Optical Monitoring of Exo- and Endocytosis

Neurotransmitter Release

Synapses form the basic computing unit in the brain, transmitting and processing information with superb time and space precision. In congenital nerve endings, neurotransmitters are stored in synaptic vesicles (SV), released, and recovered by exocytosis and endocytosis within hundreds of microseconds. This process is mainly monitored by electrophysiology and optical imaging methods.

Patch clip electrophysiological technology can detect the total external penetration and/or endocrine signals of all staphylococci in a single cell with high resolution. Unfortunately, this technique was invasive and cannot be easily used to assess monocystic exocytosis. In contrast, optical imaging techniques allow the evaluation of single-cell or multi-celled extracellular penetration of a single vesicle, which was essential for elucidating the basic exocytosis mechanism.

Methods of Optically Monitoring of Exo- and Endocytosis

• PH-sensitive fluorescence proteins
• Electron microscopy
• Optogenetics and high-pressure freezing (flash and freeze)
• Capacitance measurement
• Quantum dots
• Fluorescent styryl dyes

Fig.1 Schematic diagrams and instruments for optically monitoring exo- and endocytosis of vesicles. (Jin, 2019)

Applications of Monitoring Exo- and Endocytosis

• Fluorescence microscopy imaging tracks the transmission pathway of individual vesicles in the hippocampus of rats by attaching fluorescent labels to specific molecules in the vesicles. Changes in the composition of synaptic vesicles lead to a variety of neurological and psychiatric diseases, and the development of drugs for the recycling of synaptic vesicles through optical monitoring may be a promising therapeutic approach.

Fig.2 Immunofluorescence analysis of hippocampal neuron V0a1-fluoroprotein expression control (empty vector). (Chanaday, 2018)

• The long-term potentiation (LTP) and long-term depression (LTD) of hippocampal glutathione resection synapses are considered to be the basic cellular mechanisms of learning and memory. Changes in the functional properties of α-amino-3-hydroxy-5-methyl-4-isooxyacrylic acid (AMPA) glutamate receptor (AMPAR) exocytosis and endocytosis are considered to be the molecular mechanism of LTP or LTD. The total internal reflection fluorescence microscope (TIRFM) was used to observe individual events of AMPAR exocytosis or endocytosis with high noise (SN) ratio during culture preparation for live cell imaging study of AMPAR dynamics.

Fig.3 Postsynaptic-like membrane (PSLM) and normal synapses observed with total internal reflection fluorescence microscopy (TIRFM) or with conventional epi-fluorescence. (Hirano, 2018)

Many studies have shown that congenital dysfunction, especially the damage of SV endocytosis, is related to neurological diseases. By monitoring SV endocytosis and exocytosis to assess innate function, it is helpful for the discovery of early markers of neurological diseases.

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References

1. Jin, Y.; et al. Various approaches for measurement of synaptic vesicle endocytosis at the central nerve terminal. Archives of pharmacal research. 2019, 42(6): 455-465.
2. Chanaday, N.L.; Kavalali, E.T. Optical detection of three modes of endocytosis at hippocampal synapses. Elife. 2018, 7: e36097.
3. Hirano, T. Visualization of exo-and endocytosis of AMPA receptors during hippocampal synaptic plasticity around postsynaptic-like membrane formed on glass surface. Frontiers in cellular neuroscience. 2018, 12: 442.