RESUMEN
The hypothalamo-neurohypophysial system (HNS) consisting of arginine vasopressin (AVP) and oxytocin (OXT) magnocellular neurons shows the structural plasticity including the rearrangement of synapses, dendrites, and neurovascular contacts during chronic physiological stimulation. In this study, we examined the remodeling of chondroitin sulfate proteoglycans (CSPGs), main extracellular matrix (ECM), in the HNS after salt loading known as a chronic stimulation to cause the structural plasticity. In the supraoptic nucleus (SON), confocal microscopic observation revealed that the immunoreactivity of 6B4 proteoglycans (PG) was observed mainly at AVP-positive magnocellular neurons but that of neurocan was seen chiefly at OXT-positive magnocellular neurons. The immunoreactivity of phosphacan and aggrecan was seen at both AVP- and OXT-positive magnocellular neurons. Electron microscopic observation further showed that the immunoreactivity of phosphacan and neurocan was observed at astrocytic processes to surround somata, dendrites, and terminals, but not synaptic junctions. In the neurohypophysis (NH), the immunoreactivity of phosphacan, 6B4 PGs, and neurocan was observed at AVP-positive magnocellular terminals, but the reactivity of Wisteria floribunda agglutinin lectin was seen at OXT-positive ones. The immunoreactivity of versican was found at microvessel and that of aggrecan was not detected in the NH. Quantitative morphometrical analysis showed that the chronic physiological stimulation by 7-day salt loading decreased the level of 6B4 PGs in the SON and the level of phosphacan, 6B4 PGs, and neurocan in the NH. These results suggest that the extracellular microenvironment of CSPGs is different between AVP and OXT magnocellular neurons and activity-dependent remodeling of CSPGs could be involved in the structural plasticity of the HNS.
Asunto(s)
Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Matriz Extracelular/metabolismo , Sistema Hipotálamo-Hipofisario/metabolismo , Hipotálamo/metabolismo , Plasticidad Neuronal/fisiología , Neurohipófisis/metabolismo , Agrecanos/metabolismo , Animales , Arginina Vasopresina/metabolismo , Sistema Hipotálamo-Hipofisario/ultraestructura , Hipotálamo/ultraestructura , Inmunohistoquímica , Masculino , Microscopía Confocal , Microscopía Electrónica de Transmisión , Neurocano , Plasticidad Neuronal/efectos de los fármacos , Oxitocina/metabolismo , Neurohipófisis/ultraestructura , Lectinas de Plantas/metabolismo , Terminales Presinápticos/metabolismo , Ratas , Ratas Wistar , Proteínas Tirosina Fosfatasas Clase 5 Similares a Receptores/metabolismo , Receptores N-Acetilglucosamina/metabolismo , Cloruro de Sodio/farmacología , Núcleo Supraóptico/metabolismo , Núcleo Supraóptico/ultraestructura , Versicanos/metabolismo , Equilibrio Hidroelectrolítico/fisiologíaRESUMEN
We used immunohistochemistry to examine possible molecular interactions between the subplate and growing thalamocortical axons in rat fetuses. In the cortical anlage of embryonic day 16 (E16), the subplate first appeared below the cortical plate. Among chondroitin sulfate proteoglycans, phosphacan was uniformly distributed throughout the cortical wall, whereas neurocan was localized only in the subplate at E16. Neural cell adhesion molecules, NCAM-H, TAG-1, and L1, were detected in the cortical anlage. Both cortical neurons and growing axons were diffusely immunopositive for NCAM-H, and TAG-1 immunoreactivity was found on immature neurons and cortical efferent axons but not on thalamocortical axons. L1 immunoreactivity was specifically localized on the growing thalamocortical axons. When the locations of neurocan and L1 were compared in the developing cortex, L1-bearing axons were found to extend to neurocan-immunopositive regions; neurocan immunoreactivity was intense in the subplate at E16, when small numbers of L1-immunoreactive thalamocortical axons began to invade the cortex. At E17, many L1-positive axons were observed in the subplate that expressed neurocan specifically. Double immunostaining showed that L1-positive axons and neurocan immunoreactivity overlapped in the subplate at E17. After E18, neurocan expression gradually extended to the lower part of the cortical plate; it extended to the entire cortex by E21, 1 day before birth. By E21, L1-bearing axons had invaded the lower part of the cortical plate. The present study demonstrated that the neurocan expression precedes growth of L1-bearing thalamocortical afferent fibers. Because neurocan can bind to L1 molecule in vitro, these results suggest that neurocan and L1 play some important roles in pathfinding of the thalamocortical afferent fibers during rat corticogenesis.
