Publicaciones por año
Publicaciones del Departamento de Histología y Embriología
A Note of Caution: Gramicidin Affects Signaling Pathways Independently of Its Effects on Plasma Membrane Conductance
Biomed Res Int 2021 2021:2641068
Frances Evans 1 , Julio A Hernández 2 , Federico Cabo 3 , Silvia Chifflet 3
1 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800 Montevideo, Uruguay. 2 Sección Biofísica, Facultad de Ciencias, Universidad de la República, Iguá s/n esq. Mataojo, 11400 Montevideo, Uruguay. 3 Departamento de Bioquímica, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800 Montevideo, Uruguay.
DOI: 10.1155/2021/2641068
PMID: 34722759
Pubmed: https://pubmed.ncbi.nlm.nih.gov/34722759
Texto completo: https://doi.org/10.1155/2021/2641068
Abstract:
Gramicidin is a thoroughly studied cation ionophore widely used to experimentally manipulate the plasma membrane potential (PMP). In addition, it has been established that the drug, due to its hydrophobic nature, is capable of affecting the organization of membrane lipids. We have previously shown that modifications in the plasma membrane potential of epithelial cells in culture determine reorganizations of the cytoskeleton. To elucidate the molecular mechanisms involved, we explored the effects of PMP depolarization on some putative signaling intermediates. In the course of these studies, we came across some results that could not be interpreted in terms of the properties of gramicidin as an ionic channel. The purpose of the present work is to communicate these results and, in general, to draw attention to the fact that gramicidin effects can be misleadingly attributed to its ionic or electrical properties. In addition, this work also contributes with some novel findings of the modifications provoked on the signaling intermediates by PMP depolarization and hyperpolarization.
Neonatal androgenization in rats affects oocyte maturation
Reprod Sci 2021 28(10):2799-2806
Gabriel Anesetti 1 , Rebeca Chávez-Genaro 2
1 Laboratorio de Biología de la Reproducción, Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. ganesett@fmed.edu.uy. 2 Laboratorio de Biología de la Reproducción, Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay
DOI: 10.1007/s43032-021-00559-6
PMID: 33825168
Pubmed: https://pubmed.ncbi.nlm.nih.gov/33825168
Texto completo: https://doi.org/10.1007/s43032-021-00559-6
Abstract:
Androgens are relevant in order to achieve a normal growth and maturation of the follicle and oocyte, since both excess and absence of androgens may affect the correct ovarian function. The current study analyzes the impact of neonatal androgenization in the first ovulation and oocyte maturation in response to exogenous gonadotrophin stimulation. Neonatal rats were daily treated with testosterone, dihydrotestosterone, or vehicle during follicle assembly period (days 1 to 5). At juvenile period, rats were stimulated sequentially with PMSG and hCG. Ovulation, ovarian histology, hormonal milieu, morphological characteristics of meiotic spindle, and in vitro fertilization rate in oocytes were analyzed. Our data shows that oocytes from androgenized rats displayed a major proportion of aberrant spindles and altered meiotic advance that control animals. These alterations were accompanied with an increase in both fertilization rate and aberrant embryos after 48 h of culture. Our findings showed a direct impact of neonatal androgens on oocyte development; their effects may be recognized at adulthood, supporting the idea of a programming effect exerted by neonatal androgens. These results could be relevant to explain the low fertility rate seen in polycystic ovary syndrome patients after in vitro fertilization procedures.
Dissection of Single Skeletal Muscle Fibers for Immunofluorescent and Morphometric Analyses of Whole-Mount Neuromuscular Junctions
J Vis Exp 2021 (174)
Carmen Bolatto # 1 , Silvia Olivera-Bravo # 2 , Sofía Cerri 3
1 Developmental Biology Laboratory, Histology and Embryology Department, Faculty of Medicine, Universidad de la República; cbolatto@fmed.edu.uy. 2 Cell and Molecular Neurobiology Laboratory, Clemente Estable Biology Research Institute (IIBCE), Ministerio de Educación y Cultura. 3 Developmental Biology Laboratory, Histology and Embryology Department, Faculty of Medicine, Universidad de la República; Cell and Molecular Neurobiology Laboratory, Clemente Estable Biology Research Institute (IIBCE), Ministerio de Educación y Cultura. # Contributed equally.
