Acetylcholine measured at short time intervals during transmission of nerve impulses in the electric organ of <i>Torpedo</i>

Dunant, Yves; Jones, G. J.; Loctin, F.

doi:10.1113/jphysiol.1982.sp014161

Cited by 31 publications

(22 citation statements)

References 21 publications

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“…The particles examined in the present work were probably not donated to the membrane by synaptic vesicles after a rapid fusion for the reasons given above. Moreover, vesicular acetylcholine is apparently a stable pool that is not immediately mobilized on activity (23), and an increase in the number of large IMPs was also found in a system where acetylcholine release was elicited in the absence of synaptic vesicles (8). The momentary increase in the number of large IMPs is not simply related to the depolarization of nerve endings since it did not take place in the absence of Ca, when the action potential still reached the terminals and depolarized them.…”

Section: Discussionmentioning

confidence: 98%

Brief occurrence of a population of presynaptic intramembrane particles coincides with transmission of a nerve impulse.

Michelet¹,

García‐Segura²,

Párducz³

et al. 1987

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

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Small pieces of Torpedo electric organ were cryofixed at 1-ms time intervals in a liquid medium at -190°C before, during, and after the passage of a single nerve impulse. In contrast to studies using this or other preparations, these experiments were done without 4-aminopyridine or other drugs that potentiate transmitter release. Freeze-fracture replicas were made from the most superficial layers of the tissue, where the rate of cooling was rapid enough to retain ultrastructure in the absence of chemical fixation. We found that the transmission of an impulse was accompanied by the momentary appearance of a population of large intramembrane particles in both the protoplasmic (P) and the external (E) leaflets of the presynaptic plasma membrane. The change was very brief, appearing soon after the stimulus artifact. It lasted for 2-3 ms. Large pits denoting vesicle openings at the presynaptic membrane were found in a small proportion of nerve terminals; their number did not increase during transmission of the nerve impulse. Reducing the temperature from 16 to 5°C slowed the time course of both the electrophysiological response and the change in intramembrane particles. The number of large particles did not increase when stimulation was applied in a low-Ca medium, a condition where the nerve terminals were still depolarized by the action potential but did not release the neurotransmitter. From these and other observations, we conclude that this transient change of intramembrane particles is closely linked to the mechanism of acetylcholine release at the nerve-electroplaque junction.Transmission of a nerve impulse at synapses takes only a few milliseconds. This makes it difficult to distinguish, with biochemical or morphological techniques, the changes that accompany transmission of the nerve impulse and from the changes that take place soon thereafter. The difficulty can be overcome by using rapid-freezing methods and by treating the synapse with drugs, such as 4-aminopyridine, which potentiate and prolong the release of acetylcholine. This strategy was used by Heuser et al. (1) in their freeze-fracture work on the frog neuromuscular junction. They found that during and soon after the passage ofan impulse there is a large increase in the number of vesicle openings in the active zones of the motor nerve terminals. Active zones are specialized areas of this membrane that are near a double row of synaptic vesicles (2). This was followed by the appearance, at the same place, of large intramembrane particles (IMPs; ref. 3). Vesicle openings were also found by in cryofixed neuromuscular junctions that had been stimulated in the presence of 4-aminopyridine and a high-Ca concentration. These authors used cryosubstitution since they found that freeze-fracture did not preserve the ultrastructure of nerve terminal membranes [due to the large size of the myofibers, only muscle fragments with nerve terminals against the coolant are expected to give well-preserved presynaptic membranes, and the protoplasmic (P)-face of th...

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Section: Discussionmentioning

confidence: 98%

Brief occurrence of a population of presynaptic intramembrane particles coincides with transmission of a nerve impulse.

Michelet¹,

García‐Segura²,

Párducz³

et al. 1987

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

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“…Starting with the whole tissue, it was shown that the characteristic decay curve of the electrical discharge of a stimulated electric organ slice is associated with a characteristic variation of the free AcCho compartment (9, 12, 13). The correlation was followed down to a few stimuli (14). The release of free AcCho from isolated cholinergic synaptosomes was more recently studied (15).…”

Section: Discussionmentioning

confidence: 99%

“…Free AcCho decreases and is renewed in the course of stimulation of electric organ slices (9,(12)(13)(14). The decrease of free AcChp has also been observed with stimulated synaptosomes, where it was directly measured with a chemiluminescent AcCho assay (15).…”

Section: Introductionmentioning

confidence: 96%

Reconstitution of a functional synaptosomal membrane possessing the protein constituents involved in acetylcholine translocation.

