self-written macros  (MathWorks Inc)

Journal: Science Advances

Article Title: Dynamic regulation of vesicle pools in a detailed spatial model of the complete synaptic vesicle cycle

doi: 10.1126/sciadv.adq6477

Figure Lengend Snippet: ( A ) Snapshot of complete vesicle cycling model showing clustered, free, newly endocytosed, and docked vesicles in the reconstructed synaptic bouton. ( B ) Toy model in a cylindrical mesh used to develop the vesicle clustering model. This toy model shows vesicle clustering behavior more clearly than the complete model in the hippocampal bouton reconstruction (see movie S1 for full animated visualization)

Article Snippet: For the experiments described in the “Labeling of recycling or reserve hippocampal vesicle pools, to determine the presence of pHluorin within these pools” section, all images were analyzed using self-written macros developed in MATLAB (The Mathworks Inc., Natick, MA, USA; version R2023a).

Techniques:

Journal: Science Advances

Article Title: Dynamic regulation of vesicle pools in a detailed spatial model of the complete synaptic vesicle cycle

doi: 10.1126/sciadv.adq6477

Figure Lengend Snippet: ( A ) Fusions of vesicle types during 10-Hz model stimulation (reserve and recycling vesicles are from the initial population, and new vesicles are those formed by endocytosis). ( B ) Usage of the model reserve vesicle pool at 5, 10, 20, and 50 Hz. ( C ) Proportion of vesicle types used after 45 s of model stimulation (5 to 50 Hz). Numbers indicate the fraction of the initial population of recycling (blue) and reserve (red) vesicles used. Note: Since there is no initial pool of recycled vesicles (yellow), no number is shown. ( D ) Comparison of initial vesicle pools (including all labeled vesicles within the cytosol) usage after 30 s of stimulation at 5, 10, 20, and 50 Hz in isolated hippocampal cells with vesicles prelabeled with pHluorin (experiment) and our model (i.e., sum of initial reserve and recycling vesicles used). The experiment control condition (broken line) does not include bafilomycin. ( E ) PHluorin imaging experiments. Neurons were incubated with an antibody against the luminal domain of synaptotagmin 1, decorated with secondary nanobodies conjugated to pHluorin. This tool is incorporated into recycling vesicles during the incubation time. The top shows the response of pHluorin-labeled vesicles to stimulation, comparing baseline condition (left) to a stimulation train of 20 Hz (30 s), and lastly to a pulse of NH4Cl, to neutralize the pH inside vesicles, and thereby reveal all pHluorin molecules. The bottom shows the same experiment but in the presence of bafilomycin, which inhibits vesicle reacidification. The bottom graphs indicate the fluorescence responses (normalized to the baseline) for the different conditions. Left: A series of stimulation trains was applied, at 5, 10, 20, and 50 Hz, before a final round of NH 4 Cl. Right: The same experiment, performed in the presence of bafilomycin. Average traces from individual experiments are shown. CNQX, cyanquixaline; AP5, (2R)-amino-5-phosphonopentanoate; Baf, bafilomycin.

Article Snippet: For the experiments described in the “Labeling of recycling or reserve hippocampal vesicle pools, to determine the presence of pHluorin within these pools” section, all images were analyzed using self-written macros developed in MATLAB (The Mathworks Inc., Natick, MA, USA; version R2023a).

Techniques: Comparison, Labeling, Isolation, Control, Imaging, Incubation, Fluorescence