DOI: 10.3791/62620
PMID: 34459813
Pubmed: https://pubmed.ncbi.nlm.nih.gov/34459813
Texto completo: https://doi.org//10.3791/62620
Abstract:
The neuromuscular junction (NMJ) is a specialized point of contact between the motor nerve and the skeletal muscle. This peripheral synapse exhibits high morphological and functional plasticity. In numerous nervous system disorders, NMJ is an early pathological target resulting in neurotransmission failure, weakness, atrophy, and even in muscle fiber death. Due to its relevance, the possibility to quantitatively assess certain aspects of the relationship between NMJ components can help to understand the processes associated with its assembly/disassembly. The first obstacle when working with muscles is to gain the technical expertise to quickly identify and dissect without damaging their fibers. The second challenge is to utilize high-quality detection methods to obtain NMJ images that can be used to perform quantitative analysis. This article presents a step-by-step protocol for dissecting extensor digitorum longus and soleus muscles from rats. It also explains the use of immunofluorescence to visualize pre and postsynaptic elements of whole-mount NMJs. Results obtained demonstrate that this technique can be used to establish the microscopic anatomy of the synapsis and identify subtle changes in the status of some of its components under physiological or pathological conditions.
Decline of semen quality over the last 30 years in Uruguay
Basic Clin Androl 2021 31(1):8
Lucía Rosa-Villagrán 1 , Natalibeth Barrera 2 3 , José Montes 2 , Carlos Riso 1 , Rossana Sapiro 4
1 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800, Montevideo, Uruguay. 2 Laboratorio de Andrología, Fertilab Laboratorio de Análisis Clínicos, Montevideo, Uruguay. 3 Laboratorio de FIV, Centro de Esterilidad Montevideo (CEM), Montevideo, Uruguay. 4 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Gral. Flores 2125, 11800, Montevideo, Uruguay. rossanasapiro@fmed.edu.uy.
DOI: 10.1186/s12610-021-00128-6
PMID: 33952196
Pubmed: https://pubmed.ncbi.nlm.nih.gov/33952196
Texto completo: https://bacandrology.biomedcentral.com/articles/10.1186/s12610-021-00128-6
Abstract:
Background: Over the last years, there has been an increasing concern about a global decline in men's fertility. Specifically, some evidence indicates that sperm quality has decreased over the last years. However, reports showing the changes in sperm quality with time are inconsistent. Part of the contradictions between studies is attributed to geographical differences. Surprisingly, few studies include data from South American countries, creating a bias in the conclusions. This study aims to determine how sperm quality has evolved over the past 30 years in Uruguay. For this purpose, 317 medical records from allegedly healthy sperm donor candidates, aged between 18 and 36 years old, who voluntarily requested to be considered as sperm donors between 1988 and 2019, were analyzed. The studied variables were the following sperm parameters: semen volume, sperm cell concentration, total sperm number, progressive motility, vitality, and sperm morphology. A correlative statistical analysis was performed between seminal parameter values and the year data were collected.
Results: We found a statistically significant decrease in sperm concentration and normal sperm morphology during the studied period. There was no decrease in vitality, seminal volume, and total progressive motility. Semen parameters were not associated with tobacco, drugs, or alcohol consumption.
Conclusions: We conclude that the sperm quality of donor candidates in Uruguay decreased during this period. Further studies should be carried out to verify the occurrence of this phenomenon in the general population and find its possible causes.
Distinct neuron phenotypes may serve object feature sensing in the electrosensory lobe of Gymnotus omarorum
J Exp Biol 2021 224(9):jeb242242
Javier Nogueira 1 2 , María E Castelló 3 , Carolina Lescano 3 , Ángel A Caputi 1
1 Departamento Neurociencias Integrativas y Computacionales, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, Uruguay. 2 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Av. Gral Flores 2515, Montevideo, Uruguay. 3 Laboratorio Desarrollo y Evolución Neural, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, Uruguay.
