Inositol (1,4,5)-trisphosphate receptor links to filamentous actin are important for generating local Ca2+ signals in pancreatic acinar cells

Abstract

We explored a potential structural and functional link between filamentous actin (F-actin) and inositol (1,4,5)-trisphosphate receptors (IP3Rs) in mouse pancreatic acinar cells. Using immunocytochemistry, F-actin and type 2 and 3 IP3Rs (IP3R2 and IP3R3) were identified in a cellular compartment immediately beneath the apical plasma membrane. In an effort to demonstrate that IP3R distribution is dependent on an intact F-actin network in the apical subplasmalemmal region, cells were treated with the actin-depolymerising agent latrunculin B. Immunocytochemistry indicated that latrunculin B treatment reduced F-actin in the basolateral subplasmalemmal compartment, and reduced and fractured F-actin in the apical subplasmalemmal compartment. This latrunculin-B-induced loss of F-actin in the apical region coincided with a reduction in IP3R2 and IP3R3, with the remaining IP3Rs localized with the remaining F-actin. Experiments using western blot analysis showed that IP3R3s are resistant to extraction by detergents, which indicates a potential interaction with the cytoskeleton. Latrunculin B treatment in whole-cell patch-clamped cells inhibited Ca2+-dependent Cl– current spikes evoked by inositol (2,4,5)-trisphosphate; this is due to an inhibition of the underlying local Ca2+ signal. Based on these findings, we suggest that IP3Rs form links with F-actin in the apical domain and that these links are essential for the generation of local Ca2+ spikes.