Rapid fluctuations in transmitter release from single vesicles in bovine adrenal chromaffin cells

Zhou, Zhuan; Misler, Stanley; Chow, Robert H.

doi:10.1016/s0006-3495(96)79718-7

Cited by 188 publications

(176 citation statements)

References 29 publications

(41 reference statements)

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“…These results are in agreement with recent findings suggesting that, in the immature hippocampus, silent synapses containing an appreciable number of AMPA receptors appear silent because the peak of glutamate concentration in the synaptic cleft is too low to activate low-affinity AMPA receptors (25). Low levels of glutamate could arise either from spillover of neurotransmitter from neighboring terminals (34)(35)(36) and͞or by the trickle of neurotransmitter from a fusion pore as occurs in neuronal (37) or non-neuronal secretory cells (38)(39)(40). According to the spillover hypothesis, only high-affinity NMDA receptors, but not low-affinity AMPA receptors, would be activated by the low concentration of glutamate diffused from a neighboring synapse (35,41).…”

Section: Discussionsupporting

confidence: 92%

“…However, at physiological temperature, spillover would be limited by the higher rate of glutamate clearance from the extracellular space (34,36), and, therefore, the contribution of this mechanism should be minimal. Alternatively, release of a small amount of glutamate from a nonexpanding fusion pore may account for some of the above observations (37)(38)(39)(40). Factors that increase the release probability would modify the gating properties of presynaptic fusion pores from the nonexpanding mode to a rapidly expanding one in the same way as recently suggested for LTP at silent synapses (25).…”

Section: Discussionmentioning

confidence: 72%

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Silent synapses in the developing hippocampus: Lack of functional AMPA receptors or low probability of glutamate release?

Gasparini

Saviane

Voronin

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

171

133

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At early developmental stages, silent synapses have been commonly found in different brain areas. These synapses are called silent because they do not respond at rest but are functional at positive membrane potentials. A widely accepted interpretation is that N-methyl-D-aspartate (NMDA) but not ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors are functionally expressed on the subsynaptic membrane. Here we show that, in both CA3 and CA1 hippocampal regions, AMPA-mediated synaptic responses can be detected already at early stages of postnatal development. However, some synapses appear silent because of a very low probability of glutamate release. They can be converted into functional ones by factors that enhance release probability such as paired-pulse stimulation, increasing the temperature or cyclothiazide (CTZ), a drug that blocks AMPA receptor desensitization and increases transmitter release. Conversely, conducting synapses can be switched off by increasing the frequency of stimulation. Although we cannot exclude that ''latent AMPA receptors'' can become functional after activity-dependent processes, our results clearly indicate that, in the neonatal hippocampus, a proportion of glutamatergic synaptic connections are presynaptically rather than postsynaptically silent.

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Section: Discussionsupporting

confidence: 92%

Section: Discussionmentioning

confidence: 72%

Silent synapses in the developing hippocampus: Lack of functional AMPA receptors or low probability of glutamate release?

Gasparini

Saviane

Voronin

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

171

133

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“…The percentage of such events that did not lead to full fusion was 11.6% for 140 NaCl, 8.9% for 90 NaCl, 7% for 40 NaCl and 6.8% for 0 NaCl. These frequencies are similar to those previously reported 17 . The mean amperometric current amplitudes of these events showed a similar decrease with decreasing [NaCl] from 4.9 +/− 0.4 pA (n=33) in 140 NaCl solution to 2.1+/−0.2 pA (n=31) in 0 NaCl solution.…”

Section: Flux Of Catecholamines Through the Fusion Pore Depends On Exsupporting

confidence: 92%

Exocytotic catecholamine release is not associated with cation flux through channels in the vesicle membrane but Na+ influx through the fusion pore

