Gap junction internalization and processing in vivo: a 3D immuno-electron microscopy study

Area of Science:

• Cell Biology
• Molecular Biology
• Reproductive Biology

Background:

• Gap junctions facilitate intercellular communication via permeable channels.
• The internalization process forming connexosomes (double membrane vesicles) is not fully understood.
• Connexosomes contain cytosol and membranes from adjacent cells.

Purpose of the Study:

• To systematically investigate the fate and processing of connexosomes in intact ovarian follicles.
• To elucidate the membrane dynamics and degradation pathways of connexosomes.
• To demonstrate novel high-resolution analytical technology for gap junction processing.

Main Methods:

• High-pressure freezing and serial sectioning of ovarian follicle tissue.
• Immunogold labeling for connexin 43 (Cx43/GJA1).
• Categorization and surface area measurement of labeled structures within a defined cellular volume.

Main Results:

• Multiple connexosomes originate from larger invaginated gap junctions.
• Connexosome membranes separate, with loss of Cx43 from the outer membrane and inner membrane fission.
• Localization of cathepsin B suggests lysosomal involvement in connexosome processing.

Conclusions:

• This study provides new insights into connexosome formation and processing pathways.
• Lysosomes play a role in the degradation of processed connexosomes.
• The developed methodology enables high-resolution analysis of gap junction dynamics.