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. 1994 Dec 1;127(5):1419–1433. doi: 10.1083/jcb.127.5.1419

Preferential localization of a vesicular monoamine transporter to dense core vesicles in PC12 cells

PMCID: PMC2120259  PMID: 7962100

Abstract

Neurons and endocrine cells have two types of secretory vesicle that undergo regulated exocytosis. Large dense core vesicles (LDCVs) store neural peptides whereas small clear synaptic vesicles store classical neurotransmitters such as acetylcholine, gamma-aminobutyric acid (GABA), glycine, and glutamate. However, monoamines differ from other classical transmitters and have been reported to appear in both LDCVs and smaller vesicles. To localize the transporter that packages monoamines into secretory vesicles, we have raised antibodies to a COOH- terminal sequence from the vesicular amine transporter expressed in the adrenal gland (VMAT1). Like synaptic vesicle proteins, the transporter occurs in endosomes of transfected CHO cells, accounting for the observed vesicular transport activity. In rat pheochromocytoma PC12 cells, the transporter occurs principally in LDCVs by both immunofluorescence and density gradient centrifugation. Synaptic-like microvesicles in PC12 cells contain relatively little VMAT1. The results appear to account for the storage of monoamines by LDCVs in the adrenal medulla and indicate that VMAT1 provides a novel membrane protein marker unique to LDCVs.

