Abstract
Recently, we have shown that high molecular weight polymers inhibit cortical granule exocytosis at total osmolalities only slightly higher than that of sea water (Whitaker, M., and J. Zimmerberg. 1987. J. Physiol. 389:527-539). In this study, we visualize the step at which this inhibition occurs. Lytechinus pictus and Strongylocentrotus purpuratus eggs were exposed to 0.8 M stachyose or 40% (wt/vol) dextran (average molecular mass of 10 kD) in artificial sea water, activated with 60 microM of the calcium ionophore A23187, and then either fixed with glutaraldehyde and embedded or quick-frozen and freeze-fractured. Stachyose (2.6 osmol/kg) appears to inhibit cortical granule exocytosis by eliciting formation of a granule-free zone (GFZ) in the egg cortex which pushes granules away from the plasma membrane thus preventing their fusion. In contrast, 40% dextran (1.58 osmol/kg) does not result in a GFZ and cortical granules undergo fusion. In some specimens, the pores joining granule and plasma membranes are relatively small; in other cases, the exocytotic pocket has been stabilized in an omega configuration and the granule matrix remains intact. These observations suggest that high molecular weight polymers block exocytosis because of their inability to enter the granule matrix: they retard the water entry that is needed for matrix dispersal.
Full Text
The Full Text of this article is available as a PDF (5.6 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Anderson E. Oocyte differentiation in the sea urchin, Arbacia punctulata, with particular reference to the origin of cortical granules and their participation in the cortical reaction. J Cell Biol. 1968 May;37(2):514–539. doi: 10.1083/jcb.37.2.514. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bilinski M., Plattner H., Matt H. Secretory protein decondensation as a distinct, Ca2+-mediated event during the final steps of exocytosis in Paramecium cells. J Cell Biol. 1981 Jan;88(1):179–188. doi: 10.1083/jcb.88.1.179. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chandler D. E. Comparison of quick-frozen and chemically fixed sea-urchin eggs: structural evidence that cortical granule exocytosis is preceded by a local increase in membrane mobility. J Cell Sci. 1984 Dec;72:23–36. doi: 10.1242/jcs.72.1.23. [DOI] [PubMed] [Google Scholar]
- Chandler D. E. Exocytosis involves highly localized membrane fusions. Biochem Soc Trans. 1984 Dec;12(6):961–963. doi: 10.1042/bst0120961. [DOI] [PubMed] [Google Scholar]
- Chandler D. E., Heuser J. E. Arrest of membrane fusion events in mast cells by quick-freezing. J Cell Biol. 1980 Aug;86(2):666–674. doi: 10.1083/jcb.86.2.666. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chandler D. E., Heuser J. Membrane fusion during secretion: cortical granule exocytosis in sex urchin eggs as studied by quick-freezing and freeze-fracture. J Cell Biol. 1979 Oct;83(1):91–108. doi: 10.1083/jcb.83.1.91. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chandler D. E., Heuser J. The vitelline layer of the sea urchin egg and its modification during fertilization. A freeze-fracture study using quick-freezing and deep-etching. J Cell Biol. 1980 Mar;84(3):618–632. doi: 10.1083/jcb.84.3.618. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cohen F. S., Akabas M. H., Zimmerberg J., Finkelstein A. Parameters affecting the fusion of unilamellar phospholipid vesicles with planar bilayer membranes. J Cell Biol. 1984 Mar;98(3):1054–1062. doi: 10.1083/jcb.98.3.1054. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cohen F. S., Zimmerberg J., Finkelstein A. Fusion of phospholipid vesicles with planar phospholipid bilayer membranes. II. Incorporation of a vesicular membrane marker into the planar membrane. J Gen Physiol. 1980 Mar;75(3):251–270. doi: 10.1085/jgp.75.3.251. [DOI] [PMC free article] [PubMed] [Google Scholar]
- ENDO Y. The role of the cortical granules in the formation of the fertilization membrane in the eggs of sea urchins. II. Exp Cell Res. 1961 Dec;25:518–528. doi: 10.1016/0014-4827(61)90187-2. [DOI] [PubMed] [Google Scholar]
- Eddy E. M., Shapiro B. M. Changes in the topography of the sea urchin egg after fertilization. J Cell Biol. 1976 Oct;71(1):35–48. doi: 10.1083/jcb.71.1.35. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Edwards C., Englert D., Lotshaw D., Ye H. Z. Light microscopic observations on the release of vesicles by isolated chromaffin cells. Cell Motil. 1984;4(4):297–303. doi: 10.1002/cm.970040407. [DOI] [PubMed] [Google Scholar]
