SpringerLink
Log in

Cholesterol Depletion from the Plasma Membrane Impairs Proton and Glutamate Storage in Synaptic Vesicles of Nerve Terminals

  • Published:
Journal of Molecular Neuroscience Aims and scope Submit manuscript

Abstract

We report that cholesterol depletion with methyl-β-cyclodextrin (MβCD) acutely applied to rat brain synaptosomes is accompanied by an immediate increase in transporter-mediated glutamate release and decrease in exocytotic release. To clarify the possible mechanisms underlying these phenomena, we investigated the influence of MβCD on synaptic vesicle acidification and exo/endocytotic process in nerve terminals. As shown by acridine orange fluorescence measurements, the application of MβCD to synaptosomes, as well as to isolated synaptic vesicles, led to the gradual leakage of the protons from the vesicles, whereas the application of MβCD complexed with cholesterol stimulated additional vesicle acidification and an increase in Ca2+-dependent exocytotic response. It was found that the treatment of nerve terminals with MβCD did not block Ca2+-triggered vesicle recycling. We suggest that cholesterol depletion of the plasma membrane with MβCD induces the removal of cholesterol from the membrane of synaptic vesicles resulting in immediate dissipation of synaptic vesicle proton gradient and redistribution of the neurotransmitter between the vesicular and cytosolic pools. The latter appears to be the main cause of a dramatic decrease in exocytotic and considerable increase in transporter-mediated release of l-[14C]glutamate.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

Abbreviations

MβCD:

Methyl-β-cyclodextrin

AO:

Acridine orange

∆μ H+ :

Synaptic vesicle electrochemical H+ gradient

DL-TBOA:

DL-threo-beta-benzyloxyaspartate

LDH:

Lactate dehydrogenase

References

  • Allen JA, Halverson-Tamboli RA, Rasenick MM (2007) Lipid rafts microdomains and neurotransmitter signaling. Nat Rev Neurosci 8:128–140

    Article  CAS  PubMed  Google Scholar 

  • Amador E, Dorfman LE, Wacker WE (1963) Serum lactic dehydrogenase activity: an analytical assessment of current assays. Clin Chem 12:391–399

    CAS  PubMed  Google Scholar 

  • Barnies K, Ingram J, Bennet M, Stewart G, Baldwin S (2004) Methyl-beta-cyclodextrin stimulates glucose uptake in clone 9 cells: a possible role for lipid rafts. Biochem J 378:343–351

    Article  Google Scholar 

  • Berman SB, Hastings TG (1997) Inhibition of glutamate transport in synaptosomes by dopamine oxidation and reactive oxygen species. J Neurochem 69:1185–1195

    Article  CAS  PubMed  Google Scholar 

  • Borisova T, Krisanova N, Sivko R, Borysov A (2010) Cholesterol depletion attenuates tonic release but increases the ambient level of glutamate in rat brain synaptosomes. Neurochem Int 56:466–478

    Article  CAS  PubMed  Google Scholar 

  • Burger K, Gimpl G, Fahrenholz F (2000) Regulation of receptor function by cholesterol. Cell Mol Life Sci 57:1577–1592

    Article  CAS  PubMed  Google Scholar 

  • Butchbach M, Tian G, Guo H, Lin CG (2004) Association of excitatory amino acid transporters, especially EAAT2, with cholesterol-rich lipid raft microdomains. J Biol Chem 279:34388–34396

    Article  CAS  PubMed  Google Scholar 

  • Cevc G, Richardsen H (1999) Lipid vesicles and membrane fusion. Adv Drug Deliv Rev 38:207–232

    Article  CAS  PubMed  Google Scholar 

  • Chattopadhyay A, Paila YD (2007) Lipid-protein interactions, regulation and dysfunction of brain cholesterol. Biochem Biophys Res Commun 354:627–633

    Article  CAS  PubMed  Google Scholar 

  • Cho WJ, Jeremic A, ** H, Ren G, Jena BP (2007) Neuronal fusion pore assembly requires membrane cholesterol. Cell Biol Int 31:1301–1308

    Article  CAS  PubMed  Google Scholar 

  • Christian AE, Haynes MP, Phillips MC, Rothblat GH (1997) Use of cyclodextrins for manipulating cellular cholesterol content. J Lipid Res 38:2264–2272

    CAS  PubMed  Google Scholar 

  • Churchward MA, Rogasevskaia T, Hofgen J, Bau J, Coorseen JR (2005) Cholesterol facilitates the native mechanism of Ca2+-triggered membrane fusion. J Cell Sci 118:4833–4848

    Article  CAS  PubMed  Google Scholar 

  • Cotman CW (1974) Isolation of synaptosomal and synaptic plasma membrane fractions. Meth Enzymol 31:445–452

