Simultaneous Measurement of Intracellular Ca<sup>2+</sup> and Asynchronous Transmitter Release from the same Crayfish Bouton

Ravin, R.; Spira, Micha Ε.; Parnas, H.; Parnas, I.

doi:10.1111/j.1469-7793.1997.tb00001.x

Cited by 70 publications

(109 citation statements)

References 42 publications

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“…At both nearphysiological ( A) and cool ( B) temperatures, the amplitude of the ⌬F/F transient evoked by 500 stimuli increased as the frequency increased from 25 to 100 Hz. This frequency-dependence is similar to that measured in other nerve terminal preparations (Ravin et al, 1997;David et al, 1998). The peak amplitude of the ⌬F/F transient was always greater at the cooler temperature, but the difference was much greater at 100 Hz than at 25 Hz.…”

Section: ؉supporting

confidence: 87%

“…When certain nerve terminals are stimulated with prolonged depolarizing pulses or trains of action potentials, the average cytosolic [Ca 2ϩ ] measured with fluorescent indicators increases rapidly at first but then stabilizes at a plateau level until stimulation ceases (Steunkel, 1994;Ravin et al, 1997;David et al, 1998). This stabilization of average cytosolic [Ca 2ϩ ] during continued Ca 2ϩ influx is disrupted by agents that inhibit mitochondrial Ca 2ϩ uptake (Steunkel, 1994;David et al, 1998), suggesting that mitochondrial Ca 2ϩ uptake contributes importantly to sequestration of the Ca 2ϩ loads entering stimulated nerve terminals.…”

Section: Abstract: Mitochondria; Presynaptic Terminal; Motor Nerve Tmentioning

confidence: 99%

“…The resting potential was Ϫ75 mV. son, Ravin et al (1997) and Tang and Zucker (1997) measured cytosolic [Ca 2ϩ ] plateauing at ϳ0.66 -3.2 M during 20 -100 Hz stimulation of crayfish motor nerve terminals. Also, increases in cytosolic [Ca 2ϩ ] plateauing at ϳ0.35-0.6 M were measured in fura-2-filled rat neurohypophysial terminals (Steunkel, 1994) and bovine chromaffin cells (perforated patch; Smith, 1999) during large depolarizing pulses.…”

Section: ؉mentioning

confidence: 99%

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Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

2000

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Increases in cytosolic [Ca 2ϩ ] evoked by trains of action potentials (20-100 Hz) were recorded from mouse and lizard motor nerve terminals filled with a low-affinity fluorescent indicator, Oregon Green BAPTA 5N. In mouse terminals at nearphysiological temperatures (30-38°C), trains of action potentials at 25-100 Hz elicited increases in cytosolic [Ca 2ϩ ] that stabilized at plateau levels that increased with stimulation frequency. Depolarization of mitochondria with carbonylcyanide m-chlorophenylhydrazone (CCCP) or antimycin A1 caused cytosolic [Ca 2ϩ ] to rise to much higher levels during stimulation. ] measured with fluorescent indicators increases rapidly at first but then stabilizes at a plateau level until stimulation ceases (Steunkel, 1994;Ravin et al., 1997;David et al., 1998). This stabilization of average cytosolic [Ca 2ϩ ] during continued Ca 2ϩ influx is disrupted by agents that inhibit mitochondrial Ca 2ϩ uptake (Steunkel, 1994;David et al., 1998), suggesting that mitochondrial Ca 2ϩ uptake contributes importantly to sequestration of the Ca 2ϩ loads entering stimulated nerve terminals. Ca 2ϩ uptake via the mitochondrial uniporter is driven by the large negative potential (approximately Ϫ150 to Ϫ200 mV) created by proton transport across the inner mitochondrial membrane (for review, see Gunter and Pfeiffer, 1990).For some secretory cells, additional evidence for mitochondrial Ca 2ϩ uptake has been obtained using fluorescent or luminescent indicators localized within the mitochondrial matrix. Increases in matrix [Ca 2ϩ ] evoked by depolarization and/or hormones have been demonstrated in adrenal chromaffin cells (Babcock et al., 1997;Montero et al., 2000) and lizard motor nerve terminals (David et al., 1998). Mitochondrial Ca uptake has also been demonstrated by electron probe microanalysis of total Ca in frog sympathetic ganglion neurons fast-frozen after a 45 sec bath application of 50 mM K ϩ (Pivovarova et al., 1999). Simultaneous imaging of cytosolic and mitochondrial [Ca 2ϩ ] showed that, in lizard motor nerve terminals, mitochondrial Ca 2ϩ uptake begins after as few as 25-50 action potentials delivered at 50 -100 Hz, at approximately the same time that cytosolic [Ca 2ϩ ] reaches a plateau (David et al., 1998). In this preparation, as in crayfish motor nerve terminals, adrenal chromaffin cells, and several types of neuronal somata, mitochondria have been shown to be the dominant means of sequestering moderate to large Ca 2ϩ loads (Friel and Tsien, 1994;Werth and Thayer, 1994;White and Reynolds, 1995;Herrington et al., 1996;Park et al., 1996;Tang and Zucker, 1997;David, 1999;Colegrove et al., 2000).The present study was undertaken to measure cytosolic [Ca 2ϩ ] transients evoked by physiological stimulation in mammalian (mouse) motor nerve terminals and to determine whether mitochondrial Ca 2ϩ sequestration contributes to limiting the magnitude of these transients. We demonstrate that, at temperatures near physiological (33-38°C), the elevation of average cytosolic [Ca 2ϩ ] stab...

