JCS034603 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Three-dimensional reconstructed image of an ANF-EGFP-transfected hippocampal neuron. ANF-EGFP localized to punctate vesicular structures throughout the soma and neurites of the neurons.

• Supplemental Figure S2 -

  Fig. S2. Intracellular Ca²⁺ level elevated by 90 mM and 30 mM K⁺. Ca²⁺ levels were significantly elevated in soma and neurites under both stimulation conditions. 90 mM K⁺ led to higher Ca²⁺ levels than 30 mM K⁺.

• Movie 1 -

  Movie 1. Exocytosis of ANF-EGFP-containing DCVs in the soma and neurites of hippocampal neurons. The time point of 90 mM K⁺ stimulation was set as 0. The movie lost focus briefly during buffer exchange. The ‘footprint’ of the cell body was circled by a gray line. All exocytic events occurred within the circle. To view the soma and neurites together, the incident angle was adjusted slightly smaller than the critical TIRF angle. See Movies 2 and 3 for better resolution of single exocytic events.

• Movie 2 -

  Movie 2. Exocytosis of ANF-EGFP-containing DCVs in the soma evoked by 90 mM K⁺. Typical events are indicated by arrows and arrowheads. Arrows: examples in which about 30% of the contents were released (the majority of events); arrowheads: examples in which about 65% of the contents were released (the minority of events); arrows with asterisk: examples of single vesicles undergoing multiple exocytic events.

• Movie 3 -

  Movie 3. Exocytosis of ANF-EGFP-containing DCVs in neurites evoked by 90 mM K⁺. Events are indicated by arrows. Note the low frequency of events and the long latency between stimulation and exocytosis. Also note that events are slower but fuller compared with events in the soma shown in Movie 2.

• Movie 4 -

  Movie 4. Exocytosis of BDNF-EGFP-containing DCVs in the soma evoked by 90 mM K⁺. Representative events are indicated by arrows. Unlike ANF-EGFP release, fluorescence clouds were not detected for BDNF-EGFP-containing DCV exocytic events.