are discussed in the light of current opinions concerned with post-natal neurogenesis in vertebrates.
Synaptic activity modifies the levels of Dorsal and Cactus at the neuromuscular junction of Drosophila
J Neurobiol 2003 Feb 15;54(3):525-36
Carmen Bolatto 1 , Silvia Chifflet 2 , Aram Megighian 3 , Rafael Cantera 4
1 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Uruguay. 2 Departamento de Bioquímica, Facultad de Medicina, Universidad de la República, Uruguay 3 Aram Megighian, Department of Human Anatomy and Physiology, University of Padova, Italy 4 Zoology Department, Svante Arrheniusväg 14, Stockholm University, Stockholm, Sweden
DOI: 10.1002/neu.10179
PMID: 12532402
Pubmed: https://pubmed.ncbi.nlm.nih.gov/12532402
Texto completo: https://doi.org/10.1002/neu.10179
Abstract:
The Drosophila Rel transcription factor Dorsal and its inhibitor Cactus participate in a signal transduction pathway involved in several biologic processes, including embryonic pattern formation, immunity, and muscle development. In contrast with embryonic muscle, where Dorsal is reportedly absent, this protein and Cactus accumulates in the neuromuscular junctions in the muscle of both larvae and adults. The phenotype of homozygous dorsal mutant larvae suggested that Dorsal and Cactus maybe necessary for normal function and maintenance of the neuromuscular system. Here we investigate if these proteins can respond to synaptic activity. Using larval body wall preparations and antibodies specific for Dorsal or Cactus we show that the amount of these proteins at the neuromuscular junction is substantially decreased after electrical stimulation of the nerves or incubation in glutamate, the principal transmitter in this type of synapse. The specificity of the response was tested with a glutamate receptor antagonist (argiotoxin 636). Because the effect can be reproduced using a calcium ionophore (ionomycin treatment) as well as blocked by the inhibition of the muscle ryanodine receptor (tetracaine treatment), the involvement of calcium in this process seems likely. We also observed that the inhibition of the calcium dependent protein phosphatase calcineurin prevents the effect of glutamate on the fluorescence for Dorsal and Cactus, suggesting its participation in a signal transduction cascade that may activate Dorsal in the muscle independently of Toll. Our results are consistent with a novel function of the Rel factor Dorsal in a molecular pathway turned on by neural activity and/or contractile activity.
Involvement of nitric oxide on kainate-induced toxicity in oligodendrocyte precursors
Neurotox Res 2003 2003;5(6):399-406
Laura Martinez-Palma 1, Mariana Pehar 2, Patricia Cassina 1, Hugo Peluffo 1, Raquel Castellanos 1, Gabriel Anesetti 1, Joseph S Beckman3 , Luis Barbeito 2
1 Departamento de Histologia, Facultad de Veterinaria y Facultad de Medicina, Universidad de la República, 11600, Montevideo, Uruguay Laura Martinez-Palma, Patricia Cassina, Hugo Peluffo, Raquel Castellanos & Gabriel Anesetti 2 Departamento de Neurobiología Celular, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, 11600, Montevideo, Uruguay Mariana Pehar & Luis Barbeito 3 Linus Pauling Institute, Department of Biochemistry and Biophysics, Oregon State University, 97331, Corvallis, OR, USA Joseph S. Beckman
DOI: 10.1007/BF03033168
PMID: 14715442
Pubmed: https://pubmed.ncbi.nlm.nih.gov/14715442
Texto completo: https://dx.doi.org/10.1007/BF03033168
Abstract:
The vulnerability of oligodendrocytes to excitatory amino acids may account for the pathology of white matter occurring following hypoxia/ischemia or autoimmune attack. Here, we examined the vulnerability of immature oligodendrocytes (positively labeled by galactocerobroside-C and not expressing myelin basic protein) from neonatal rat spinal cord to kainate, an agonist of excitatory amino acid receptors that induces long-lasting inward currents in immature oligodendrocytes. In particular, we studied whether kainate toxicity was linked to the endogenous production of nitric oxide. We found cultured oligodendrocytes to be highly sensitive to 24-48 h exposure to 0.5-1 mM kainate. The toxin induced striking morphological changes in oligodendrocytes, characterized by the disruption of the process network around the cell body and the growth of one or two long, thick and non-branched processes. A longer exposure to kainate resulted in massive death of oligodendrocytes, which was prevented by 6,7, dinitroquinoxaline-2,3-dione (DNQX) (30 micro M), the antagonist of AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic/kainate receptors. Remarkably, we found that those oligodendrocytes displaying bipolar morphology following kainate exposure, also expressed the inducible form of nitric oxide synthase (iNOS) and nitrotyrosine immunoreactivity, suggesting that peroxynitrite could be formed by the reaction of nitric oxide with superoxide. Moreover, kainate toxicity was significantly prevented by addition of the NOS inhibitor nitro-L-arginine methyl ester (L-NAME), further suggesting that nitric oxide-derived oxidants contribute to excitotoxic mechanisms in immature oligodendrocytes.