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Section: ؉supporting

confidence: 87%

Section: Abstract: Mitochondria; Presynaptic Terminal; Motor Nerve Tmentioning

confidence: 99%

Section: ؉mentioning

confidence: 99%

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Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

2000

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“…Although most of the studies have used mammalian preparations, this form of transmission is also observed at the frog and crayfish neuromuscular junction (Rahamimoff and Yaari, 1973;Ravin et al, 1997). Because of the ubiquitous nature of this form of transmission, it likely has an important role in information processing distinct from synchronous release.…”

Section: Discussionmentioning

confidence: 99%

Integration of Asynchronously Released Quanta Prolongs the Postsynaptic Spike Window

Iremonger¹,

Bains²

2007

Journal of Neuroscience

105

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Classically, the release of glutamate in response to a presynaptic action potential causes a brief increase in postsynaptic excitability. Previous reports indicate that at some central synapses, a single action potential can elicit multiple, asynchronous release events. This raises the possibility that the temporal dynamics of neurotransmitter release may determine the duration of altered postsynaptic excitability. In response to physiological challenges, the magnocellular neurosecretory cells (MNCs) in the paraventricular nucleus of the hypothalamus (PVN) exhibit robust and prolonged increases in neuronal activity. Although the postsynaptic conductances that may facilitate this form of activity have been investigated thoroughly, the role of presynaptic release has been largely overlooked. Because the specific patterns of activity generated by MNCs require the activation of excitatory synaptic inputs, we sought to characterize the release dynamics at these synapses and determine whether they contribute to prolonged excitability in these cells. We obtained whole-cell recordings from MNCs in brain slices of postnatal day 21-44 rats. Stimulation of glutamatergic inputs elicited large and prolonged postsynaptic events that resulted from the summation of multiple, asynchronously released quanta. Asynchronous release was selectively inhibited by the slow calcium buffer EGTA-AM and potentiated by brief high-frequency stimulus trains. These trains caused a prolonged increase in postsynaptic spike activity that could also be eliminated by EGTA-AM. Our results demonstrate that glutamatergic terminals in PVN exhibit asynchronous release, which is important in generating large postsynaptic depolarizations and prolonged spiking in response to brief, high-frequency bursts of presynaptic activity.

