Minimum essential factors required for vesicle mobilization at hippocampal synapses

被引:12
作者
Mozhayeva, MG
Matos, MF
Liu, XR
Kavalali, ET
机构
[1] Univ Texas, SW Med Ctr, Ctr Basic Neurosci, Dallas, TX 75390 USA
[2] Univ Texas, SW Med Ctr, Dept Mol Genet, Dallas, TX 75390 USA
[3] Univ Texas, SW Med Ctr, Dept Physiol, Dallas, TX 75390 USA
关键词
synaptic vesicle mobilization; synaptic vesicle recycling; cell permeabilization; SNARE; FM1-43; hippocampal culture;
D O I
10.1523/JNEUROSCI.3801-03.2004
中图分类号
Q189 [神经科学];
学科分类号
071006 ;
摘要
Studies on the mechanisms that underlie the function of small central presynaptic terminals have been hampered by the inaccessibility of these synapses to soluble reagents. Here, we permeabilized hippocampal synapses in culture, manipulated their interior, and monitored the resulting changes in vesicle mobilization with the styryl dye FM2-10. Using this method, we found that 1 muM Ca2+ after incubation with GTP or GTP-gamma-S could mobilize similar to90% of the total recycling pool, whereas 1 muM Ca2+ application after dialysis of permeabilized synapses with GDP-beta-S mobilized similar to30% of the recycling vesicles, presumably corresponding to the readily releasable pool. In electron micrographs of permeabilized hippocampal synapses stimulated with 1 muM Ca2+, we could detect significant vesicle depletion after preincubation with GTP-gamma-S, whereas preincubation with GDP-gamma-S left the total vesicle pool relatively intact. Taken together, in this system replenishment of the readily releasable pool by the reserve vesicles was strictly GTP dependent. In contrast, vesicle replenishment and release did not require ATP or N-ethylmaleimide-sensitive factor (NSF); however, this process involved formation of new soluble NSF-attachment protein receptor (SNARE) complexes as judged by its sensitivity to tetanus toxin. These results suggest that in hippocampal synapses, vesicle mobilization and replenishment of the readily releasable pool require GTP and Ca2+ but do not necessitate ATP-dependent priming and SNARE recycling.
引用
收藏
页码:1680 / 1688
页数:9
相关论文
共 49 条
[1]   GRADUAL AND STEPWISE CHANGES IN THE MEMBRANE CAPACITANCE OF RAT PERITONEAL MAST-CELLS [J].
ALMERS, W ;
NEHER, E .
JOURNAL OF PHYSIOLOGY-LONDON, 1987, 386 :205-217
[2]   Reconstitution of regulated exocytosis in cell-free systems: A critical appraisal [J].
Avery, J ;
Jahn, R ;
Edwardson, JM .
ANNUAL REVIEW OF PHYSIOLOGY, 1999, 61 :777-807
[3]   2 ROLES FOR GUANINE-NUCLEOTIDES IN THE STIMULUS-SECRETION SEQUENCE OF NEUTROPHILS [J].
BARROWMAN, MM ;
COCKCROFT, S ;
GOMPERTS, BD .
NATURE, 1986, 319 (6053) :504-507
[4]  
BEAN BP, 1992, METHOD ENZYMOL, V207, P181
[5]   Imaging exocytosis and endocytosis [J].
Betz, WJ ;
Mao, F ;
Smith, CB .
CURRENT OPINION IN NEUROBIOLOGY, 1996, 6 (03) :365-371
[6]  
BITTNER MA, 1986, J BIOL CHEM, V261, P182
[7]   A reevaluation of the role of mitochondria in neuronal Ca2+ homeostasis [J].
Budd, SL ;
Nicholls, DG .
JOURNAL OF NEUROCHEMISTRY, 1996, 66 (01) :403-411
[8]   Rabphilin-3A: A multifunctional regulator of synaptic vesicle traffic [J].
Burns, ME ;
Sasaki, T ;
Takai, Y ;
Augustine, GJ .
JOURNAL OF GENERAL PHYSIOLOGY, 1998, 111 (02) :243-255
[9]   Synaptic vesicle endocytosis [J].
Cremona, O ;
DeCamilli, P .
CURRENT OPINION IN NEUROBIOLOGY, 1997, 7 (03) :323-330
[10]   Molecular determinants of presynaptic active zones [J].
Garner, CC ;
Kindler, S ;
Gundelfinger, ED .
CURRENT OPINION IN NEUROBIOLOGY, 2000, 10 (03) :321-327