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Selected References

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  1. Amara S. G., Kuhar M. J. Neurotransmitter transporters: recent progress. Annu Rev Neurosci. 1993;16:73–93. doi: 10.1146/annurev.ne.16.030193.000445. [DOI] [PubMed] [Google Scholar]
  2. Anderson D. C., King S. C., Parsons S. M. Proton gradient linkage to active uptake of [3H]acetylcholine by Torpedo electric organ synaptic vesicles. Biochemistry. 1982 Jun 22;21(13):3037–3043. doi: 10.1021/bi00256a001. [DOI] [PubMed] [Google Scholar]
  3. Andersson P. O., Bloom S. R., Edwards A. V., Järhult J. Effects of stimulation of the chorda tympani in bursts on submaxillary responses in the cat. J Physiol. 1982 Jan;322:469–483. doi: 10.1113/jphysiol.1982.sp014050. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bauerfeind R., Régnier-Vigouroux A., Flatmark T., Huttner W. B. Selective storage of acetylcholine, but not catecholamines, in neuroendocrine synaptic-like microvesicles of early endosomal origin. Neuron. 1993 Jul;11(1):105–121. doi: 10.1016/0896-6273(93)90275-v. [DOI] [PubMed] [Google Scholar]
  5. 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]
  6. Burger P. M., Mehl E., Cameron P. L., Maycox P. R., Baumert M., Lottspeich F., De Camilli P., Jahn R. Synaptic vesicles immunoisolated from rat cerebral cortex contain high levels of glutamate. Neuron. 1989 Dec;3(6):715–720. doi: 10.1016/0896-6273(89)90240-7. [DOI] [PubMed] [Google Scholar]
  7. Burgess T. L., Kelly R. B. Constitutive and regulated secretion of proteins. Annu Rev Cell Biol. 1987;3:243–293. doi: 10.1146/annurev.cb.03.110187.001331. [DOI] [PubMed] [Google Scholar]
  8. Clift-O'Grady L., Linstedt A. D., Lowe A. W., Grote E., Kelly R. B. Biogenesis of synaptic vesicle-like structures in a pheochromocytoma cell line PC-12. J Cell Biol. 1990 May;110(5):1693–1703. doi: 10.1083/jcb.110.5.1693. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cutler D. F., Cramer L. P. Sorting during transport to the surface of PC12 cells: divergence of synaptic vesicle and secretory granule proteins. J Cell Biol. 1990 Mar;110(3):721–730. doi: 10.1083/jcb.110.3.721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. De Camilli P., Jahn R. Pathways to regulated exocytosis in neurons. Annu Rev Physiol. 1990;52:625–645. doi: 10.1146/annurev.ph.52.030190.003205. [DOI] [PubMed] [Google Scholar]
  11. Edwards R. H., Selby M. J., Mobley W. C., Weinrich S. L., Hruby D. E., Rutter W. J. Processing and secretion of nerve growth factor: expression in mammalian cells with a vaccinia virus vector. Mol Cell Biol. 1988 Jun;8(6):2456–2464. doi: 10.1128/mcb.8.6.2456. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Edwards R. H. The transport of neurotransmitters into synaptic vesicles. Curr Opin Neurobiol. 1992 Oct;2(5):586–594. doi: 10.1016/0959-4388(92)90023-e. [DOI] [PubMed] [Google Scholar]
  13. Erickson J. D., Eiden L. E., Hoffman B. J. Expression cloning of a reserpine-sensitive vesicular monoamine transporter. Proc Natl Acad Sci U S A. 1992 Nov 15;89(22):10993–10997. doi: 10.1073/pnas.89.22.10993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Feany M. B., Yee A. G., Delvy M. L., Buckley K. M. The synaptic vesicle proteins SV2, synaptotagmin and synaptophysin are sorted to separate cellular compartments in CHO fibroblasts. J Cell Biol. 1993 Nov;123(3):575–584. doi: 10.1083/jcb.123.3.575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fiedler K., Parton R. G., Kellner R., Etzold T., Simons K. VIP36, a novel component of glycolipid rafts and exocytic carrier vesicles in epithelial cells. EMBO J. 1994 Apr 1;13(7):1729–1740. doi: 10.1002/j.1460-2075.1994.tb06437.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Forgac M. Structure and function of vacuolar class of ATP-driven proton pumps. Physiol Rev. 1989 Jul;69(3):765–796. doi: 10.1152/physrev.1989.69.3.765. [DOI] [PubMed] [Google Scholar]
  17. Green S. A., Setiadi H., McEver R. P., Kelly R. B. The cytoplasmic domain of P-selectin contains a sorting determinant that mediates rapid degradation in lysosomes. J Cell Biol. 1994 Feb;124(4):435–448. doi: 10.1083/jcb.124.4.435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hell J. W., Maycox P. R., Jahn R. Energy dependence and functional reconstitution of the gamma-aminobutyric acid carrier from synaptic vesicles. J Biol Chem. 1990 Feb 5;265(4):2111–2117. [PubMed] [Google Scholar]
  19. Hsu S. M., Raine L., Fanger H. Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem. 1981 Apr;29(4):577–580. doi: 10.1177/29.4.6166661. [DOI] [PubMed] [Google Scholar]
  20. Isambert M. F., Gasnier B., Botton D., Henry J. P. Characterization and purification of the monoamine transporter of bovine chromaffin granules. Biochemistry. 1992 Feb 25;31(7):1980–1986. doi: 10.1021/bi00122a012. [DOI] [PubMed] [Google Scholar]
  21. Johnson R. G., Jr Accumulation of biological amines into chromaffin granules: a model for hormone and neurotransmitter transport. Physiol Rev. 1988 Jan;68(1):232–307. doi: 10.1152/physrev.1988.68.1.232. [DOI] [PubMed] [Google Scholar]
  22. Johnston P. A., Cameron P. L., Stukenbrok H., Jahn R., De Camilli P., Südhof T. C. Synaptophysin is targeted to similar microvesicles in CHO and PC12 cells. EMBO J. 1989 Oct;8(10):2863–2872. doi: 10.1002/j.1460-2075.1989.tb08434.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kanner B. I., Schuldiner S. Mechanism of transport and storage of neurotransmitters. CRC Crit Rev Biochem. 1987;22(1):1–38. doi: 10.3109/10409238709082546. [DOI] [PubMed] [Google Scholar]
  24. Kelly R. B. Secretory granule and synaptic vesicle formation. Curr Opin Cell Biol. 1991 Aug;3(4):654–660. doi: 10.1016/0955-0674(91)90037-y. [DOI] [PubMed] [Google Scholar]
  25. Kent U. M., Fleming P. J. Cytochrome b561 is fatty acylated and oriented in the chromaffin granule membrane with its carboxyl terminus cytoplasmically exposed. J Biol Chem. 1990 Sep 25;265(27):16422–16427. [PubMed] [Google Scholar]
  26. Liu Y., Peter D., Roghani A., Schuldiner S., Privé G. G., Eisenberg D., Brecha N., Edwards R. H. A cDNA that suppresses MPP+ toxicity encodes a vesicular amine transporter. Cell. 1992 Aug 21;70(4):539–551. doi: 10.1016/0092-8674(92)90425-c. [DOI] [PubMed] [Google Scholar]
  27. Liu Y., Roghani A., Edwards R. H. Gene transfer of a reserpine-sensitive mechanism of resistance to N-methyl-4-phenylpyridinium. Proc Natl Acad Sci U S A. 1992 Oct 1;89(19):9074–9078. doi: 10.1073/pnas.89.19.9074. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Martin J. L., Magistretti P. J. Pharmacological studies of voltage-sensitive Ca2+-channels involved in the release of vasoactive intestinal peptide evoked by K+ in mouse cerebral cortical slices. Neuroscience. 1989;30(2):423–431. doi: 10.1016/0306-4522(89)90262-5. [DOI] [PubMed] [Google Scholar]
  29. 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]
  30. Maycox P. R., Deckwerth T., Hell J. W., Jahn R. Glutamate uptake by brain synaptic vesicles. Energy dependence of transport and functional reconstitution in proteoliposomes. J Biol Chem. 1988 Oct 25;263(30):15423–15428. [PubMed] [Google Scholar]
  31. Milgram S. L., Eipper B. A., Mains R. E. Differential trafficking of soluble and integral membrane secretory granule-associated proteins. J Cell Biol. 1994 Jan;124(1-2):33–41. doi: 10.1083/jcb.124.1.33. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Milgram S. L., Mains R. E., Eipper B. A. COOH-terminal signals mediate the trafficking of a peptide processing enzyme in endocrine cells. J Cell Biol. 1993 Apr;121(1):23–36. doi: 10.1083/jcb.121.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. 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]
  34. Perin M. S., Fried V. A., Slaughter C. A., Südhof T. C. The structure of cytochrome b561, a secretory vesicle-specific electron transport protein. EMBO J. 1988 Sep;7(9):2697–2703. doi: 10.1002/j.1460-2075.1988.tb03123.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Rane S. G., Holz G. G., 4th, Dunlap K. Dihydropyridine inhibition of neuronal calcium current and substance P release. Pflugers Arch. 1987 Aug;409(4-5):361–366. doi: 10.1007/BF00583789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rudnick G. ATP-driven H+ pumping into intracellular organelles. Annu Rev Physiol. 1986;48:403–413. doi: 10.1146/annurev.ph.48.030186.002155. [DOI] [PubMed] [Google Scholar]
  37. Régnier-Vigouroux A., Tooze S. A., Huttner W. B. Newly synthesized synaptophysin is transported to synaptic-like microvesicles via constitutive secretory vesicles and the plasma membrane. EMBO J. 1991 Dec;10(12):3589–3601. doi: 10.1002/j.1460-2075.1991.tb04925.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Schweitzer E. S., Kelly R. B. Selective packaging of human growth hormone into synaptic vesicles in a rat neuronal (PC12) cell line. J Cell Biol. 1985 Aug;101(2):667–676. doi: 10.1083/jcb.101.2.667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Schweitzer E. S., Paddock S. Localization of human growth hormone to a sub-set of cytoplasmic vesicles in transfected PC12 cells. J Cell Sci. 1990 Jul;96(Pt 3):375–381. doi: 10.1242/jcs.96.3.375. [DOI] [PubMed] [Google Scholar]
  40. Stern-Bach Y., Greenberg-Ofrath N., Flechner I., Schuldiner S. Identification and purification of a functional amine transporter from bovine chromaffin granules. J Biol Chem. 1990 Mar 5;265(7):3961–3966. [PubMed] [Google Scholar]
  41. Südhof T. C., Jahn R. Proteins of synaptic vesicles involved in exocytosis and membrane recycling. Neuron. 1991 May;6(5):665–677. doi: 10.1016/0896-6273(91)90165-v. [DOI] [PubMed] [Google Scholar]
  42. Thureson-Klein A. Exocytosis from large and small dense cored vesicles in noradrenergic nerve terminals. Neuroscience. 1983 Oct;10(2):245–259. doi: 10.1016/0306-4522(83)90132-x. [DOI] [PubMed] [Google Scholar]
  43. Trimble W. S., Linial M., Scheller R. H. Cellular and molecular biology of the presynaptic nerve terminal. Annu Rev Neurosci. 1991;14:93–122. doi: 10.1146/annurev.ne.14.030191.000521. [DOI] [PubMed] [Google Scholar]
  44. Vincent M. S., Near J. A. Purification of a [3H]dihydrotetrabenazine-binding protein from bovine adrenal medulla. Mol Pharmacol. 1991 Dec;40(6):889–894. [PubMed] [Google Scholar]
  45. Walch-Solimena C., Takei K., Marek K. L., Midyett K., Südhof T. C., De Camilli P., Jahn R. Synaptotagmin: a membrane constituent of neuropeptide-containing large dense-core vesicles. J Neurosci. 1993 Sep;13(9):3895–3903. doi: 10.1523/JNEUROSCI.13-09-03895.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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