    Article  CAS  PubMed  Google Scholar 

  • De Lorenzo RJ, Freedman SD (1978) Calcium dependent neurotransmitter release and protein phosphorylation in synaptic vesicles. Biochem Biophys Res Commun 80:183–192

    Article  Google Scholar 

  • Deutsch JW, Kelly RB (1981) Lipids of synaptic vesicles: relevance to the mechanism of membrane fusion. Biochemistry 20:378–385

    Article  CAS  PubMed  Google Scholar 

  • Dietschy JM, Turley SD (2001) Cholesterol metabolism in the brain. Curr Opin Lipidol 12:105–112

    Article  CAS  PubMed  Google Scholar 

  • Dröse S, Altendorf K (1997) Bafilomycins and concanamycins as inhibitors of V-ATPases and P-ATPases. J Exp Biol 200:1–8

    PubMed  Google Scholar 

  • Eroglu C, Brugger B, Wieland F, Sinning I (2003) Glutamate-binding affinity of Drosophila metabotropic glutamate receptor is modulated by association with lipid rafts. Proc Natl Acad Sci U S A 100:10219–10224

    Article  CAS  PubMed  Google Scholar 

  • Floor E, Leventhal PS, Schaeffer SF (1990) Partial purification and characterization of the vacuolar H(+)-ATPase of mammalian synaptic vesicles. J Neurochem 55:1663–1670

    Article  CAS  PubMed  Google Scholar 

  • Fong TM, McNamee MG (1986) Correlation between acetylcholine receptor function and structural properties of membranes. Biochemistry 25:830–840

    Article  CAS  PubMed  Google Scholar 

  • Gil C, Soler-Jover A, Blasi J (2005) Aguilera synaptic proteins and SNARE complexes are localized in lipid rafts from brain synaptosomes. Biochem Biophys Res Commun 329:117–124

    Article  CAS  PubMed  Google Scholar 

  • Gonzáles MI, Susarla BT, Fournier KM, Sheldon AL, Robinson MB (2007) Constitutive endocytosis and recycling of the neuronal glutamate transporter, excitatory amino acid carrier 1. J Neurochem 103:1917–1931

    Article  CAS  Google Scholar 

  • Hering H, Lin CC, Sheng M (2003) Lipid rafts in the maintenance of synapses, dendritic spines, and surface AMPA receptor stability. J Neurosci 23:3262–3271

    CAS  PubMed  Google Scholar 

  • Hill W, An B, Johnson J (2002) Endogenously expressed epithelial sodium channel is present in lipid rafts in A6 cells. J Biol Chem 277:33541–33544

    Article  CAS  PubMed  Google Scholar 

  • Huss M, Wieczorek H (2009) Inhibitors of V-ATPases: old and new players. J Exp Biol 212:341–346

    Article  CAS  PubMed  Google Scholar 

  • Jadot M, Andrianaivo F, Dubois F, Wattiaux R (2001) Effects of methylcyclodextrin on lysosomes. Eur J Biochem 268:1392–1399

    Article  CAS  PubMed  Google Scholar 

  • Jennings LJ, Xu QW, Firth TA, Nelson MT, Mawe GM (1999) Cholesterol inhibits spontaneous action potentials and calcium currents in guinea pig gallbladder smooth muscle. Am J Physiol 277:1017–1026

    Google Scholar 

  • Kato N, Nakanishi M, Hirashima N (2003) Cholesterol depletion inhibits store-operated calcium currents and exocytotic membrane fusion in RBL-2H3 cells. Biochemistry 42:11808–11814

    Article  CAS  PubMed  Google Scholar 

  • Klein U, Gimpl G, Fahrenholz F (1995) Alteration of the myometrial plasma membrane cholesterol content with β-cyclodextrin modulates the binding affinity of the oxytocin receptor. Biochemistry 34:13784–13793

    Article  CAS  PubMed  Google Scholar 

  • Krysanova NV, Sivko RV, Krupko OA, Borisova TA (2007) Methyl-beta-cyclodextrin influences glutamate transport in the rat brain nerve terminals by depletion of membrane cholesterol. Ukr Biokhim Zh 79:29–37

    CAS  PubMed  Google Scholar 

  • Lang T, Bruns D, Wenzel D et al (2001) SNAREs are concentrated in cholesterol-dependent clusters that define docking and fusion sites for exocytosis. EMBO J 20:2202–2213

    Article  CAS  PubMed  Google Scholar 

  • Lange Y, Ye J, Rigney M, Steck TL (1999) Regulation of endoplasmic reticulum cholesterol by plasma membrane cholesterol. J Lipid Res 40:2264–2269

