Freeze-fracture study of the zymogen granule membrane of pancreas: two novel types of intramembrane particles.
The ultrastructure of the zymogen granule (ZG) membrane has been observed in vitro by rapid freezing and freeze-fracture techniques. Unidirectional shadowing of the plasmic fracture (PF) leaflet of the intact granule reveals a relatively smooth surface uniformly studded by intramembrane particles (IMP; 360 microns2) their diameters ranging from 5 to 18 nm (mean = 10.2 nm) but does not allow a clear visualization of the particles on the external fracture (EF) leaflet. Indeed, rotary shadowing reveals that the EF leaflet presents a highly textured subparticle background with a significantly lower frequency of IMP (44 microns2) showing diameters from 9 to 18 nm and a shift to larger IMP (mean = 12.3 nm). Two hitherto undescribed types of IMP are found on both leaflets of the membrane: first a population of 13-nm particles with an electron-lucent center or "pore", the most frequent type on the EF face (26%), is a second population of large IMP (15 nm) characterized by a large "pore" (5.0 nm diameter) subdivided by a delicate cross-shaped structure. In alkaline conditions, pH 8.2, ZG lysis occurs rapidly and membrane ghosts thus obtained were rapidly frozen or suspended in dextran and filtered immediately. Transmission electron microscopy (TEM) shows many opened ghosts with adhering amorphous material and numerous small vesicles near or still attached to openings in the ghosts. Freeze-fracture preparations show that granule lysis is accompanied by major alterations of membrane ultrastructure; the subparticle background on the EF leaflet is now visible only as a cap or linear crest at one pole of the ghosts. These two newly formed zones are demarcated by a row of 13-nm particles, whereas the other IMP are confined to the subparticle background. Some images suggest that the subparticle background and 13-nm IMP necklace give rise to vesicles, some of them occasionally attached to the ghosts. The subparticle background on the EF leaflet shows a complementary imprint on the PF leaflet which is similarly modified. This study shows the presence of hitherto undescribed types of IMP and also demonstrates alterations of certain domains of zymogen granule membranes that occur at the moment of lysis, associated with a redistribution of different particle populations.
Cabana
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C
Centre de recherche sur les mécanismes de sécrétion, Université de Sherbrooke, Québec, Canada.
Magny
P
P
Nadeau
D
D
Grondin
G
G
Beaudoin
A
A
D000818:Animals; D003594:Cytoplasmic Granules / Q000648:ultrastructure*; D004792:Enzyme Precursors; D005614:Freeze Fracturing; D007425:Intracellular Membranes / Q000648:ultrastructure*; D008297:Male; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning; D010179:Pancreas / Q000032:analysis / Q000648:ultrastructure*; D051381:Rats; D011919:Rats, Inbred Strains
D004792:Enzyme Precursors