Asunto(s)
Moléculas de Adhesión Celular Neuronal , Corteza Cerebral/embriología , Proteoglicanos Tipo Condroitín Sulfato/biosíntesis , Regulación del Desarrollo de la Expresión Génica , Glicoproteínas de Membrana/biosíntesis , Proteínas del Tejido Nervioso/biosíntesis , Neuronas/fisiología , Tálamo/embriología , Animales , Axones/fisiología , Corteza Cerebral/citología , Corteza Cerebral/metabolismo , Proteoglicanos Tipo Condroitín Sulfato/análisis , Contactina 2 , Desarrollo Embrionario y Fetal , Inmunohistoquímica/métodos , Lectinas Tipo C , Complejo de Antígeno L1 de Leucocito , Glicoproteínas de Membrana/análisis , Proteínas del Tejido Nervioso/análisis , Moléculas de Adhesión de Célula Nerviosa/análisis , Moléculas de Adhesión de Célula Nerviosa/biosíntesis , Neurocano , Neuronas/citología , Ratas , Ratas Sprague-Dawley , Tálamo/citología , Tálamo/metabolismoRESUMEN
Two rare cases of strabismus resulting from contracture of the extraocular rectus muscles after retrobulbar anesthesia for cataract surgery are described. Clinical signs in both cases suggested that the development of the impaired function of the lateral and superior rectus muscles followed the same pattern: initial stimulation followed by paretic and restrictive stages. Abnormal enlargement of the muscles was identified by computed tomography (CT) and magnetic resonance imaging (MRI). The data indicate that the strabismus was the result of direct injection of anesthetics into the rectus muscle.
Asunto(s)
Anestesia Local/efectos adversos , Anestésicos Locales/efectos adversos , Músculos Oculomotores/efectos de los fármacos , Estrabismo/inducido químicamente , Anestésicos Locales/administración & dosificación , Extracción de Catarata , Diagnóstico Diferencial , Femenino , Humanos , Inyecciones , Imagen por Resonancia Magnética , Persona de Mediana Edad , Músculos Oculomotores/diagnóstico por imagen , Músculos Oculomotores/patología , Órbita , Complicaciones Posoperatorias , Estrabismo/diagnóstico , Tomografía Computarizada por Rayos XRESUMEN
Monoclonal antibodies were raised to membrane-bound proteoglycans derived from rat brain, and four monoclonal antibodies that recognized a 150-kDa chondroitin sulfate proteoglycan with a core glycoprotein of 120 kDa were obtained. Immunohistological study revealed that the proteoglycan was associated with developing neurons. We screened rat brain cDNA libraries using the four monoclonal antibodies and isolated overlapping cDNA clones that encoded the entire core protein of 514 amino acids plus a 30-residue signal peptide. The deduced amino acid sequence suggested an integral membrane protein divided into five structurally different domains: an N-terminal domain to which chondroitin sulfate chains might be attached, a basic amino acid cluster consisting of seven arginine and two lysine residues, a cysteine-containing domain, a membrane-spanning segment, and a C-terminal cytoplasmic domain of 95 amino acids. On Northern blots, the cDNA hybridized with a single mRNA of 3.1 kilobases that was detectable in brains of neonatal and adult rats but not in kidney, liver, lung, and muscle of either. The sequence of the proteoglycan did not exhibit significant homology to any other known protein, indicating that the proteoglycan, designated neuroglycan C, is a novel integral membrane proteoglycan.
Asunto(s)
Encéfalo/metabolismo , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Proteínas de la Matriz Extracelular , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Agrecanos , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales , Secuencia de Bases , Proteoglicanos Tipo Condroitín Sulfato/genética , Proteoglicanos Tipo Condroitín Sulfato/inmunología , Clonación Molecular , Cartilla de ADN/genética , ADN Complementario/genética , Lectinas Tipo C , Proteínas de la Membrana/genética , Proteínas de la Membrana/inmunología , Datos de Secuencia Molecular , Estructura Molecular , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/inmunología , Proteoglicanos/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Distribución TisularRESUMEN
The effects of soluble chondroitin sulfate proteoglycans (CSPGs) purified from the rat brain on proliferation of and neurite outgrowth from PC12D cells (Katoh-Semba et al., J Neurosci Res 17:36, 1987) were investigated. When PC12D cells are cultured under standard conditions, they proliferate with a doubling time of about 2 days, irrespective of the presence or absence of NGF. However, the addition of a mixture of several types of purified soluble brain CSPG (50 nmol uronic acid/ml) to the culture medium prevented the increase in the number of PC12D cells as well as the nerve growth factor (NGF)-induced neurite extension. The dose for 50% inhibition (ID50) was 1.6 nmol/ml for cell proliferation and 2.7 nmol/ml for neurite elongation. The increase in cell number seemed to stop around 6 h after exposure to culture medium supplemented with brain-derived CSPGs, and even substratum-attached CSPGs were able to exert such inhibitory effects. Only brain-type CSPGs, not a cartilage-derived CSPG (PGH) or a hyaluronate-binding PGH, had such inhibitory effects. Furthermore, these inhibitory activities were associated only with the core proteins of brain-derived CSPGs, and not with polysaccharide chains from brain-derived CSPGs. Incorporation of [3H]thymidine into DNA did not decrease for at least the first 12 h. Consequently, the amount of DNA per cell after 48 h of culture was about twofold higher in cells treated with brain CSPGs than in nontreated cells after exposure to the medium with CSPGs. Microspectrophotometry revealed that the population of cells with a high DNA content was greater in the culture treated with brain-derived CSPGs than in the control culture. These findings indicate that purified soluble brain CSPGs block the cell cycle of PC12D cells at the G2 phase with resultant cessation of cell proliferation and the inhibition of neurite outgrowth.