Skip to main content
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1989 Dec 1;109(6):3425–3433. doi: 10.1083/jcb.109.6.3425

Microvesicles of the neurohypophysis are biochemically related to small synaptic vesicles of presynaptic nerve terminals

PMCID: PMC2115912  PMID: 2513331

Abstract

Nerve endings of the posterior pituitary are densely populated by dense- core neurosecretory granules which are the storage sites for peptide neurohormones. In addition, they contain numerous clear microvesicles which are the same size as small synaptic vesicles of typical presynaptic nerve terminals. Several of the major proteins of small synaptic vesicles of presynaptic nerve terminals are present at high concentration in the posterior pituitary. We have now investigated the subcellular localization of such proteins. By immunogold electron microscopy carried out on bovine neurohypophysis we have found that three of these proteins, synapsin I, Protein III, and synaptophysin (protein p38) were concentrated on microvesicles but were not detectable in the membranes of neurosecretory granules. In addition, we have studied the distribution of the same proteins and of the synaptic vesicle protein p65 in subcellular fractions of bovine posterior pituitaries obtained by sucrose density centrifugation. We have found that the intrinsic membrane proteins synaptophysin and p65 had an identical distribution and were restricted to low density fractions of the gradient which contained numerous clear microvesicles with a size range the same as that of small synaptic vesicles. The peripheral membrane proteins synapsin I and Protein III exhibited a broader distribution extending into the denser part of the gradient. However, the amount of these proteins clearly declined in the fractions preceding the peak of neurosecretory granules. Our results suggest that microvesicles of the neurohypophysis are biochemically related to small synaptic vesicles of all other nerve terminals and argue against the hypothesis that such vesicles represent an endocytic byproduct of exocytosis of neurosecretory granules.

