Super-Resolution Analysis of the Origins of the Elementary Events of ER Calcium Release in Dorsal Root Ganglion Neurons.

Coordinated events of calcium (Ca) released from the endoplasmic reticulum (ER) are key second messengers in excitable cells. In pain-sensing dorsal root ganglion (DRG) neurons, these events can be observed as Ca sparks, produced by a combination of ryanodine receptors (RyR) and inositol 1,4,5-triphosphate receptors (IP3R1). These microscopic signals offer the neuronal cells with a possible means of modulating the subplasmalemmal Ca handling, initiating vesicular exocytosis.

A correlative super-resolution protocol to visualise structural underpinnings of fast second-messenger signalling in primary cell types.

Nanometre-scale cellular information obtained through super-resolution microscopies are often unaccompanied by functional information, particularly transient and diffusible signals through which life is orchestrated in the nano-micrometre spatial scale. We describe a correlative imaging protocol which allows the ubiquitous intracellular second messenger, calcium (Ca), to be directly visualised against nanoscale patterns of the ryanodine receptor (RyR) Ca channels which give rise to these Ca signals in wildtype primary cells. This was achieved by combining total internal reflection fluorescence (TIRF) imaging of the elementary Ca signals, with the subsequent DNA-PAINT imaging of the RyRs.