cAMP-induced Cytoskeleton Rearrangement Increases Calcium Transients through the Enhancement of Capacitative Calcium Entry

Abstract

In this report we investigated the correlation between cell morphology and regulation of cytosolic calcium homeostasis. Type I astrocytes were differentiated to stellate process-bearing cells by a 100-min exposure to cAMP. Differentiation of cortical astrocytes increased the magnitude and duration of calcium transients elicited by phospholipase C-activating agents as measured by single cell Fura-2-based imaging. Calcium imaging showed differences in the spatial pattern of the response. In both differentiated and the control cells, the response originated in the periphery and gradually extended into the center of the cell. However, the elevation of cytosolic calcium concentration ([Ca 2؉ ] i ) was particularly evident within the processes and adjacent to the inner cell membrane of the differentiated astrocytes. In addition, differentiation significantly prolonged the duration of the [Ca 2؉ ] i elevation. Potentiation of the calcium transients was mimicked by forskolin-induced differentiation and abolished by a specific protein kinase-A blocker. Conversely, the enhancement of the calcium transients was not mimicked by brief exposure to cAMP not causing morphological differentiation, and in PC12 cells that did not undergo morphological changes after 100 min of cAMP treatment. Impairing cAMP-induced cytoskeleton re-organization, by means of cytochalasin D and nocodazole, prevented the potentiation of the calcium transients in cAMPtreated astrocytes. Phospholipase C activity and sensitivity to inositol (1,4,5)-trisphosphate were not involved in the enhancement of the calcium responses. Also, potentiation of the calcium transients was dependent on extracellular calcium. Calcium storage and thapsigargin-depletable intracellular calcium reservoirs were analogously not increased in differentiated astrocytes. Rearrangement of the cell shape also caused a condensation of the endoplasmic reticulum and altered the spatial relationship between the endoplasmic reticulum and the cell membrane. In conclusion, morphological rearrangements of type I astrocytes increase the magnitude and the duration of agonist-induced calcium transients via enhancement of capacitative calcium entry and is associated with a spatial reorganization of the relationship between cell membrane and the endoplasmic reticulum structures.

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

“…However, the method used in our study delivers (2,4,5)IP 3 to elicit local Ca 2+ spikes directly, and therefore the effects of latrunculin B cannot be mediated through an action on cell-surface receptors (Shin et al, 2001). Disruption of the actin cytokeleton has been shown to inhibit Ca 2+ entry in vascular endothelial cells (Holda and Blatter, 1997), astrocytes (Grimaldi et al, 1999) and platelets (Rosado and Sage, 2000). However, it is unlikely that the effects of latrunculin B on the local Ca 2+ response that were measured in our study are due to a block of Ca 2+ entry as the local Ca 2+ response is largely independent of extracellular Ca 2+ (Wakui et al, 1989).…”

Cyclic Nucleotides Modulate Store-mediated Calcium Entry through the Activation of Protein-tyrosine Phosphatases and Altered Actin Polymerization in Human Platelets

“…In the secretion-like coupling model for SMCE, the actin cytoskeleton plays an important role, which has been demonstrated in several cell types (11,12), including platelets (5,9). Platelet activation is accompanied by a dramatic increase in tyrosine phosphorylation of many cellular proteins (13).…”

Microtubules Regulate Local Ca2+ Spiking in Secretory Epithelial Cells

“…For example, in acinar cells it is clear that agonist action at receptors on the basal pole leads to global elevation in IP 3 and a primary effect on Ca 2ϩ release sites at the opposite, secretory (apical) pole (7). In fact, in contrast to Ribeiro et al, evidence from endothelial cells (30,31,34), type 1 astrocytes (32), and the work we now present in acinar cells (studying the global Ca 2ϩ response) shows that agonists are capable of inducing responses after cytoskeletal disruption.…”

“…However, it is not clear at the moment if microtubules or microfilaments play a specific role in this process or if their action is an indirect effect of changes in cell shape. In type 1 astrocytes, microfilament or microtubule disruption abolished a cAMPmediated up-regulation of Ca 2ϩ entry (32), and cytoskeletal disruption was associated with changes in the endoplasmic reticulum, as visualized with ER-Tracker. Abolition of storeoperated Ca 2ϩ entry has also been seen in HEK293 cells by a calyculin A-mediated production of a cortical actin network (33).…”