Analytical Methods to Determine the Dynamic Chemical Anatomy of Signalling Organelles

Cellular communication is mediated by nanometer-sized organelles (vesicles, stress granules, exosomes) that are responsible for carrying and releasing signalling molecules.

I propose to develop new technologies to chemically dissect vesicles, stress granules, and exosomes both dynamically and in fixed samples with spatial discrimination across living cells - the next frontier in cell analysis. The following four aims are proposed.

We will develop 1) a method with a chaotropic anion in a carbon nanopipette electrode to discriminate and quantify the contents of suborganelle chemical contents, 2) novel fluorescent nanoelectrochemical probes and employ STED microscopy to spatially track the position of the probes across cells and subcellular regions while quantifying organelle chemical content, 3) analytical methods and approaches to measure the content and location of exosomes from catecholamine cells and determine their role in neural communication, and 4) nanotip and nanopipette electrodes, STED localisation, and NanoSIMS imaging to determine if intracellular vesicles and exosomes interchange and transfer their messenger content and the mechanism by which they do so.

We discovered that most exocytosis is partial and this opens a window of discovery concerning the structural and dynamical mechanisms of organelles that regulate neural communication. Ultimately, this will allow us to unravel the chemical mechanisms of initial plasticity and brain cellular disease.