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“…Crayfish. The L1 bundle of the deep extensor abdominal muscles or the opener muscle of the first two walking legs of 4 -8-cm-long (head-to-tail) Procambarus clarkii crayfish (Shalom Hayat Biological Preparatory) were exposed and submerged in a modified Van Harreveled solution (10 Ϯ 1°C, unless otherwise stated) as described before (49,50).…”

Section: Methodsmentioning

confidence: 99%

Molecular mechanisms that control initiation and termination of physiological depolarization-evoked transmitter release

Kupchik

Rashkovan

Ohana

et al. 2008

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

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Ca 2؉ is essential for physiological depolarization-evoked synchronous neurotransmitter release. But, whether Ca 2؉ influx or another factor controls release initiation is still under debate. The time course of ACh release is controlled by a presynaptic inhibitory G protein-coupled autoreceptor (GPCR), whose agonist-binding affinity is voltage-sensitive. However, the relevance of this property for release control is not known. To resolve this question, we used pertussis toxin (PTX), which uncouples GPCR from its Gi/o and in turn reduces the affinity of GPCR toward its agonist. We show that PTX enhances ACh and glutamate release (in mice and crayfish, respectively) and, most importantly, alters the time course of release without affecting Ca 2؉ currents. These effects are not mediated by G␤␥ because its microinjection into the presynaptic terminal did not alter the time course of release. Also, PTX reduces the association of the GPCR with the exocytotic machinery, and this association is restored by the addition of agonist. We offer the following mechanism for control of initiation and termination of physiological depolarization-evoked transmitter release. At rest, release is under tonic block achieved by the transmitter-bound high-affinity presynaptic GPCR interacting with the exocytotic machinery. Upon depolarization, the GPCR uncouples from its G protein and consequently shifts to a low-affinity state toward the transmitter. The transmitter dissociates, the unbound GPCR detaches from the exocytotic machinery, and the tonic block is alleviated. The free machinery, together with Ca 2؉ that had already entered, initiates release. Release terminates when the reverse occurs upon repolarization.G protein-coupled receptor ͉ neurotransmitter release ͉ pertussis toxin ͉ presynaptic receptors C a 2ϩ influx is essential for physiological depolarizationinduced neurotransmitter (NT) release (1, 2). A broader, Ca 2ϩ voltage, hypothesis suggests that two factors control release: Ca 2ϩ and G protein-coupled receptors (GPCRs), whose agonist-binding affinity is voltage-dependent (3). The mechanism suggested for this control is as follows. (i) At resting potential and rest concentration (nMs) of transmitter, the release machinery (SNARE proteins and synaptotagmin) is under tonic block imposed by the transmitter-bound high-affinity (nMs) GPCR. (ii) Depolarization shifts the GPCR to a lowaffinity state ( Ms), resulting in rapid transmitter dissociation (it should be emphasized that at this stage release of NT did not occur yet, and the concentration of NT in the synapse is still in the nM range). (iii) The unbound GPCR detaches from the release machinery to relieve the tonic block. The free-release machinery together with Ca 2ϩ , which had already entered, initiates release. (iv) Upon repolarization, release terminates because the receptor returns to its high-affinity state and the tonic block is reinstated.Much of this suggested mechanism was supported experimentally (3-8) by using mainly the cholinergic neuromuscular junction (N...

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Simultaneous Measurement of Intracellular Ca²⁺ and Asynchronous Transmitter Release from the same Crayfish Bouton

Cited by 70 publications

References 42 publications

Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

Integration of Asynchronously Released Quanta Prolongs the Postsynaptic Spike Window

Molecular mechanisms that control initiation and termination of physiological depolarization-evoked transmitter release

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Simultaneous Measurement of Intracellular Ca2+ and Asynchronous Transmitter Release from the same Crayfish Bouton

Cited by 70 publications

References 42 publications

Stimulation-Evoked Increases in Cytosolic [Ca2+] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

Stimulation-Evoked Increases in Cytosolic [Ca2+] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

Integration of Asynchronously Released Quanta Prolongs the Postsynaptic Spike Window

Molecular mechanisms that control initiation and termination of physiological depolarization-evoked transmitter release

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Product

Resources

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Simultaneous Measurement of Intracellular Ca²⁺ and Asynchronous Transmitter Release from the same Crayfish Bouton

Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent

Stimulation-Evoked Increases in Cytosolic [Ca²⁺] in Mouse Motor Nerve Terminals Are Limited by Mitochondrial Uptake and Are Temperature-Dependent