- Gilligan D. M., Satir B. H. Stimulation and inhibition of secretion in Paramecium: role of divalent cations. J Cell Biol. 1983 Jul;97(1):224–234. doi: 10.1083/jcb.97.1.224. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hampton R. Y., Holz R. W. Effects of changes in osmolality on the stability and function of cultured chromaffin cells and the possible role of osmotic forces in exocytosis. J Cell Biol. 1983 Apr;96(4):1082–1088. doi: 10.1083/jcb.96.4.1082. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Heuser J. E., Reese T. S., Dennis M. J., Jan Y., Jan L., Evans L. Synaptic vesicle exocytosis captured by quick freezing and correlated with quantal transmitter release. J Cell Biol. 1979 May;81(2):275–300. doi: 10.1083/jcb.81.2.275. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kazilek C. J., Merkle C. J., Chandler D. E. Hyperosmotic inhibition of calcium signals and exocytosis in rabbit neutrophils. Am J Physiol. 1988 May;254(5 Pt 1):C709–C718. doi: 10.1152/ajpcell.1988.254.5.C709. [DOI] [PubMed] [Google Scholar]
- Lubbock R., Gupta B. L., Hall T. A. Novel role of calcium in exocytosis: mechanism of nematocyst discharge as shown by x-ray microanalysis. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3624–3628. doi: 10.1073/pnas.78.6.3624. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ornberg R. L., Reese T. S. Beginning of exocytosis captured by rapid-freezing of Limulus amebocytes. J Cell Biol. 1981 Jul;90(1):40–54. doi: 10.1083/jcb.90.1.40. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pollard H. B., Pazoles C. J., Creutz C. E., Scott J. H., Zinder O., Hotchkiss A. An osmotic mechanism for exocytosis from dissociated chromaffin cells. J Biol Chem. 1984 Jan 25;259(2):1114–1121. [PubMed] [Google Scholar]
- Pollard H. B., Tack-Goldman K., Pazoles C. J., Creutz C. E., Shulman N. R. Evidence for control of serotonin secretion from human platelets by hydroxyl ion transport and osmotic lysis. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5295–5299. doi: 10.1073/pnas.74.12.5295. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Steinhardt R., Zucker R., Schatten G. Intracellular calcium release at fertilization in the sea urchin egg. Dev Biol. 1977 Jul 1;58(1):185–196. doi: 10.1016/0012-1606(77)90084-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Swann K., Whitaker M. The part played by inositol trisphosphate and calcium in the propagation of the fertilization wave in sea urchin eggs. J Cell Biol. 1986 Dec;103(6 Pt 1):2333–2342. doi: 10.1083/jcb.103.6.2333. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Verdugo P. Hydration kinetics of exocytosed mucins in cultured secretory cells of the rabbit trachea: a new model. Ciba Found Symp. 1984;109:212–225. doi: 10.1002/9780470720905.ch15. [DOI] [PubMed] [Google Scholar]
- Veron M., Foerder C., Eddy E. M., Shapiro Sequential biochemical and morphological events during assembly of the fertilization membrane of the sea urchin. Cell. 1977 Feb;10(2):321–328. doi: 10.1016/0092-8674(77)90226-4. [DOI] [PubMed] [Google Scholar]
- Whitaker M. J., Baker P. F. Calcium-dependent exocytosis in an in vitro secretory granule plasma membrane preparation from sea urchin eggs and the effects of some inhibitors of cytoskeletal function. Proc R Soc Lond B Biol Sci. 1983 Jul 22;218(1213):397–413. doi: 10.1098/rspb.1983.0047. [DOI] [PubMed] [Google Scholar]
- Whitaker M., Zimmerberg J. Inhibition of secretory granule discharge during exocytosis in sea urchin eggs by polymer solutions. J Physiol. 1987 Aug;389:527–539. doi: 10.1113/jphysiol.1987.sp016670. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zimmerberg J., Curran M., Cohen F. S., Brodwick M. Simultaneous electrical and optical measurements show that membrane fusion precedes secretory granule swelling during exocytosis of beige mouse mast cells. Proc Natl Acad Sci U S A. 1987 Mar;84(6):1585–1589. doi: 10.1073/pnas.84.6.1585. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zimmerberg J. Molecular mechanisms of membrane fusion: steps during phospholipid and exocytotic membrane fusion. Biosci Rep. 1987 Apr;7(4):251–268. doi: 10.1007/BF01121447. [DOI] [PubMed] [Google Scholar]
- Zimmerberg J., Parsegian V. A. Polymer inaccessible volume changes during opening and closing of a voltage-dependent ionic channel. Nature. 1986 Sep 4;323(6083):36–39. doi: 10.1038/323036a0. [DOI] [PubMed] [Google Scholar]
- Zimmerberg J., Sardet C., Epel D. Exocytosis of sea urchin egg cortical vesicles in vitro is retarded by hyperosmotic sucrose: kinetics of fusion monitored by quantitative light-scattering microscopy. J Cell Biol. 1985 Dec;101(6):2398–2410. doi: 10.1083/jcb.101.6.2398. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zimmerberg J., Whitaker M. Irreversible swelling of secretory granules during exocytosis caused by calcium. Nature. 1985 Jun 13;315(6020):581–584. doi: 10.1038/315581a0. [DOI] [PubMed] [Google Scholar]