A fluorescence recovery after photobleaching protocol to measure surface diffusion of DAGLα in primary cultured cortical mouse neurons

Abstract

Summary: This protocol describes using fluorescence recovery after photobleaching (FRAP) of a superecliptic pHluorin (SEP)-diacylglycerol lipase α (DAGLα) to measure membrane-bound DAGLα mobility in dendritic shafts of primary cultured cortical mouse neurons. This could serve as an excellent tool to analyze endocannabinoid-mediated synaptic plasticity. We have used this protocol to show that DAGLα surface dynamics play an integral role in regulating the dendritic spine. We also detail how we test the qualities of generated SEP-DAGLα in HEK293T cells by FRAP assay.For complete details on the use and execution of this profile, please refer to Yoon et al. (2021a).

Keywords

• Cell Biology
• Cell culture
• Cell Membrane
• Cell-based Assays
• Microscopy
• Molecular Biology