| Modeling of quantitative nano-amperometric measurement of sub-quantal glutamate release by living neurons Giovanni Pireddu1; 0; Xiaoke Yang2; Fu-li Zhang2; Yan-ling Liu2; Irina Svir3; Alexander Oleinick4; Wei-hua Huang2; Christian Andre Amatore5; 1CNRS, ECOLE NORMALE SUPERIEURE, SORBONNE UNIVERSITY, Paris, France; 2WUHAN UNIVERSITY, Wuhan, China; 3ECOLE NORMALE SUPERIEURE, DEPARTMENT CHEMISTRY, PARIS, France; 4CNRS, Paris, France; 5CNRS & PSL, FRENCH ACADEMY OF SCIENCES, Paris, France; PAPER: 447/Oxidative/Regular (Oral) SCHEDULED: 17:10/Mon. 28 Nov. 2022/Ballroom B ABSTRACT: Glutamate (Glu) is a crucial fundamental excitatory neurotransmitter released through vesicular exocytosis in the central nervous system. Dysregulation of the glutamate uptake by neurons and glial cells result in increase of the glutamate extracellular concentration leading eventually to excitotoxicity associated with increased oxidative stress and neurodegeneration [1]. Hence, quantitative measurements and interpretation of intravesicular Glu and of transient exocytotic release contents directly from individual living neurons are highly desired for understanding the mechanisms (full or sub-quantal release?) of synaptic transmission and plasticity. However, this could not be achieved so far due to the lack of adequate experimental strategies relying on selective and sensitive Glu nanosensors. We will show that a novel electrochemical Glu nanobiosensor based on a single SiC nanowire [2] is prone to selectively measure in real-time Glu fluxes released via exocytosis by large Glu vesicles (ca. 125 nm diameter) present in single hippocampal axonal varicosities as well as their intravesicular content before exocytosis by IVIEC. Combination of these two series of measurements revealed a sub-quantal release mode in living hippocampal neurons, viz., only ca. one third to one half of intravesicular Glu molecules are released by individual vesicles during exocytotic events. Importantly, this fraction remained practically the same when hippocampal neurons were pretreated with L-Glu-precursor L-glutamine, while it significantly increased after zinc treatment, although in both cases the intravesicular contents before release were drastically affected. Finally, the simulations of the electrochemical monitoring of the glutamate release events will be presented. The obtained theoretical results support the quantitative measurements with the enzymatic electrode. In addition, simulation results will also serve to discuss the meaning and adequacy of pre-calibrations performed in bulk solutions [3] to assess the analytical properties of enzyme-based electrochemical nanosensors aimed to detect fast transient release events. References: [1] A.A. Kritis et al. Front. Cell. Neurosci. 9 (2015) 91. [2] X. Yang, et al. Angew. Chem. Int. Ed. 60 (2021) 15803–15808. [3] C.P. McMahon, et al. Analyst 131 (2006) 68–72. |
