期刊
CELL CALCIUM
卷 53, 期 5-6, 页码 338-348出版社
CHURCHILL LIVINGSTONE
DOI: 10.1016/j.ceca.2013.03.001
关键词
Profilin; Actin; Astrocyte; Morphological plasticity; Peripheral processes; Filopodia; Membrane tracing; PAP; Calcium; Photolysis; Uncaging; Cytoskeleton
类别
Peripheral astrocytic processes (PAPs) are highly motile structures that are strategically positioned in close proximity to synapses. Long-lasting PAP retraction in hypothalamus is known to alter synaptic transmission [1]. The PAP motility is likely to be actin-based because they are known to contain actin-related proteins such as Ezrin [2]. However, the link between dynamic activity-dependent changes in astrocytic morphology and the synaptic function has not been established experimentally, presumably due to lack of appropriate tools. To selectively suppress activity-dependent morphological plasticity of astrocytes, we developed a bicistronic construct that allows simultaneous tracing and manipulating the morphology of PAPs. The construct is designed for co-expression of (i) the mutant actin binding protein Profilin-1 (abdProf-1) with a single amino acid substitution (H119E) that prevents its binding to actin monomers [3] with (ii) the membrane-targeted morphological tracer LckGFP [4]. Cultured cortical astrocytes transfected with this construct showed abdProf-1 overexpression at a 5-fold level compared to the endogenous Profilin-1. The cells also expressed LckGFP at a level sufficient for precise morphological tracing. We found that photolysis of caged Ca2+ induced a pronounced outgrowth of PAPs, which was suppressed by abdProf-1 overexpression in terms of PAP number, growth rate and maximal length. In contrast, the morphological complexity of astrocytes, basal motility of their PAPs and major cytoskeletal structures were not affected by abdProf-1 overexpression. In summary, we identified the actin binding by Profilin-1 as a pivotal mechanism in activity-dependent morphological plasticity of PAPs in cultured astrocytes. (C) 2013 Elsevier Ltd. All rights reserved.
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