Full Text

The Full Text of this article is available as a PDF (3.8 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alonso G., Assenmacher I. The smooth endoplasmic reticulum in neurohypophysial axons of the rat: possible involvement in transport, storage and release of neurosecretory material. Cell Tissue Res. 1979 Jul 17;199(3):415–429. doi: 10.1007/BF00236080. [DOI] [PubMed] [Google Scholar]
  2. Baker P. F., Ravazzola M., Malaisse-Lagae F. Secretion-dependent uptake of extracellular fluid by the rat neurohypophysis. Nature. 1974 Jul 12;250(462):155–157. doi: 10.1038/250155a0. [DOI] [PubMed] [Google Scholar]
  3. Baumert M., Maycox P. R., Navone F., De Camilli P., Jahn R. Synaptobrevin: an integral membrane protein of 18,000 daltons present in small synaptic vesicles of rat brain. EMBO J. 1989 Feb;8(2):379–384. doi: 10.1002/j.1460-2075.1989.tb03388.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Borgese N., Macconi D., Parola L., Pietrini G. Rat erythrocyte NADH-cytochrome b5 reductase. Quantitation and comparison between the membrane-bound and soluble forms using an antibody against the rat liver enzyme. J Biol Chem. 1982 Nov 25;257(22):13854–13861. [PubMed] [Google Scholar]
  5. Browning M. D., Huang C. K., Greengard P. Similarities between protein IIIa and protein IIIb, two prominent synaptic vesicle-associated phosphoproteins. J Neurosci. 1987 Mar;7(3):847–853. doi: 10.1523/JNEUROSCI.07-03-00847.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brownstein M. J., Russell J. T., Gainer H. Synthesis, transport, and release of posterior pituitary hormones. Science. 1980 Jan 25;207(4429):373–378. doi: 10.1126/science.6153132. [DOI] [PubMed] [Google Scholar]
  7. Buckley K., Kelly R. B. Identification of a transmembrane glycoprotein specific for secretory vesicles of neural and endocrine cells. J Cell Biol. 1985 Apr;100(4):1284–1294. doi: 10.1083/jcb.100.4.1284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ceccarelli B., Hurlbut W. P., Mauro A. Turnover of transmitter and synaptic vesicles at the frog neuromuscular junction. J Cell Biol. 1973 May;57(2):499–524. doi: 10.1083/jcb.57.2.499. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Coulter H. D. Vesicular localization of immunoreactive [Met5]enkephalin in the globus pallidus. Proc Natl Acad Sci U S A. 1988 Sep;85(18):7028–7032. doi: 10.1073/pnas.85.18.7028. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. De Camilli P., Cameron R., Greengard P. Synapsin I (protein I), a nerve terminal-specific phosphoprotein. I. Its general distribution in synapses of the central and peripheral nervous system demonstrated by immunofluorescence in frozen and plastic sections. J Cell Biol. 1983 May;96(5):1337–1354. doi: 10.1083/jcb.96.5.1337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. De Camilli P., Harris S. M., Jr, Huttner W. B., Greengard P. Synapsin I (Protein I), a nerve terminal-specific phosphoprotein. II. Its specific association with synaptic vesicles demonstrated by immunocytochemistry in agarose-embedded synaptosomes. J Cell Biol. 1983 May;96(5):1355–1373. doi: 10.1083/jcb.96.5.1355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. De Camilli P., Ueda T., Bloom F. E., Battenberg E., Greengard P. Widespread distribution of protein I in the central and peripheral nervous systems. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5977–5981. doi: 10.1073/pnas.76.11.5977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. De Camilli P., Vitadello M., Canevini M. P., Zanoni R., Jahn R., Gorio A. The synaptic vesicle proteins synapsin I and synaptophysin (protein P38) are concentrated both in efferent and afferent nerve endings of the skeletal muscle. J Neurosci. 1988 May;8(5):1625–1631. doi: 10.1523/JNEUROSCI.08-05-01625.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dean C. R., Hope D. B. The isolation of purified neurosecretory granules from bovine pituitary posterior lobes. Comparison of granule protein constituents with those of neurophysin. Biochem J. 1967 Sep;104(3):1082–1088. doi: 10.1042/bj1041082. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fournier S., Trifaró J. M. A similar calmodulin-binding protein expressed in chromaffin, synaptic, and neurohypophyseal secretory vesicles. J Neurochem. 1988 Jan;50(1):27–37. doi: 10.1111/j.1471-4159.1988.tb13225.x. [DOI] [PubMed] [Google Scholar]
  16. GERSCHENFELD H. M., TRAMEZZANI J. H., DE ROBERTIS E. Ultrastructure and function in neurohypophysis of the toad. Endocrinology. 1960 May;66:741–762. doi: 10.1210/endo-66-5-741. [DOI] [PubMed] [Google Scholar]
  17. Heuser J. E., Reese T. S. Evidence for recycling of synaptic vesicle membrane during transmitter release at the frog neuromuscular junction. J Cell Biol. 1973 May;57(2):315–344. doi: 10.1083/jcb.57.2.315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Holtzman E., Freeman A. R., Kashner L. A. Stimulation-dependent alterations in peroxidase uptake at lobster neuromuscular junctions. Science. 1971 Aug 20;173(3998):733–736. doi: 10.1126/science.173.3998.733. [DOI] [PubMed] [Google Scholar]
