Synaptopodin Regulates Spine Plasticity: Mediation by Calcium Stores

Abstract

The role of synaptopodin (SP), an actin-binding protein residing in dendritic spines, in synaptic plasticity was studied in dissociated cultures of hippocampus taken from control and SP knock-out (SPKO) mice. Unlike controls, SPKO cultures were unable to express changes in network activity or morphological plasticity after intense activation of their NMDA receptors. SPKO neurons were transfected with SP-GFP, such that the only SP resident in these neurons is the fluorescent species. The localization and intensity of the transfected SP were similar to that of the native one. Because less than half of the spines in the transfected neurons contained SP, comparisons were made between SP-containing (SP(ϩ)) and SP lacking (SP(Ϫ)) spines in the same dendritic segments. Synaptic plasticity was induced either in the entire network by facilitation of the activation of the NMDA receptor, or specifically by local flash photolysis of caged glutamate. After activation, spines that were endowed with SP puncta were much more likely to expand than SP(Ϫ) spines. The spine expansion was suppressed by thapsigargin, which disables calcium stores. The mechanism through which SP may promote plasticity is indicated by the observations that STIM-1, the sensor of calcium concentration in stores, and Orai-1, the calcium-induced calcium entry channel, are colocalized with SP, in the same dendritic spines. The structural basis of SP is likely to be the spine apparatus, found in control but not in SPKO cells. These results indicate that SP has an essential, calcium store-related role in regulating synaptic plasticity in cultured hippocampal neurons.

Synaptic plasticity at the interface of health and disease: New insights on the role of endoplasmic reticulum intracellular calcium stores

“…Synaptopodin (SP; Mundel et al, 1997;Asanuma et al, 2005) was recognized to be a marker (Deller et al, 2000) and essential component of the SA (Deller et al, 2003). SP-deficient mice lack SAs (and COs; Bas and show deficits in synaptic plasticity (Deller et al, 2003;Jedlicka et al, 2009;Zhang et al, 2013;Vlachos et al, 2013a;Korkotian et al, 2014). Meanwhile, experimental evidence has been provided that SP/SA Ca 2+ stores regulate different forms of synaptic plasticity: (1) RyR-dependent long-term potentiation (LTP) of synaptic strength was reported (Vlachos et al, 2009), as well as (2) mGluR-triggered IP3R-mediated synaptic depression, i.e., long-term depression (LTD) of excitatory synaptic strength (Holbro et al, 2009).…”

Chronic exposure to alcohol alters network activity and morphology of cultured hippocampal neurons

“…Imaging was conducted as detailed elsewhere (Korkotian et al, 2014;Cohen et al, 2008). Glass coverslips containing the cultures were incubated for 1 h at room temperature with the standard recording medium (in mM, 129 NaCl; 4 KCl; 2 CaCl 2 ; 1 MgCl 2 ; 10 Glucose; 10 HEPES; pH 7.4, 320 mOsm), containing 2 mM Fluo-2 High Aff (AM) (TEFLabs, USA).…”

“…Cells were transfected with green fluorescent protein (GFP) at the age of 7 days in vitro, using lipofectamine 2000 (Korkotian et al, 2014). Control and 1% ethanol-treated cells were fixed with 4% paraformaldehyde at the age of 14-21 days in vitro, and individual neurons were imaged on the stage of a confocal microscope, 3D reconstructed, and analyzed morphologically in a blind procedure.…”

“…Dendritic spines were classified into mushroom type, when the spine head was large (>1 mm) compared to the spine neck, and to filopodia which lacked a distinct head. This was based on previous classifications (see Korkotian and Segal, 2011;Korkotian et al, 2014).…”

Orai1 regulates calcium entry into dendritic spines

“…This study follows our interest in the role of calcium stores in spine plasticity, where we found that dendritic spines contain ryanodine receptor-type calcium stores. 5 In the more recent study, 6 we employed plasmids that encode the Orai1 protein, as well as plasmids that encode the dominant negative (DN) Orai1. We also knocked down Orai1, using selective siRNA for this protein.…”