Presynaptic control of inhibitory neurotransmitter content in VIAAT containing synaptic vesicles

Unique pH dynamics in GABAergic synaptic vesicles illuminates the mechanism and kinetics of GABA loading

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1604527113 ◽

2016 ◽

Vol 113 (38) ◽

pp. 10702-10707 ◽

Cited By ~ 18

Author(s):

Yoshihiro Egashira ◽

Miki Takase ◽

Shoji Watanabe ◽

Junji Ishida ◽

Akiyoshi Fukamizu ◽

...

Keyword(s):

Synaptic Vesicles ◽

Hippocampal Neurons ◽

Glutamate Uptake ◽

Ph Sensor ◽

Mammalian Brain ◽

Gaba Uptake ◽

Inhibitory Neurotransmitter ◽

Luminal Ph ◽

Time Required ◽

Kinetics Of

GABA acts as the major inhibitory neurotransmitter in the mammalian brain, shaping neuronal and circuit activity. For sustained synaptic transmission, synaptic vesicles (SVs) are required to be recycled and refilled with neurotransmitters using an H+ electrochemical gradient. However, neither the mechanism underlying vesicular GABA uptake nor the kinetics of GABA loading in living neurons have been fully elucidated. To characterize the process of GABA uptake into SVs in functional synapses, we monitored luminal pH of GABAergic SVs separately from that of excitatory glutamatergic SVs in cultured hippocampal neurons. By using a pH sensor optimal for the SV lumen, we found that GABAergic SVs exhibited an unexpectedly higher resting pH (∼6.4) than glutamatergic SVs (pH ∼5.8). Moreover, unlike glutamatergic SVs, GABAergic SVs displayed unique pH dynamics after endocytosis that involved initial overacidification and subsequent alkalization that restored their resting pH. GABAergic SVs that lacked the vesicular GABA transporter (VGAT) did not show the pH overshoot and acidified further to ∼6.0. Comparison of luminal pH dynamics in the presence or absence of VGAT showed that VGAT operates as a GABA/H+ exchanger, which is continuously required to offset GABA leakage. Furthermore, the kinetics of GABA transport was slower (τ > 20 s at physiological temperature) than that of glutamate uptake and may exceed the time required for reuse of exocytosed SVs, allowing reuse of incompletely filled vesicles in the presence of high demand for inhibitory transmission.

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The Lateral Giant Fiber to Motor Giant Fiber Synapse in Crayfish

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100094103 ◽

1972 ◽

Vol 30 ◽

pp. 170-171

Author(s):

Charles A. Stirling

Keyword(s):

Synaptic Vesicles ◽

Procambarus Clarkii ◽

Synaptic Contact ◽

Giant Fiber ◽

Synaptic Junctions

The lateral giant (LG) to motor giant (MoG) synapses in crayfish (Procambarus clarkii) abdominal ganglia are the classic electrotonic synapses. They have previously been described as having synaptic vesicles and as having them on both the pre- and postsynaptic sides of symmetrical synaptic junctions. This positioning of vesicles would make these very atypical synapses, but in the present work on the crayfish Astacus pallipes the motor giant has never been found to contain any type of vesicle at its synapses with the lateral giant fiber.The lateral to motor giant fiber synapses all occur on short branches off the main giant fibers. Closely associated with these giant fiber synapses are two small presynaptic nerves which make synaptic contact with both of the giant fibers and with their small branches.

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Opening images of synaptic vesicles and calcium localization by means of microwave fixation method

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100134508 ◽

1990 ◽

Vol 48 (2) ◽

pp. 182-183

Author(s):

Vinci Mizuhira ◽

Hiroshi Hasegawa

Keyword(s):

Synaptic Vesicles ◽

Room Temperature ◽

Osmium Tetroxide ◽

Sampling Procedure ◽

Fixation Method ◽

Potassium Oxalate ◽

X Ray ◽

Cacodylate Buffer ◽

Ultrathin Sections ◽

Microwave Fixation

Microwave irradiation (MWI) was applied to 0.3 to 1 cm3 blocks of rat central nervous system at 2.45 GHz/500W for about 20 sec in a fixative, at room temperature. Fixative composed of 2% paraformaldehyde, 0.5% glutaraldehyde in 0.1 M cacodylate buffer at pH 7.4, also contained 2 mM of CaCl2 , 1 mM of MgCl2, and 0.1% of tannic acid for conventional observation; and fuether 30-90 mM of potassium oxalate containing fixative was applied for the detection of calcium ion localization in cells. Tissue blocks were left in the same fixative for 30 to 180 min after MWI at room temperature, then proceeded to the sampling procedure, after postfixed with osmium tetroxide, embedded in Epon. Ultrathin sections were double stained with an useal manner. Oxalate treated sections were devided in two, stained and unstained one. The later oxalate treated unstained sections were analyzed with electron probe X-ray microanalyzer, the EDAX-PU-9800, at 40 KV accelerating voltage for 100 to 200 sec with point or selected area analyzing methods.

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Evaluation of hippocampus in rhesus monkeys after chronic (1-year) inhalation exposure to marijuana: I. Electron microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100159539 ◽

1990 ◽

Vol 48 (3) ◽

pp. 398-399

Author(s):

A.M. Andrews ◽

S.W. Wilson ◽

A.C. Scallet ◽

S.F. Ali ◽

J. Bailey ◽

...

Keyword(s):

Synaptic Vesicles ◽

Rhesus Monkeys ◽

Low Dose ◽

Inhalation Exposure ◽

Synaptic Cleft ◽

High Dose ◽

Withdrawal Time ◽

Treatment Groups ◽

Ultrastructural Evidence ◽

Male Rhesus

Exposure of rhesus monkeys (Macaca mulatta) to marijuana via inhalation or to intravenous delta-9-tetrahydrocannabinol (THC), reportedly caused ultrastructural evidence of increased synaptic width. Chronic marijuana smoke in a single rhesus monkey examined after a six month withdrawal time caused ultrastructure changes in the septal, hippocampal and amygdala regions; the synaptic cleft was widened, electron opaque material was found in the cleft and in the pre- and postsynaptic regions, with some clumping of the synaptic vesicles. The objective of our study was to assess neuropathological alterations produced by chronic inhalation of marijuana smoke.Nineteen male rhesus monkeys, 3-5 years of age and weighing 3-8 kg, were divided into four treatment groups: a) sham control, b) placebo smoke (7 days/ week) c) low dose marijuana (2 times/week with 5 days/week sham) and d) high dose marijuana (7 times/week). A smoke exposure consisted of smoke from one cigarette (2.6% THC) burned down to 10 mm butt length. Smoke was administered via smoke generator (ADL II, Arthur D. Little, Inc. Cambridge, MA) and nose-mouth only masks (local production) equipped with one-way valves.

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