Abstract
BackgroundAstrocytoma are known to have altered glutamate machinery that results in the release of large amounts of glutamate into the extracellular space but the precise role of glutamate in favoring cancer processes has not yet been fully established. Several studies suggested that glutamate might provoke active killing of neurons thereby producing space for cancer cells to proliferate and migrate. Previously, we observed that calcium promotes disassembly of integrin-containing focal adhesions in astrocytoma, thus providing a link between calcium signaling and cell migration. The aim of this study was to determine how calcium signaling and glutamate transmission cooperate to promote enhanced astrocytoma migration.MethodsThe wound-healing model was used to assay migration of human U87MG astrocytoma cells and allowed to monitor calcium signaling during the migration process. The effect of glutamate on calcium signaling was evaluated together with the amount of glutamate released by astrocytoma during cell migration.ResultsWe observed that glutamate stimulates motility in serum-starved cells, whereas in the presence of serum, inhibitors of glutamate receptors reduce migration. Migration speed was also reduced in presence of an intracellular calcium chelator. During migration, cells displayed spontaneous Ca2+ transients. L-THA, an inhibitor of glutamate re-uptake increased the frequency of Ca2+ oscillations in oscillating cells and induced Ca2+ oscillations in quiescent cells. The frequency of migration-associated Ca2+ oscillations was reduced by prior incubation with glutamate receptor antagonists or with an anti-β1 integrin antibody. Application of glutamate induced increases in internal free Ca2+ concentration ([Ca2+]i). Finally we found that compounds known to increase [Ca2+]i in astrocytomas such as thapsigagin, ionomycin or the metabotropic glutamate receptor agonist t-ACPD, are able to induce glutamate release.ConclusionOur data demonstrate that glutamate increases migration speed in astrocytoma cells via enhancement of migration-associated Ca2+ oscillations that in turn induce glutamate secretion via an autocrine mechanism. Thus, glutamate receptors are further validated as potential targets for astrocytoma cancer therapy.
Highlights
Astrocytoma are known to have altered glutamate machinery that results in the release of large amounts of glutamate into the extracellular space but the precise role of glutamate in favoring cancer processes has not yet been fully established
We demonstrate that the human astrocytoma cell line U87MG is able to release glutamate in the extracellular space which in turn, activates glutamate receptors in an autocrine/ paracrine manner, leading to calcium signaling involved in both cell migration and enhanced glutamate release
Glutamate-enhanced migration of astrocytoma cells Initially, using the wound-healing model of cell migration, we measured the migration speed of U87MG cells plated on matrigel-coated dishes
Summary
Astrocytoma are known to have altered glutamate machinery that results in the release of large amounts of glutamate into the extracellular space but the precise role of glutamate in favoring cancer processes has not yet been fully established. We observed that calcium promotes disassembly of integrin-containing focal adhesions in astrocytoma, providing a link between calcium signaling and cell migration. Astrocytomas represent the most common type of glial tumors and are generally associated with poor prognosis as these tumor cells often diffusely infiltrate neighboring brain structures by migrating along defined pathways such as blood vessels or myelinated nerves. This characteristic makes surgical resection rarely efficient because by Migration is a cellular process by which motile cells interact with different adhesion molecules presented by other cell types and extracellular matrix. We demonstrated that in astroctytoma, increases in [Ca2+]i are associated with increased phosphorylation of FAK at Tyr-397 [8]
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