Methods to detect cell surface expression and constitutive activity of GPR6.

GPR6 is a constitutively active Gs-coupled receptor that can signal from intracellular compartments. We present different methods used to study cell surface expression of receptors and other membrane proteins. A comparison of these methods shows that methods based on susceptibility to proteolytic enzymes are more efficient at providing estimates of cell surface expression than the commonly used cell surface biotinylation method. We also present different methods that can be used to detect constitutive activity of Gs-coupled receptors. Imaging-based assays to detect intracellular cyclic AMP accumulation are well suited to study signaling at a single cell level. These assays are particularly useful when the cells of interest form a small fraction of the culture such as primary cultures with low transfection efficiency.