RESUMEN
In the developing cortex, axons and dendrites extend progressively in response to environmental cues attracting or repelling growing processes. Recent evidence suggests the existence of a functional link between guidance molecules and metalloproteinases. Here, we analyzed the putative functional interaction of matrix metalloproteinases (MMPs) with guidance cues of the semaphorin family during growth and guidance of cortical axons. Our results demonstrate that the expression pattern and the proteolytic activity of MMP-3 are consistent with a role of this particular MMP during cortical axon outgrowth. We found that MMP-3 is required for an optimal axon extension and is involved in the Sema3C-dependent chemoattraction of cortical axons by modulating both the growth capacity and the orientation of growth. Interestingly, the inhibitory Sema3A decreased both the expression and activity of MMP-3. Taken together, our results reveal a molecular interaction between MMPs and semaphorins providing new insight into the molecular mechanism allowing axonal growth cone to respond to environmental guidance cues in the context of cortical development.
Asunto(s)
Axones/fisiología , Corteza Cerebral/embriología , Corteza Cerebral/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Metaloproteinasa 3 de la Matriz/metabolismo , Semaforinas/metabolismo , Animales , Axones/ultraestructura , Células Cultivadas , Corteza Cerebral/citología , Ratones , Mapeo de Interacción de ProteínasRESUMEN
Semaphorins are multifunctional factors implicated in various developmental processes. Little is known about the intracellular pathways ensuring appropriate signal transduction that encode the diverse functions observed. In this study, we investigated whether mitogen-activated protein kinases (MAPK), which are key elements of signal transduction in eukaryotic cells, were activated during semaphorin 3A (Sema3A)-induced repulsion or apoptosis of neural progenitor cells. We found that selective recruitment of the ERK1/2 pathway occurred during Sema3A-induced neural progenitor cell repulsion, whereas p38 MAPK activation was necessary for induction of apoptosis. Moreover, we provide evidence for the involvement of vascular endothelial growth factor receptor 1 (VEGFR1) in the activation of ERK1/2. Additional experiments performed with native cerebellar progenitors confirmed such a selective recruitment of MAPK during Sema3A-dependent migration or apoptosis. Altogether, our results suggest a model to explain how a single factor can exert different functions for a given cell type by the selective recruitment of intracellular pathways.
Asunto(s)
Apoptosis/fisiología , Sistema de Señalización de MAP Quinasas/fisiología , Neuronas/enzimología , Semaforina-3A/metabolismo , Células Madre/enzimología , Animales , Apoptosis/efectos de los fármacos , Comunicación Celular/efectos de los fármacos , Comunicación Celular/fisiología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Activación Enzimática/efectos de los fármacos , Activación Enzimática/fisiología , Inhibidores Enzimáticos/farmacología , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Proteína Quinasa 3 Activada por Mitógenos , Proteínas Quinasas Activadas por Mitógenos/efectos de los fármacos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Sistema Nervioso/citología , Sistema Nervioso/embriología , Sistema Nervioso/enzimología , Neuronas/citología , Semaforina-3A/farmacología , Células Madre/citología , Células Madre/efectos de los fármacos , Receptor 1 de Factores de Crecimiento Endotelial Vascular/efectos de los fármacos , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Astrocytes release glutamate and aspartate in response to elevated intracellular calcium levels, and it has been proposed that this occurs by a vesicular release mechanism, in which SNARE proteins are implicated. Although syntaxin, synaptobrevin, and cellubrevin have been shown to be expressed by cultured astrocytes, SNAP-25 has not been detected. By using immunocytochemical, immunoblotting, and polymerase chain reaction techniques, the present study demonstrates that SNAP-23, an analogue of SNAP-25, is expressed by astrocytes both in culture and in rat cerebellum. These findings provide additional evidence that astrocytes release excitatory amino acids by a vesicular mechanism involving SNARE proteins. SNAP-23 and also syntaxin 1 and cellubrevin were found to be expressed in glial precursor cells, oligodendrocytes, and microglia. These data suggest that the t-SNAREs SNAP-23 and syntaxin 1 and the v-SNARE cellubrevin participate in general membrane insertion mechanisms involved in diverse glial cell functions such as secretion, phagocytosis, and myelinogenesis.
Asunto(s)
Antígenos de Superficie/biosíntesis , Proteínas Portadoras/biosíntesis , Proteínas de la Membrana/biosíntesis , Proteínas del Tejido Nervioso/biosíntesis , Neuroglía/metabolismo , Animales , Animales Recién Nacidos , Antígenos de Superficie/genética , Astrocitos/metabolismo , Western Blotting , Proteínas Portadoras/genética , Membrana Celular/metabolismo , Membrana Celular/fisiología , Células Cultivadas , Técnica del Anticuerpo Fluorescente Indirecta , Proteínas de la Membrana/genética , Microglía/metabolismo , Proteínas del Tejido Nervioso/genética , Oligodendroglía/metabolismo , Especificidad de Órganos , Reacción en Cadena de la Polimerasa , Proteínas Qb-SNARE , Proteínas Qc-SNARE , Ratas , Ratas Wistar , Células Madre/metabolismo , Proteína 25 Asociada a Sinaptosomas , Sintaxina 1 , Proteína 3 de Membrana Asociada a VesículasRESUMEN
Enzymatic antioxidant defense systems, like superoxide dismutase (SOD), may protect neuronal and glial cells from reactive oxygen species (ROS) damage. Beside the cytosolic constitutive CuZn SOD, mitochondrial manganese SOD (Mn SOD) represents a ROS inducible enzyme which should allow the adaptation of brain cells to variation in ROS concentrations resulting from their oxidative metabolism. Using immunocytochemistry, the distribution of Mn SOD among the various representatives of the rat brain glial population (astroglia and microglia in primary culture as well as oligodendroglia in secondary culture) has been examined. Among astroglial cells, only a population of flat polygonal-shaped astrocytes, highly immunostained for glial fibrillary acid protein (GFAP) express Mn SOD immunoreactivity. Microglial cells defined by their shape and OX-42 immunoreactivity also express an intense Mn SOD signal. Exposure of the primary culture to reactive oxygen species generated by a xanthine/xanthine oxidase mixture (X/XO) accentuates the Mn SOD signal in astroglial and microglial cells. On the contrary, oligodendroglial cells grown in secondary culture in a serum-free chemically defined or a serum-containing medium and well characterized by their 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) immunoreactivity never express any immunostaining for Mn SOD, even in response to an extracellular reactive oxygen species generating source like X/XO. Likewise, a population of A2B5-positive glial cells which may represent bipotential O-2A progenitor precursors does not express Mn SOD immunostaining. These results point out that in addition to the well known ability of microglial and astroglial cells to secrete ROS, they also express a high mitochondrial oxygen superoxide decomposition potential. On the contrary, the absence of any observable Mn SOD signal in precursors and in more differentiated oligodendroglial cells could be related to their great sensitivity to ROS damage and could therefore play an important role in the development of various dysmyelinating disorders.