DOI: 10.1242/jeb.242242
PMID: 33707195
Pubmed: https://pubmed.ncbi.nlm.nih.gov/33707195
Texto completo: https://journals.biologists.com/jeb/article-lookup/doi/10.1242/jeb.242242
Abstract:
Early sensory relay circuits in the vertebrate medulla often adopt a cerebellum-like organization specialized for comparing primary afferent inputs with central expectations. These circuits usually have a dual output, carried by center ON and center OFF neurons responding in opposite ways to the same stimulus at the center of their receptive fields. Here, we show in the electrosensory lateral line lobe of Gymnotiform weakly electric fish that basilar pyramidal neurons, representing 'ON' cells, and non-basilar pyramidal neurons, representing 'OFF' cells, have different intrinsic electrophysiological properties. We used classical anatomical techniques and electrophysiological in vitro recordings to compare these neurons. Basilar neurons are silent at rest, have a high threshold to intracellular stimulation, delayed responses to steady-state depolarization and low pass responsiveness to membrane voltage variations. They respond to low-intensity depolarizing stimuli with large, isolated spikes. As stimulus intensity increases, the spikes are followed by a depolarizing after-potential from which phase-locked spikes often arise. Non-basilar neurons show a pacemaker-like spiking activity, smoothly modulated in frequency by slow variations of stimulus intensity. Spike-frequency adaptation provides a memory of their recent firing, facilitating non-basilar response to stimulus transients. Considering anatomical and functional dimensions, we conclude that basilar and non-basilar pyramidal neurons are clear-cut, different anatomo-functional phenotypes. We propose that, in addition to their role in contrast processing, basilar pyramidal neurons encode sustained global stimuli such as those elicited by large or distant objects while non-basilar pyramidal neurons respond to transient stimuli due to movement of objects with a textured surface.
A nuclear fluorescent dye identifies pericytes at the neurovascular unit
J Neurochem 2021 157(4):1377-1391
Sandra P Mai-Morente 1 , Virginia M Marset 1 , Fabiana Blanco 2 , Eugenia E Isasi 3 , Verónica Abudara 1
1 Departamento de Fisiología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 2 Departamento de Biofísica, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 3 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay.
DOI: 10.1111/jnc.15193
PMID: 32974913
Pubmed: https://pubmed.ncbi.nlm.nih.gov/32974913
Texto completo: https://doi.org/10.1111/jnc.15193
Abstract:
Perivascular pericytes are key regulators of the blood-brain barrier, vascular development, and cerebral blood flow. Deciphering pericyte roles in health and disease requires cellular tracking; yet, pericyte identification remains challenging. A previous study reported that the far-red fluorophore TO-PRO-3 (642/661), usually employed as a nuclear dye in fixed tissue, was selectively captured by live pericytes from the subventricular zone. Herein, we validated TO-PRO-3 as a specific pericyte tracer in the nervous system (NS). Living pericytes from ex vivo murine hippocampus, cortex, spinal cord, and retina robustly incorporated TO-PRO-3. Classical pericyte immunomarkers such as chondroitin sulphate proteoglycan neuron-glial antigen 2 (NG2) and platelet-derived growth factor receptor beta antigen (PDGFrβ) and the new pericyte dye NeuroTrace 500/525 confirmed cellular specificity of dye uptake. The TO-PRO-3 signal enabled quantification of pericytes density and morphometry; likewise, TO-PRO-3 labeling allowed visualization of pericytes associated with other components of the neurovascular unit. A subset of TO-PRO-3 stained cells expressed the contractile protein α-SMA, indicative of their ability to control the capillary diameter. Uptake of TO-PRO-3 was independent of connexin/pannexin channels but was highly sensitive to temperature and showed saturation, suggesting that a yet unidentified protein-mediated active transport sustained dye incorporation. We conclude that TO-PRO-3 labeling provides a reliable and simple tool for the bioimaging of pericytes in the murine NS microvasculature.
Glial Metabolic Reprogramming in Amyotrophic Lateral Sclerosis
Neuroimmunomodulation 2021 28(4):204-212
Patricia Cassina 1 2 , Ernesto Miquel 1 2 , Laura Martínez-Palma 1 2 , Adriana Cassina 2 3
1 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 2 Centro de Investigaciones Biomédicas (CEINBIO), Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 3 Departamento de Bioquímica, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay.