Israël¹,

Lesbats²,

Morel³

et al. 1984

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

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Reconstitution of a functional presynaptic membrane possessing calcium-dependent acetylcholine release properties has been achieved. The proteoliposomal membrane obtained gains its acetyicholine-releasing capabilities from presynaptic membrane proteins. At the peak of acetylcholine release, intramembrane particles became more numerous in one of the proteoliposomal membrane faces. This phenomenon resembles the intramembrane particle rearrangements found in stimulated synaptosomes. No visible structures capable of releasing acetylcholine as a result of the calcium influx were found inside the proteoliposomes. This supports the view that the release of free cytosolic acetylcholine from stimulated nerve terminals can be directly attributed to presynaptic membrane proteins. These proteins were extracted in a functional form from the synaptosomal membrane.The intramembrane particles found in the presynaptic membrane undergo important changes in the course of synaptic activity (1-8). They wvere analyzed during acetylcholine (AcCho) release from Torpedo electric organ synaptosomes stimulated with a variety of agents. The only common ultrastructural change found was the appearance of a category of large (8-to 18-nm) particles while smaller particles (5-11 nm) disappeared (6-8). The large particles can be pinched-off with the E or P faces of the membrane according to the conditions used. The hypothesis that Ca2t causes the assembly of large particles in the membrane and that these particles ensure Ca2+-dependent translocation of AcCho has been put forward by Israel et al. (6)(7)(8).At present, the view is generally accepted that there is, in the nerve terminal, a genuine cytosolic free AcCho compartment (20-50%) of total AcCho (9, 10) and that the enzyme cholineacetylase, which synthesizes it, is also located in the cytosol (11). Free AcCho decreases and is renewed in the course of stimulation of electric organ slices (9,(12)(13)(14). The decrease of free AcChp has also been observed with stimulated synaptosomes, where it was directly measured with a chemiluminescent AcCho assay (15). Cytosolic AcCho also seems to be involved at Aplysia synapses (16) or at neuromuscular junctions (17). As for the vesicular AcCho pool (bound AcCho), it is mobilized during intense stimulation in Torpedo electric organ (9) and at neuromuscular synapses (17). The possibility of direct release of free AcCho from the cytosolic compartment through a presynaptic membrane element (the operator) (13) is supported by recent experiments in which synaptosomes were depleted of their contents and filled with AcCho under controlled ionic conditions. These sacs were able to release AcCho in proportion to the Ca2t influx and the internal AcCho concentration (18). It therefore became possible to try to recoyer several essential presynaptic membrane mechanisms such as choline uptake, or AcCho release, in proteoliposomes derived from the presynaptic membrane (19-21). Until recently, proteoliposomes filled with AcCho or choline had been found to exhibi...

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“…Some of the results presented here have been published as short notes (Dunant, 1980;Dunant, Corthay, Eder & Loctin, 1980b).…”

Section: Introductionmentioning

confidence: 99%

“…It was therefore of interest to investigate whether significant changes in the level of tissue ACh could be detected during and after the course of such a giant and single e.p.p. This was done in the present study, in which we used the freezing technique described in the previous paper (Dunant, Jones & Loctin, 1982) to arrest metabolic processes at different precise moments after the stimulus.…”

Section: Introductionmentioning

confidence: 99%

Acetylcholine changes underlying transmission of a single nerve impulse in the presence of 4‐aminopyridine in Torpedo

Corthay¹,

Dunant²,

Loctin³

1982

The Journal of Physiology

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SUMMARY1. Transmission of a single nerve impulse has been investigated at the nerveelectroplaque junction of Torpedo marmorata in the presence of 4-aminopyridine (4-AP), a drug which powerfully potentiates evoked transmitter release.2. Three methodological approaches were used conjointly. These were (i) electrophysiological recording of the compound electroplaque potential (e.p.p.), (ii) radiochemical measurement of evoked acetylcholine (ACh) release and (iii) analysis of the content of ACh and ATP in the tissue at brief time intervals during the course of the e.p.p. and soon after. The last was achieved by using a stimulator coupled to a rapid tissue freezer.3. In the response to a single stimulus, 4-AP enhanced in a dose-dependent manner the size of the e.p.p., increasing the duration much more than the amplitude. At

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Acetylcholine measured at short time intervals during transmission of nerve impulses in the electric organ of Torpedo

Cited by 31 publications

References 21 publications

Brief occurrence of a population of presynaptic intramembrane particles coincides with transmission of a nerve impulse.

Brief occurrence of a population of presynaptic intramembrane particles coincides with transmission of a nerve impulse.

Reconstitution of a functional synaptosomal membrane possessing the protein constituents involved in acetylcholine translocation.

Acetylcholine changes underlying transmission of a single nerve impulse in the presence of 4‐aminopyridine in Torpedo

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