Infoscience

Journal article

Investigating cellular signaling reactions in single attoliter vesicles

Understanding cellular signaling mediated by cell surface receptors is key to modern biomedical research and drug development. The discovery of a growing no. of potential mol. targets and therapeutic compds. requires downscaling and accelerated functional screening. Receptor-mediated cellular responses are typically investigated on single cells or cell populations. Here, we show how to monitor cellular signaling reactions at a yet unreached miniaturization level. On the basis of our observations, cytochalasin induces mammalian cells to extrude from their plasma membrane submicrometer-sized native vesicles. They comprise functional cell surface receptors correctly exposing their extracellular ligand binding sites on the outer vesicle surface and retaining cytosolic proteins in the vesicle interior. As a prototypical example, ligand binding to the ionotropic 5-HT3 receptor and subsequent transmembrane Ca2+ signaling were monitored in single attoliter vesicles. Thus, native vesicles are the smallest autonomous containers capable of performing cellular signaling reactions under physiol. conditions. Because a single cell delivers about 50 native vesicles, which can be isolated and addressed as individuals, our concept allows multiple functional analyses of individual cells having a limited availability and opens new vistas for miniaturized bioanalytics. [on SciFinder (R)]

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