2007

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Release of charged neurotransmitter molecules through a narrow fusion pore requires charge compensation by other ions. It has been proposed that this may occur by ion flow from the cytosol through channels in the vesicle membrane, which would generate a net outward current. We tested this hypothesis in chromaffin cells using cell-attached patch amperometry measuring simultaneously catecholamine release from single vesicles and ionic current across the patch membrane. No detectable current was associated with catecholamine release indicating that <2% (if any) of cations enter the vesicle through its membrane. Instead we show that flux of catecholamines through the fusion pore, measured as an amperometric foot signal, decreases when the extracellular cation concentration is reduced. The results reveal that the rate of transmitter release through the fusion pore is coupled to net Na + influx through the fusion pore as predicted by electrodiffusion theory applied to fusion pore permeation and suggest a prefusion rather than postfusion role for vesicular cation channels.Neurotransmitter and hormone release occurs by exocytosis, which begins with the formation of a narrow fusion pore 1,2 . Fusion pores in mast cells and chromaffin cells have typically an initial conductance of ~330 pS 1,3 through which vesicular serotonin and catecholamines are released, respectively 4,5 . The initial fusion pore of a synaptic vesicle is typically >280 pS and release of neurotransmitter should occur with a time constant <500 μs due to the small vesicle volume 6 . Many neurotransmitters and hormones are organic ions that carry charge with them. Among these acetylcholine, serotonin and catecholamines are mostly monovalent cations at physiological or more acidic vesicular pH. Efflux of charged molecules through a narrow fusion pore would rapidly charge the small capacitance of the vesicle and a mechanism of charge compensation is required.Release of catecholamines from a single vesicle can be detected electrochemically 7 . The initial flux of catecholamines through the early narrow fusion pore can be measured as an amperometric foot signal preceding the amperometric spike 3,5,8 . The amplitude of amperometric foot currents is typically ~5 pA. Since most catecholamine molecules will be in monovalent cationic form but two electrons are transferred per molecule in the oxidation giving rise to the amperometric current 9 , the catecholamines released during the foot signal carry an ionic current of ~2.5 pA with them. This would charge a typical bovine chromaffin granule 4correspondence should be addressed to M.L. (ml95@cornell.edu).

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“…Early fusion pores may fluctuate, producing flickering amperometric foot currents in recordings from mast cells (Alvarez de Toledo et al, 1993) and chromaffin cells (Zhou et al, 1996), which reflect fluctuations in transmitter release due to fluctuations in fusion pore conductance (Albillos et al, 1997;Gong et al, 2007). The fusion pore conductance measurements showed three types of fusion pore expansion modes (Fig.…”

Section: Early Fusion Pore Conductance and Dynamicsmentioning

confidence: 99%

Positively Charged Amino Acids at the SNAP-25 C Terminus Determine Fusion Rates, Fusion Pore Properties, and Energetics of Tight SNARE Complex Zippering

et al. 2015

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SNAP-25 is a Q-SNARE protein mediating exocytosis of neurosecretory vesicles including chromaffin granules.Previous results with a SNAP-25 construct lacking the nine C terminal residues (SNAP-25⌬9) showed changed fusion pore properties (Fang et al., 2008), suggesting a model for fusion pore mechanics that couple C terminal zipping of the SNARE complex to the opening of the fusion pore. The deleted fragment contains the positively charged residues R198 and K201, adjacent to layers 7 and 8 of the SNARE complex. To determine how fusion pore conductance and dynamics depend on these residues, single exocytotic events in bovine chromaffin cells expressing R198Q, R198E, K201Q, or K201E mutants were investigated by carbon fiber amperometry and cell-attached patch capacitance measurements. Coarse grain molecular dynamics simulations revealed spontaneous transitions between a loose and tightly zippered state at the SNARE complex C terminus. The SNAP-25 K201Q mutant showed no changes compared with SNAP-25 wild-type. However, K201E, R198Q, and R198E displayed reduced release frequencies, slower release kinetics, and prolonged fusion pore duration that were correlated with reduced probability to engage in the tightly zippered state. The results show that the positively charged amino acids at the SNAP-25 C terminus promote tight SNARE complex zippering and are required for high release frequency and rapid release in individual fusion events.

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Rapid fluctuations in transmitter release from single vesicles in bovine adrenal chromaffin cells

Cited by 188 publications

References 29 publications

Silent synapses in the developing hippocampus: Lack of functional AMPA receptors or low probability of glutamate release?

Silent synapses in the developing hippocampus: Lack of functional AMPA receptors or low probability of glutamate release?

Exocytotic catecholamine release is not associated with cation flux through channels in the vesicle membrane but Na+ influx through the fusion pore

Positively Charged Amino Acids at the SNAP-25 C Terminus Determine Fusion Rates, Fusion Pore Properties, and Energetics of Tight SNARE Complex Zippering

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