    CAS  PubMed  Google Scholar 

  • Lange Y, Ye J, Steck TL (2004) How cholesterol homeostasis is regulated by plasma membrane cholesterol in excess of phospholipids. Proc Natl Acad Sci U S A 101:11664–11667

    Article  CAS  PubMed  Google Scholar 

  • Lange Y, Ye J, Steck TL (2005) Activation of membrane cholesterol by displacement from phospholipids. J Biol Chem 280:36126–36131

    Article  CAS  PubMed  Google Scholar 

  • Larson E, Howlett B, Jagendorf A (1986) Artificial reductant enhancement of the Lowry method for protein determination. Anal Biochem 155:243–248

    Article  CAS  PubMed  Google Scholar 

  • Launikonis BS, Stephenson DG (2001) Effects of membrane cholesterol manipulation on excitation-contraction coupling in skeletal muscle of the toad. J Physiol 534:71–85

    Article  CAS  PubMed  Google Scholar 

  • Linetska MV, Storchak LG, Tarasenko AS, Himmelreich NH (2004) Involvement of membrane GABA transporter in alpha-latrotoxin-stimulated [3H]GABA release. Neurochem Int 44:303–312

    Article  CAS  PubMed  Google Scholar 

  • Martens J, O’Connell K, Tamkun M (2004) Targeting of ion channels to membrane microdomains: localization of Kv channels to lipid rafts. Trends Pharmacol Sci 25:16–21

    Article  CAS  PubMed  Google Scholar 

  • Mauch DH, Nägler K, Schumacher S et al (2001) CNS synaptogenesis promoted by glia-derived cholesterol. Science 294:1354–1357

    Article  CAS  PubMed  Google Scholar 

  • Mitter D, Reisinger C, Hinz B et al (2003) The synaptophysin/synaptobrevin interaction critically depends on the cholesterol content. J Neurochem 84:35–42

    Article  CAS  PubMed  Google Scholar 

  • Pfrieger FW (2003) Cholesterol homeostasis and function in neurons of the central nervous system. Cell Mol Life Sci 60:1158–1171

    CAS  PubMed  Google Scholar 

  • Rodal SK, Skretting G, Garred O, Vilhardt F, van Deurs B, Sandvig K (1999) Extraction of cholesterol with methyl-beta-cyclodextrin perturbs formation of clathrin-coated endocytic vesicles. Mol Biol Cell 10:961–974

    CAS  PubMed  Google Scholar 

  • Rohrbough J, Broadie K (2005) Lipid regulation of the synaptic vesicle cycle. Nat Rev Neurosci 6:139–150

    Article  CAS  PubMed  Google Scholar 

  • Romanenko VG, Rothblat GH, Levitan I (2002) Modulation of endothelial inward-rectifier K+ current by optical isomers of cholesterol. Biophys J 83:3211–3222

    Article  CAS  PubMed  Google Scholar 

  • Roseth S, Fykse EM, Fonnum F (1995) Uptake of L-glutamate into rat brain synaptic vesicles: effect of inhibitors that bind specifically to the glutamate transporter. J Neurochem 65:96–103

    CAS  PubMed  Google Scholar 

  • Salaun C, James DJ, Chamberlain LH (2004) Lipid rafts and the regulation of exocytosis. Traffic 5:1–10

    Article  Google Scholar 

  • Salaun C, Gould GW, Chamberlain LH (2005) Lipid raft association of SNARE proteins regulates exocytosis in PC12 cells. J Biol Chem 280:19449–19453

    Article  PubMed  CAS  Google Scholar 

  • Sivko R, Krisanova N, Borisova T (2009) Reduced cholesterol content and the effects of inhibitors on Na+-dependent glutamate transport in rat brain nerve terminals. Neurobiol. Lipids 8(2):6–11. http://neurobiologyoflipids.org/content/8/2/

    Google Scholar 

  • Sooksawate T, Simmonds MA (2001) Effects of membrane cholesterol on the sensitivity of the GABA(A) receptor to GABA in acutely dissociated rat hippocampal neurones. Neuropharmacology 40:178–184

    Article  CAS  PubMed  Google Scholar 

  • Steck TL, Ye J, Lange Y (2002) Probing red cell membrane cholesterol movement with cyclopdextrin. Biophys J 83:2118–2125

    Article  CAS  PubMed  Google Scholar 

  • Subtil A, Gaidarov I, Kobylarz K, Lampson MA, Keen JH, McGraw TE (1999) Acute cholesterol depletion inhibits clathrin-coated pit budding. Proc Natl Acad Sci U S A 96:6775–6780

    Article  CAS  PubMed  Google Scholar 

  • Tarasenko AS, Linetska MV, Storchak LG, Himmelreich NH (2006) Effectiveness of extracellular lactate/pyruvate for sustaining synaptic vesicle proton gradient generation and vesicular accumulation of GABA. J Neurochem 99:787–796