  19. Huttner W. B., Schiebler W., Greengard P., De Camilli P. Synapsin I (protein I), a nerve terminal-specific phosphoprotein. III. Its association with synaptic vesicles studied in a highly purified synaptic vesicle preparation. J Cell Biol. 1983 May;96(5):1374–1388. doi: 10.1083/jcb.96.5.1374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jahn R., Schiebler W., Ouimet C., Greengard P. A 38,000-dalton membrane protein (p38) present in synaptic vesicles. Proc Natl Acad Sci U S A. 1985 Jun;82(12):4137–4141. doi: 10.1073/pnas.82.12.4137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kelly R. B. The cell biology of the nerve terminal. Neuron. 1988 Aug;1(6):431–438. doi: 10.1016/0896-6273(88)90174-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  23. Lowe A. W., Madeddu L., Kelly R. B. Endocrine secretory granules and neuronal synaptic vesicles have three integral membrane proteins in common. J Cell Biol. 1988 Jan;106(1):51–59. doi: 10.1083/jcb.106.1.51. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Matteoli M., Haimann C., Torri-Tarelli F., Polak J. M., Ceccarelli B., De Camilli P. Differential effect of alpha-latrotoxin on exocytosis from small synaptic vesicles and from large dense-core vesicles containing calcitonin gene-related peptide at the frog neuromuscular junction. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7366–7370. doi: 10.1073/pnas.85.19.7366. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Matthew W. D., Tsavaler L., Reichardt L. F. Identification of a synaptic vesicle-specific membrane protein with a wide distribution in neuronal and neurosecretory tissue. J Cell Biol. 1981 Oct;91(1):257–269. doi: 10.1083/jcb.91.1.257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Morris J. F., Nordmann J. J., Dyball R. E. Structure-function correlation in mammalian neurosecretion. Int Rev Exp Pathol. 1978;18:1–95. [PubMed] [Google Scholar]
  27. Morris J. F., Nordmann J. J. Membrane recapture after hormone release from nerve endings in the neural lobe of the rat pituitary gland. Neuroscience. 1980;5(3):639–659. doi: 10.1016/0306-4522(80)90061-5. [DOI] [PubMed] [Google Scholar]
  28. Nagasawa J., Douglas W. W., Schulz R. A. Micropinocytotic origin of coated and smooth microvesicles ("synaptic vesicles") in neurosecretory terminals of posterior pituitary glands demonstrated by incorporation of horseradish peroxidase. Nature. 1971 Jul 30;232(5309):341–342. doi: 10.1038/232341a0. [DOI] [PubMed] [Google Scholar]
  29. Navone F., Greengard P., De Camilli P. Synapsin I in nerve terminals: selective association with small synaptic vesicles. Science. 1984 Dec 7;226(4679):1209–1211. doi: 10.1126/science.6438799. [DOI] [PubMed] [Google Scholar]
  30. Navone F., Jahn R., Di Gioia G., Stukenbrok H., Greengard P., De Camilli P. Protein p38: an integral membrane protein specific for small vesicles of neurons and neuroendocrine cells. J Cell Biol. 1986 Dec;103(6 Pt 1):2511–2527. doi: 10.1083/jcb.103.6.2511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Nordmann J. J., Chevallier J. The role of microvesicles in buffering [Ca2+]i in the neurohypophysis. Nature. 1980 Sep 4;287(5777):54–56. doi: 10.1038/287054a0. [DOI] [PubMed] [Google Scholar]
  32. Nordmann J. J., Morris J. F. Membrane retrieval at neurosecretory axon endings. Nature. 1976 Jun 24;261(5562):723–725. doi: 10.1038/261723a0. [DOI] [PubMed] [Google Scholar]
  33. Nordmann J. J., Schmid D. W., Chauveau J., Legros J. J. Neurophysin heterogeneity. Difference between newly formed and aged neurosecretory granules. J Biol Chem. 1984 Nov 10;259(21):13111–13116. [PubMed] [Google Scholar]
  34. Obata K., Kojima N., Nishiye H., Inoue H., Shirao T., Fujita S. C., Uchizono K. Four synaptic vesicle-specific proteins: identification by monoclonal antibodies and distribution in the nervous tissue and the adrenal medulla. Brain Res. 1987 Feb 24;404(1-2):169–179. doi: 10.1016/0006-8993(87)91368-0. [DOI] [PubMed] [Google Scholar]
  35. Rosa P., Hille A., Lee R. W., Zanini A., De Camilli P., Huttner W. B. Secretogranins I and II: two tyrosine-sulfated secretory proteins common to a variety of cells secreting peptides by the regulated pathway. J Cell Biol. 1985 Nov;101(5 Pt 1):1999–2011. doi: 10.1083/jcb.101.5.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Russell J. T., Thorn N. A. Adenosine triphosphate dependent calcium uptake by subcellular fractions from bovine neurohypophyses. Acta Physiol Scand. 1975 Mar;93(3):364–377. doi: 10.1111/j.1748-1716.1975.tb05825.x. [DOI] [PubMed] [Google Scholar]
  37. Scarfone E., Demêmes D., Jahn R., De Camilli P., Sans A. Secretory function of the vestibular nerve calyx suggested by presence of vesicles, synapsin I, and synaptophysin. J Neurosci. 1988 Dec;8(12):4640–4645. doi: 10.1523/JNEUROSCI.08-12-04640.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Südhof T. C., Czernik A. J., Kao H. T., Takei K., Johnston P. A., Horiuchi A., Kanazir S. D., Wagner M. A., Perin M. S., De Camilli P. Synapsins: mosaics of shared and individual domains in a family of synaptic vesicle phosphoproteins. Science. 1989 Sep 29;245(4925):1474–1480. doi: 10.1126/science.2506642. [DOI] [PubMed] [Google Scholar]
  39. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Tweedle C. D. Ultrastructural manifestations of increased hormone release in the neurohypophysis. Prog Brain Res. 1983;60:259–272. doi: 10.1016/S0079-6123(08)64395-2. [DOI] [PubMed] [Google Scholar]
  41. Wiedenmann B., Franke W. W. Identification and localization of synaptophysin, an integral membrane glycoprotein of Mr 38,000 characteristic of presynaptic vesicles. Cell. 1985 Jul;41(3):1017–1028. doi: 10.1016/s0092-8674(85)80082-9. [DOI] [PubMed] [Google Scholar]
  42. Wiedenmann B., Rehm H., Knierim M., Becker C. M. Fractionation of synaptophysin-containing vesicles from rat brain and cultured PC12 pheochromocytoma cells. FEBS Lett. 1988 Nov 21;240(1-2):71–77. doi: 10.1016/0014-5793(88)80342-9. [DOI] [PubMed] [Google Scholar]
  43. Winkler H. The biogenesis of adrenal chromaffin granules. Neuroscience. 1977;2(5):657–683. doi: 10.1016/0306-4522(77)90022-7. [DOI] [PubMed] [Google Scholar]
  44. de Camilli P., Navone F. Regulated secretory pathways of neurons and their relation to the regulated secretory pathway of endocrine cells. Ann N Y Acad Sci. 1987;493:461–479. doi: 10.1111/j.1749-6632.1987.tb27231.x. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES