Atomic Force Microscopy for the Cellular Localization of Secretory Granules and Coated Pits in Rat Basophilic Leukemia Cells after Antigen Stimulation

• Nakamura Ryosuke

  Faculty of Pharmaceutical Sciences, Nagoya City University

• Furuno Tadahide

  Faculty of Pharmaceutical Sciences, Nagoya City University

• Nakanishi Mamoru

  Faculty of Pharmaceutical Sciences, Nagoya City University

抄録

Atomic force microscopy was used to study the cellular localization of secretory granules and coated pits in rat basophilic leukemia (RBL-2H3) cells after antigen stimulation. RBL-2H3 cells were first primed with anti-DNP-IgE and thereafter stimulated with DNP-conjugated bovine serum albumin (DNP₇-BSA) in HEPES buffer solution with 1.0 mM Ca²⁺ (pH 7.4). After that the cells were fixed at different time intervals (2, 5, and 15 min) with 3 % formaldehyde solution, and the structures of the cell surfaces were observed by atomic force microscopy. It was found that a remarkable transformation in RBL-2H3 cell surface topography occurred 15 min after antigen stimulation. The cell shape became much more irregular, and there were various pits on the cell surfaces. These pits were classified as at least two kinds, larger and smaller. The diameter of the larger pits was about 1 μm, and that of the smaller ones was about 0.1 μm. A timecourse of the appearance of larger pits was mostly consistent with that of β-hexosaminidase secretion from RBL-2H3 cells. In the absence of external calcium ions the larger pits were never observed on the surface of RBL-2H3 cells. As the degranulation from RBL-2H3 cells did not occur without external calcium ions, the larger pits were assigned to be secretory granules in RBL-2H3 cells. However, several smaller pits were observed in the absence of calcium ions after antigen stimulation. The size of the smaller pits observed here was well consistent with that of the coated pits observed by electron microscopy. When cells pretreated with wortmannin (an inhibitor of PI3-kinase) were stimulated with antigen in the presence of external calcium ions (1.0 mM), neither the larger nor the smaller pits were observed on the surfaces of RBL-2H3 cells. This suggested the possibility that the smaller pits observed here may be due to coated pits or coated vesicles close to the cell membranes.

収録刊行物

• bioimages

  bioimages 6 (2), 69-75, 1998

  日本バイオイメージング学会

キーワード

• atomic force microscopy
• RBL-2H3 cells
• degranulation
• coated pits
• wortmannin

詳細情報

• CRID: 1390001277365792256
• NII論文ID: 10002038562
• NII書誌ID: AA11084187
• DOI: 10.11169/bioimages.6.69
• ISSN: 09192719
• 本文言語コード: en
• データソース種別:
  • JaLC
  • CiNii Articles
• 抄録ライセンスフラグ: 使用不可