    Article  CAS  PubMed  Google Scholar 

  • Taverna E, Saba E, Rowe J, Francolini M, Clementi F, Rosa P (2004) Role of lipid microdomains in p/q-type calcium channel (cav2.1) clustering and function in presynaptic membranes. J Biol Chem 279:5127–5134

    Article  CAS  PubMed  Google Scholar 

  • Thiele C, Hannah MJ, Fahrenholz F, Huttner WB (2000) Cholesterol binds to synaptophysin and is required for biogenesis of synaptic vesicles. Nat Cell Biol 2:42–49

    Article  CAS  PubMed  Google Scholar 

  • Trikash IO, Volynets GP, Remenyak OV, Gorchev VF (2008) Docking and fusion of synaptic vesicles in cell-free model system of exocytosis. Neurochem Int 53:401–407

    Article  CAS  PubMed  Google Scholar 

  • Waseem TV, Kolos VA, Lapatsina LP, Fedorovich SV (2006) Influence of cholesterol depletion in plasma membrane of rat brain synaptosomes on calcium-dependent and calcium-independent exocytosis. Neurosci Lett 405:106–110

    Article  CAS  PubMed  Google Scholar 

  • Wasser CR, Ertunc M, Liu X, Kavalali ET (2007) Cholesterol-dependent balance between evoked and spontaneous vesicle recycling. J Physiol 579.2:413–429

    Google Scholar 

  • **a F, Gao X, Kwan E et al (2004) Disruption of pancreatic β-cells lipid rafts modifies Kv2.1 channel gating and insulin exocyrtosis. J Biol Chem 279:24685–24691

    Article  CAS  PubMed  Google Scholar 

  • **a F, Leung YM, Gaisano G et al (2007) Targeting of Kv4, Cav1.2 and SNARE proteins to cholesterol-rich lipid rafts in pancreatic a-cells: effects on glucagons stimulus-secretion coupling. Endocrinology 148:2157–2167

    Article  CAS  PubMed  Google Scholar 

  • Yancey PG, Rodrigueza WV, Kilsdonk EP et al (1996) Cellular cholesterol efflux mediated by cyclodextrins. J Biol Chem 271:16026–16034

    Article  CAS  PubMed  Google Scholar 

  • Yoshinaka K, Kumanogoh H, Nakamura S, Maekawa S (2004) Identification of V-ATPase as a major component in the raft fraction prepared from the synaptic plasma membrane and the synaptic vesicle of rat brain. Neurosci Lett 363:168–172

    Article  CAS  PubMed  Google Scholar 

  • Zamir O, Charlton MP (2006) Cholesterol and synaptic transmitter release at crayfish neuromuscular junctions. J Physiol 571:83–99

    Article  CAS  PubMed  Google Scholar 

  • Zhang J, Feng Y, Forgac M (1994) Proton conduction and bafilomycin binding by the V0 domain of the coated vesicle V-ATPase. J Biol Chem 269:23518–23523

    CAS  PubMed  Google Scholar 

  • Zhou Q, Petersen CC, Nicoll RA (2000) Effects of reduced vesicular filling on synaptic transmission in rat hippocampal neurons. J Physiol 525:195–206

    Article  CAS  PubMed  Google Scholar 

  • Zidovetzki R, Levitan I (2007) Use of cyclodextrins to manipulate plasma membrane cholesterol content: evidence, misconceptions and control strategies. Biochim Biophys Acta 1768:1311–1324

    Article  CAS  PubMed  Google Scholar 

  • Zoccarato F, Cavallini L, Alexandre A (1999) The pH-sensitive dye acridine orange as a tool to monitor exocytosis/endocytosis in synaptosomes. J Neurochem 72:625–633

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We would like to thank Dr. Sergey Karakhim for the excellent technical assistance and help in confocal microscopy studies.

Author information

Authors and Affiliations

  1. Department of Neurochemistry, Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kiev, Ukraine

    Alla S. Tarasenko, Roman V. Sivko, Natalia V. Krisanova, Nina H. Himmelreich & Tatiana A. Borisova

Authors
  1. Alla S. Tarasenko
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Roman V. Sivko
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Natalia V. Krisanova
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nina H. Himmelreich
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Tatiana A. Borisova
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tatiana A. Borisova.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tarasenko, A.S., Sivko, R.V., Krisanova, N.V. et al. Cholesterol Depletion from the Plasma Membrane Impairs Proton and Glutamate Storage in Synaptic Vesicles of Nerve Terminals. J Mol Neurosci 41, 358–367 (2010). https://doi.org/10.1007/s12031-010-9351-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12031-010-9351-z

Keywords

Search

Navigation