Ryanodine receptor-mediated Ca2+ release and atlastin-2 GTPase activity contribute to IP3-induced dendritic Ca2+ signals in primary hippocampal neurons

Abstract

Neuronal Ca²⁺ signals are fundamental for synaptic transmission and activity-dependent changes in gene expression. Voltage-gated Ca²⁺ channels and N-methyl-d-aspartate receptors play major roles in mediating external Ca²⁺ entry during action potential firing and glutamatergic activity. Additionally, the inositol-1,4,5-trisphosphate receptor (IP₃R) and the ryanodine receptor (RyR) channels expressed in the endoplasmic reticulum (ER) also contribute to the generation of Ca²⁺ signals in response to neuronal activity. The ER forms a network that pervades the entire neuronal volume, allowing intracellular Ca²⁺ release in dendrites, soma and presynaptic boutons. Despite its unique morphological features, the contributions of ER structure and of ER-shaping proteins such as atlastin - an ER enriched GTPase that mediates homotypic ER tubule fusion - to the generation of Ca²⁺ signals in dendrites remains unreported. Here, we investigated the contribution of RyR-mediated Ca²⁺ release to IP₃-generated Ca²⁺ signals in dendrites of cultured hippocampal neurons. We also employed GTPase activity-deficient atlastin-2 (ATL2) mutants to evaluate the potential role of atlastin on Ca²⁺ signaling and ER-resident Ca²⁺ channel distribution. We found that pharmacological suppression of RyR channel activity increased the rising time and reduced the magnitude and propagation of IP₃-induced Ca²⁺ signals. Additionally, ATL2 mutants induced specific ER morphological alterations, delayed the onset and increased the rising time of IP₃-evoked Ca²⁺ signals, and caused RyR2 and IP₃R1 aggregation and RyR2 redistribution. These results indicate that both RyR and ATL2 activity regulate IP₃-induced Ca²⁺ signal dynamics through RyR-mediated Ca²⁺-induced Ca²⁺ release, ER shaping and RyR2 distribution.

Keywords: Ca(2+) release channels; Ca(2+) transients; Ca(2+)-induced Ca(2+) release; Endoplasmic reticulum; Rat hippocampus.