RESUMEN
Gephyrin is thought to play a critical role in clustering glycine receptors at synapses within the central nervous system (CNS). The main in vivo evidence for this comes from Gephyrin (Gphn)-null mice, where glycine receptors are depleted from synaptic regions. However, these mice die at birth, possibly due to impaired molybdenum cofactor (MoCo) synthesis, an essential role Gephyrin assumes throughout an animal. This complicates the interpretation of synaptic phenotypes in Gphn-null mice and raises the question whether the synaptic and enzymatic functions of Gephyrin can be investigated separately. Here, we generated a gephyrinb zebrafish mutant, vo84, that almost entirely lacks Gephyrin staining in the spinal cord. gephyrinbvo84 mutants exhibit normal gross morphology at both larval and adult stages. In contrast to Gphn-null mice, gephyrinbvo84 mutants exhibit normal motor activity and MoCo-dependent enzyme activity. Instead, gephyrinbvo84 mutants display impaired rheotaxis and increased mortality in late development. To investigate what may mediate these defects in gephyrinbvo84 mutants, we examined the cell density of neurons and myelin in the spinal cord and found no obvious changes. Surprisingly, in gephyrinbvo84 mutants, glycine receptors are still present in the synaptic regions. However, their abundance is reduced, potentially contributing to the observed defects. These findings challenge the notion that Gephyrin is absolutely required to cluster glycine receptors at synapses and reveals a new role of Gephyrin in regulating glycine receptor abundance and rheotaxis. They also establish a powerful new model for studying the mechanisms underlying synaptic, rather than enzymatic, functions of Gephyrin.
Asunto(s)
Proteínas Portadoras , Proteínas de la Membrana , Mutación , Médula Espinal , Sinapsis , Pez Cebra , Animales , Sinapsis/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Médula Espinal/metabolismo , Mutación/genética , Proteínas Portadoras/metabolismo , Proteínas Portadoras/genética , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Receptores de Glicina/metabolismo , Receptores de Glicina/genética , Cofactores de Molibdeno , Pteridinas , Neuronas/metabolismo , Vaina de Mielina/metabolismo , Actividad Motora/fisiología , Actividad Motora/genética , Animales Modificados GenéticamenteRESUMEN
In the nervous system, only one type of neuron-glial synapse is known to exist: that between neurons and oligodendrocyte precursor cells (OPCs), yet their composition, assembly, downstream signaling and in vivo functions remain largely unclear. Here, we address these questions using in vivo microscopy in zebrafish spinal cord and identify postsynaptic molecules PSD-95 and gephyrin in OPCs. The puncta containing these molecules in OPCs increase during early development and decrease upon OPC differentiation. These puncta are highly dynamic and frequently assemble at 'hotspots'. Gephyrin hotspots and synapse-associated Ca2+ activity in OPCs predict where a subset of myelin sheaths forms in differentiated oligodendrocytes. Further analyses reveal that spontaneous synaptic release is integral to OPC Ca2+ activity, while evoked synaptic release contributes only in early development. Finally, disruption of the synaptic genes dlg4a/dlg4b, gphnb and nlgn3b impairs OPC differentiation and myelination. Together, we propose that neuron-OPC synapses are dynamically assembled and can predetermine myelination patterns through Ca2+ signaling.
Asunto(s)
Vaina de Mielina , Células Precursoras de Oligodendrocitos , Animales , Vaina de Mielina/fisiología , Pez Cebra , Oligodendroglía/fisiología , Neuronas/fisiología , Diferenciación Celular/fisiologíaRESUMEN
Astrocytes play crucial roles in regulating neural circuit function by forming a dense network of synapse-associated membrane specializations, but signaling pathways regulating astrocyte morphogenesis remain poorly defined. Here, we show the Drosophila lipid-binding G protein-coupled receptor (GPCR) Tre1 is required for astrocytes to establish their intricate morphology in vivo. The lipid phosphate phosphatases Wunen/Wunen2 also regulate astrocyte morphology and, via Tre1, mediate astrocyte-astrocyte competition for growth-promoting lipids. Loss of s1pr1, the functional analog of Tre1 in zebrafish, disrupts astrocyte process elaboration, and live imaging and pharmacology demonstrate that S1pr1 balances proper astrocyte process extension/retraction dynamics during growth. Loss of Tre1 in flies or S1pr1 in zebrafish results in defects in simple assays of motor behavior. Tre1 and S1pr1 are thus potent evolutionarily conserved regulators of the elaboration of astrocyte morphological complexity and, ultimately, astrocyte control of behavior.