Imaging GPCRs trafficking and signaling with total internal reflection fluorescence microscopy in cultured neurons.

Total internal reflection fluorescence (TIRF) microscopy allows probing the cellular events occurring close and at the plasma membrane. Over the last decade, we have seen a significant increase in the number of publications applying TIRF microscopy to unravel some of the fundamental biological questions regarding G protein-coupled receptors (GPCRs) function such as the mechanisms controlling receptor trafficking, quaternary structure, and signaling among others. Most of the published work has been performed in heterologous systems such as HEK293 and CHO cells, where the imaging surface available is higher and smoother when compared with the narrow processes or the smaller cell bodies of neurons.

Imaging of kiss-and-run exocytosis of surface receptors in neuronal cultures.

Transmembrane proteins are continuously shuttled from the endosomal compartment to the neuronal plasma membrane by highly regulated and complex trafficking steps. These events are involved in many homeostatic and physiological processes such as neuronal growth, signaling, learning and memory among others. We have previously shown that endosomal exocytosis of the B2 adrenergic receptor (B2AR) and the GluR1-containing AMPA receptor to the neuronal plasma membrane is mediated by two different types of vesicular fusion.