EXOCYTOSIS IN THE LINGUAL MUCUS CELLS OF RANA-ESCULENTA EVOKED BY ACETYLCHOLINE - OBSERVATION OF LIVING CELLS BY CONFOCAL LASER-SCANNING MICROSCOPY

Exocytosis of a secretory cell is believed to occur when a part of the limiting membrane of the secretory granules fuses with the luminal pl asma membrane to release their contents. Using a confocal laser scanni ng microscope, we have studied these events in living frog lingual gla nd cells. In a differential interference contrast (DIC) mode, secretor y granules were found to be stable in general, but after stimulation w ith acetylcholine chloride (ACh) of more than 1 mu M concentration, th ere were abrupt changes in the light intensity of granules as describe d in other cells. We found that with the same stimulation, two differe nt types of secretion occurred. In the first type, a small vacuole was formed and granule contents were released from various regions of the cell. In the second type, a large vacuole was formed and granule cont ents were released all at once. Using two fluorescent dyes BCECF and L ucifer Yellow, we were able to perform positive and negative staining of the cell as well as its granules. Confocal images of the fluorescen ce mode clearly demonstrated the release process which corresponded to the DIC image observed simultaneously. The electron microscopic image also showed two different types of secretion.