Biophysical Society Thematic Meeting| Padova 2019

Quantitative Aspects of Membrane Fusion and Fission

Poster Abstracts

34-POS Board 34 CA 2+ -INDEPENDENT AND VOLTAGE-DEPENDENT EXOCYTOSIS IN MOUSE CHROMAFFIN CELLS José Moya-Díaz 1 ; Lucas Bayonés 1 ; Mauricio Montenegro 1 ; Fernando D. Marengo 1 ; 1 Instituto de Fisiología, Biología Molecular y Neurociencias. Consejo Nacional de Investigaciones Científicas y Técnicas. Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Departamento de Fisiología y Biología Molecular y Celular, Buenos Aires, Capital Federal, Argentina It is widely demonstrated that the exocytosis of synaptic and secretory vesicles is triggered by a localized Ca 2+ increase associated to the activation of voltage dependent Ca 2+ channels. However, in neurons and neuroendocrine cells there are evidences of a different mode of fast exocytosis, induced in response to membrane depolarization but lacking Ca2+ current and intracellular Ca 2+ increase. In this work we investigated if such a mechanism can contribute to secretory vesicle exocytosis in mouse chromaffin cells. The application of brief depolatizations on cells bathed in total absence of extracellular Ca +2 induced moderate but consistent exocytosis, evaluated by patch-clamp membrane capacitance measurements and amperometry. This phenomenon was also reproduced when the release of Ca +2 from intracellular stores was pharmacologically inhibited and when high concentrations of BAPTA were introduced in the cytosol. Moreover, no increase in cytosolic Ca 2+ was observed when measured with Fluo-8. This exocytosis process is dependent on the applied membrane potential, reaching the 50% of the saturating value at approximately -30 mV, and was inhibited by the neurotoxin Bont-B that specifically cleaves the SNARE protein synaptobrevin. This Ca 2+ -independent and voltage- dependent exocytosis is very fast, saturating with a time constant < 10 ms, and recovers completely after depletion in less than 5 s. Due to these characteristics this mechanism is able to maintain synchronous fast exocytosis during action potential like stimuli applied at low frequencies. Finally, our data suggest that P/Q-type voltage dependent Ca +2 channels are suitable voltage sensors coupled to the activation of this mechanism of exocytosis.

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