DOI: 10.1159/000516926
PMID: 34175843
Pubmed: https://pubmed.ncbi.nlm.nih.gov/34175843
Texto completo: https://doi.org/10.1159/000516926
Abstract:
ALS is a human neurodegenerative disorder that induces a progressive paralysis of voluntary muscles due to motor neuron loss. The causes are unknown, and there is no curative treatment available. Mitochondrial dysfunction is a hallmark of ALS pathology; however, it is currently unknown whether it is a cause or a consequence of disease progression. Recent evidence indicates that glial mitochondrial function changes to cope with energy demands and critically influences neuronal death and disease progression. Aberrant glial cells detected in the spinal cord of diseased animals are characterized by increased proliferation rate and reduced mitochondrial bioenergetics. These features can be compared with cancer cell behavior of adapting to nutrient microenvironment by altering energy metabolism, a concept known as metabolic reprogramming. We focus on data that suggest that aberrant glial cells in ALS undergo metabolic reprogramming and profound changes in glial mitochondrial activity, which are associated with motor neuron death in ALS. This review article emphasizes on the association between metabolic reprogramming and glial reactivity, bringing new paradigms from the area of cancer research into neurodegenerative diseases. Targeting glial mitochondrial function and metabolic reprogramming may result in promising therapeutic strategies for ALS.
Sex-dependent role of CD300f immune receptor in generalized anxiety disorder
Brain Behav Immun Health 2020 11:100191
Fernanda N Kaufmann 1 , Natalia Lago 2 , Daniela Alí-Ruiz 2 , Karen Jansen 3 , Luciano D M Souza 3 , Ricardo A Silva 3 , Diogo R Lara 4 , Gabriele Ghisleni 3 , Hugo Peluffo 2 5 , Manuella P Kaster 6
1 Department of Psychiatry and Neuroscience, Faculty of Medicine and CERVO Brain Research Center, Université Laval, Quebec City, Canada. 2 Neuroinflammation and Gene Therapy Laboratory, Institut Pasteur de Montevideo, Montevideo, Uruguay. 3 Department of Life and Health Sciences, Catholic University of Pelotas, Rio Grande do Sul, Brazil. 4 Department of Cellular and Molecular Biology, Pontifical Catholic University of Rio Grande do Sul, Porto Alegre, Brazil. 5 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 6 Department of Biochemistry, Federal University of Santa Catarina, Florianópolis, Santa Catarina, Brazil.
DOI: 10.1016/j.bbih.2020.100191
PMID: 34589728
Pubmed: https://pubmed.ncbi.nlm.nih.gov/34589728
Texto completo: https://linkinghub.elsevier.com/retrieve/pii/S2666-3546(20)30156-3
Abstract:
Generalized Anxiety Disorder (GAD) presents a high prevalence in the population, leading to distress and disability. Immune system alterations have been associated with anxiety-related behaviors in rodents and GAD patients. CD300f immune receptors are highly expressed in microglia and participate not only in the modulation of immune responses but also in pruning and reshaping synapses. It was recently demonstrated that CD300f might be influential in the pathogenesis of depression in a sex-dependent manner. Here, we evaluated the role of CD300f immune receptor in anxiety, using CD300f knockout mice (CD300f-/-) and patients with GAD. We observed that male CD300f-/- mice had numerous behavioral changes associated with a low-anxiety phenotype, including increased open field central locomotion and rearing behaviors, more exploration in the open arms of the elevated plus-maze test, and decreased latency to eat in the novelty suppressed feeding test. In a cross-sectional population-based study, including 1111 subjects, we evaluated a common single-nucleotide polymorphism rs2034310 (C/T) in the cytoplasmatic tail of CD300f gene in individuals with GAD. Notably, we observed that the T allele of the rs2034310 polymorphism conferred protection against GAD in men, even after adjusting for confounding variables. Overall, our data demonstrate that CD300f immune receptors are involved in the modulation of pathological anxiety behaviors in a sex-dependent manner. The biological basis of these sex differences is still poorly understood, but it may provide significant clues regarding the neuropathophysiological mechanisms of GAD and can pave the way for future specific pharmacological interventions.
