Home About us Contact
|
Home About us Contact | ||
|
|
||
Receptor Clustering (receptor + clustering)
Selected AbstractsOrganization of GABAA receptor ,-subunit clustering in the developing rat neocortex and hippocampusEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 9 2004B. Hutcheon Abstract We compared the expression and co-expression of ,1, ,2, ,3, and ,5-subunit protein clusters of the ,-aminobutyric acid (GABA)A receptor in the neocortex and hippocampus of rat at postnatal days (PND) 5,10 and 30,40 in order to understand how inhibitory receptors reorganize during brain maturation. The size, intensity, density and pattern of co-localization of fluorescently tagged subunit clusters were determined in deconvolved digital images using a novel 2D cross-correlational analysis. The cross-correlation analysis allowed an unbiased identification of GABAA receptor subunit clusters based on staining intensity. Cluster size increased through development; only the ,2 clusters in dentate gyrus (DG) decreased in size. ,5-subunit cluster density either increased or decreased with maturation depending on the brain region. For the other subunits, the cluster density remained rather constant, with noted exceptions (increase in ,2 clusters in cortical layer 5 but a decrease of ,3 clusters in hilus). The co-localization of ,1-subunit with the others was unique and not correlated to overall changes in subunit abundance between developmental époques. So, although ,2-subunit expression went up in the DG, the clusters became less co-localized with ,1. In contrast, ,5-subunit clusters became more co-localized with ,1 as the ,5-subunit expression declined in cortex and CA1. The co-localization of ,3 with ,1 also became greater in layer 6. In the adult brain not all clustering was associated with synapses, as many ,-subunit clusters did not co-localize with synaptophysin. Overall, these data indicate that the regulation of GABAA receptor clustering is both synaptic and extrasynaptic, presumably reflecting complex cellular trafficking mechanisms. [source] Contact-dependent aggregation of functional Ca2+ channels, synaptic vesicles and postsynaptic receptors in active zones of a neuromuscular junctionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 3 2001David A. DiGregorio Abstract To examine whether Ca2+ channels aggregate in a contact-dependent manner, we characterized the distribution of synaptic vesicles and postsynaptic receptors, and compared it to the location of Ca2+ entry sites, in a Xenopus laevis nerve-muscle coculture preparation using a localized Ca2+ detection method. The majority (75%) of Ca2+ entry sites at spontaneously formed nerve,muscle contacts were associated with enhanced immunofluorescence to the synaptic vesicle protein, SV2. In contrast, only 11% of recorded sites without Ca2+ transients exhibited significant SV2 immunofluorescence. When comparing the spatial distribution of synaptic markers with that of Ca2+ entry sites, we found that the majority of Ca2+ entry sites (61%) were associated with both enhanced SV2 immunofluorescence and R-BTX fluorescence, thereby identifying putative neurotransmitter release sites where Ca2+ channels, synaptic vesicles and postsynaptic receptors are colocalized. Using polystyrene beads coated with a heparin binding protein known to mediate in vitro postsynaptic receptor clustering, we show that the location of Ca2+ domains was associated with enhanced SV2 immunofluorescence at neurite-to-bead contacts. We conclude that the localization of functional Ca2+ channels to putative active zones follows a contact-dependent signalling mechanism similar to that known to mediate vesicle aggregation and AChR clustering. [source] Glycoprotein Ib,IX-mediated activation of integrin ,IIb,3: effects of receptor clustering and von Willebrand factor adhesionJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 6 2003M. Arya Summary., The interaction between the platelet glycoprotein (GP) Ib,IX complex and von Willebrand factor (VWF) initiates both hemostasis and pathological thrombosis. This interaction is not only the first adhesive event of platelets at sites of vessel injury, but also facilitates fibrinogen binding to ,IIb,3, which subsequently results in platelet aggregation. Since it has been suggested that GP Ib,IX clustering may promote platelet activation, we investigated the effect of such clustering on both VWF,GP Ib,IX and fibrinogen,,IIb,3 bonds using optical tweezers. In our system, fusion of tandem repeats of FK506-binding protein (FKBP) to the cytoplasmic tail of the GP IX subunit of the GP Ib,IX complex allowed subsequent receptor clustering within the plasma membrane by the bivalent, cell-permeant small molecule ligand AP20187. We measured binding forces between polystyrene beads coated with either plasma-derived VWF or the VWF A1 domain and GP Ib,IX(FKBP)2, and those between fibrinogen-coated beads and ,IIb,3 expressed on Chinese hamster ovary cells. The minimal detachment force between GP Ib,IX(FKBP)2 and A1 or plasma-derived VWF doubled after AP20187 was added. The binding force between immobilized fibrinogen and ,IIb,3 was not changed by the clustering agent; however, the strength of single fibrinogen,,IIb,3 bonds increased significantly after ligation of GP Ib,IX(FKBP)2 by A1. These results demonstrate that GP Ib,IX clustering increases the overall strength of its interaction with VWF. Furthermore, signals from GP Ib,IX can activate ,IIb,3, thereby increasing the strength of its interaction with fibrinogen. [source] Self-association of EPEC intimin mediated by the ,-barrel-containing anchor domain: a role in clustering of the Tir receptorMOLECULAR MICROBIOLOGY, Issue 1 2004Thierry Touzé Summary Outer membrane intimin directs attachment of enteropathogenic Escherichia coli (EPEC) via its Tir receptor in mammalian target cell membranes. Phosphorylation of Tir triggers local actin polymerization and the formation of ,pedestal-like' pseudopods. We demonstrate that the intimin protein contains three domains, a flexible N-terminus (residues 40,188), a central membrane-integrated ,-barrel (189,549), and a tightly folded Tir-binding domain (550,939). Intimin was shown by electron microscopy to form ring-like structures with a ,7 nm external diameter and an electron dense core, and to form channels of 50picoSiemens conductance in planar lipid bilayers. Gel filtration, multiangle light scattering and cross-linking showed that this central ,-barrel membrane-anchoring domain directs intimin dimerization. Isothermal titration calorimetry revealed a high affinity, single-binding site interaction of 2 : 1 stoichiometry between dimeric intimin and Tir, and modelling suggests that this interaction determines a reticular array-like superstructure underlying receptor clustering. In support of this model, actin rearrangement induced in Tir-primed cultured cells by intimin-containing proteoliposomes was dependent on the concentration of both intimin and Tir, and co-localized with clustered phosphorylated Tir. [source] Early expression of AMPA receptors and lack of NMDA receptors in developing rat climbing fibre synapsesTHE JOURNAL OF PHYSIOLOGY, Issue 3 2005Philippe Lachamp Whether nascent glutamatergic synapses acquire their AMPA receptors constitutively or via a regulated pathway triggered by pre-existing NMDA receptor activation is still an open issue. Here, we provide evidence that some glutamatergic synapses develop without expressing NMDA receptors. Using immunocytochemistry, we showed that synapses between developing rat climbing fibres and Purkinje cells expressed GluR2-containing AMPA receptors as soon as they were formed (i.e. on embryonic day 19) but never carried detectable NMDA receptors. This was confirmed by electrophysiological recordings. Excitatory synaptic currents were recorded in Purkinje cells as early as P0. However, no NMDA receptor-mediated component was found in either spontaneous or evoked synaptic responses. In addition, we ruled out a possible role of extrasynaptic NMDA receptors by showing that AMPA receptor clustering at nascent climbing fibre synapses was not modified by chronic in utero NMDA receptor blockade. [source] | ||||||||||