Mitochondrial bioenergetics, glial reactivity, and pain-related behavior can be restored by dichloroacetate treatment in rodent pain models
Pain 2020 161(12):2786-2797
Valentina Lagos-Rodríguez 1 , Laura Martínez-Palma 1 , Soledad Marton 1 , Ernesto Miquel 1 , Ricardo Escobar-Pintos 1 , Adriana Cassina 2 , Natalia Lago 3 , Patricia Cassina 1
1 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 2 Departamento de Bioquímica, Centro de Investigaciones Biomédicas (CEINBIO), Facultad de Medicina, Universidad de la República, Montevideo, Uruguay. 3 Neuroinflammation and Gene Therapy Laboratory, Institut Pasteur de Montevideo, Montevideo, Uruguay.
DOI: 10.1097/j.pain.0000000000001992
PMID: 32658145
Pubmed: https://pubmed.ncbi.nlm.nih.gov/32658145
Texto completo: https://doi.org/10.1097/j.pain.0000000000001992
Abstract:
Glial reactivity in the dorsal horn of the spinal cord is a hallmark in most chronic pain conditions. Neuroinflammation-associated reactive glia, in particular astrocytes, have been shown to exhibit reduced mitochondrial respiratory function. Here, we studied the mitochondrial function at the lumbar spinal cord tissue from complete Freund's adjuvant-induced inflammatory pain rat and chronic constriction injury mouse models by high-resolution respirometry. A significant decrease in mitochondrial bioenergetic parameters at the injury-related spinal cord level coincided with highest astrocytosis. Oral administration of dichloroacetate (DCA) significantly increased mitochondrial respiratory function by inhibiting pyruvate dehydrogenase kinase and decreased glial fibrillary acidic protein and Iba-1 immunoreactivity in spinal cord. Importantly, DCA treatment significantly reduced the ipsilateral pain-related behavior without affecting contralateral sensitivity in both pain models. Our results indicate that mitochondrial metabolic modulation with DCA may offer an alternative therapeutic strategy to alleviate chronic and persistent inflammatory pain.
Reorganization of perineuronal nets in the medial Preoptic Area during the reproductive cycle in female rats
Sci Rep 2020 10(1):5479
Natalia Uriarte 1 , Marcela Ferreño 1 , Diego Méndez 2 , Javier Nogueira 3
1 Laboratorio de Neurociencias, Facultad de Ciencias, Universidad de la República, Montevideo, 11400, Uruguay. 2 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, 11800, Uruguay. 3 Departamento de Histología y Embriología, Facultad de Medicina, Universidad de la República, Montevideo, 11800, Uruguay. nogueira@fmed.edu.uy.
DOI: 10.1038/s41598-020-62163-z
PMID: 32214157
Pubmed: https://pubmed.ncbi.nlm.nih.gov/32214157
Texto completo: https://doi.org/10.1038/s41598-020-62163-z
Abstract:
Perineuronal nets (PNNs) are aggregations of extracellular matrix associated with specific neuronal populations in the central nervous system, suggested to play key roles in neural development, synaptogenesis and experience-dependent synaptic plasticity. Pregnancy and lactation are characterized by a dramatic increase in neuroplasticity. However, dynamic changes in the extracellular matrix associated with maternal circuits have been mostly overlooked. We analyzed the structure of PNNs in an essential nucleus of the maternal circuit, the medial preoptic area (mPOA), during the reproductive cycle of rats, using the Wisteria floribunda (WFA) label. PNNs associated to neurons in the mPOA start to assemble halfway through gestation and become highly organized prior to parturition, fading through the postpartum period. This high expression of PNNs during pregnancy appears to be mediated by the influence of estrogen, progesterone and prolactin, since a hormonal simulated-gestation treatment induced the expression of PNNs in ovariectomized females. We found that PNNs associated neurons in the mPOA express estrogen receptor α and progesterone receptors, supporting a putative role of reproductive hormones in the signaling mechanisms that trigger the assembly of PNNs in the mPOA. This is the first report of PNNs presence and remodeling in mPOA during adulthood induced by physiological variables.