Acetylcholine receptor kinetics

Adams, Patrick W.

doi:10.1007/bf01870902

Cited by 206 publications

(126 citation statements)

References 108 publications

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“…2A. Similarly, Anderson and Stevens (17) (6,16,(18)(19)(20)(21)(22)(23), our previously derived value for k+ (2) gives kL < 5 ms-1 and thus (1 -g) > 0.1. We therefore adopt a value for 1 -g between 0.1 and 0.5 and for k-between 0.7 and 5 ms 1.…”

Section: Chemical Kinetics Schemesupporting

confidence: 62%

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

Land¹,

Harris²,

Salpeter³

et al. 1984

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

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In previous papers we studied the rising phase of a miniature endplate current (MEPC) to derive diffusion and forward rate constants controlling acetylcholine (AcCho) in the intact neuromuscular junction. The present study derives similar values (but with smaller error ranges) for these constants by including experimental results from the fallins phase of the MEPC. We find diffusion to be 4 x 10-6 cm2 slightly slower than free diffusion, forward binding to be 3.3 x 107 M-s-, and the distance from an average release site to the nearest exit from the cleft to be 1.6 jtm. We also estimate the back reaction rates. Heidmann and Changeux (7).] In our approach we use the shape of the MEPC as a function of AcChoR concentration to derive forward binding rates as well as information on AcCho diffusion in the intact neuromuscular cleft. We thus complement the results obtained by chemical techniques and by single-channel recording. The derived parameters support our motivating assumptions of the "saturating disk model" used by Salpeter and colleagues (8-11) to explain the generation of the MEPC. METHODSWe recorded MEPCs by voltage clamp from the intercostal muscle of the lizard Anolis carolinensis, using experimental techniques as in the first two papers of this series (1, 2) with minor modifications.Three groups were used: (i) control muscle (esterases and receptor concentration unaltered); (ii) receptors intact and esterases fully inactivated [with 1 mM diisopropyl fluorophosphate (iPr2P-F) for 1 hr]; (iii) receptors partially inactivated [with 40 nM a-bungarotoxin (BTX) for 40 min] to -0.36 its normal value (papers 1 and 2) and esterases inactivated with iPr2P-F as in ii. The filtering conditions of the voltage clamp were modified from those used in papers 1 and 2 to improve the signal-to-noise ratio. Because the falling phase of MEPCs is slow relative to the rise time, the signals were low-pass-filtered at 1 kHz (two-pole Butterworth filter) and the digitizer sampling rate was reduced to 6.25 kHz. Even though the Nyquist frequency is only a factor of 3 above our filter cutoff, aliasing was judged to be less than 4% of peak current. All experiments were done at 22 ± 10C and -100 mV holding potential. Fall time (tf) values were obtained from the experimental MEPC as 1.23 x the 90-40% fall time. The factor 1.23 is the conversion from 90-40% to efolding time for a pure exponential. Fig. 1 shows six examples of measured MEPC traces, illustrating the three conditions used in the present study. Fig. 2 plots the relation between tf and MEPC amplitude Ac, and Table 1 gives the mean values of tf and Ac under these conditions. RESULTSWe confirm (12-15) that inactivation of esterases lengthens tf appreciably and that, when cr is reduced by BTX treatment, tf is somewhat shortened again (12, 15). Our additional observations are that, when esterases are intact, tf does not depend on Ac ( Fig. 2A), but after esterase inactivation tf increases with increasing Ac (Fig. 2B). Finally, decreasing a, with esterases inactivated, alm...

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Section: Chemical Kinetics Schemesupporting

confidence: 62%

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

Land¹,

Harris²,

Salpeter³

et al. 1984

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

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“…The degree of symmetry of the AChR molecule inferred from morphological and primary structure data, can thus be experimentally tested at this level. Furthermore, electrophysiological (review in [27][28][29]) as well as biochemical (review in [6,30]) evidence supports the view that the high-affinity NCB site is located within the ion channel. Photoactivatable NCBs are thus useful tools for direct identification of potential channel-forming elements within each of the AChR subunits [1,2,12,13].…”

Section: Discussionmentioning

confidence: 75%

The noncompetitive blocker [³H]chlorpromazine labels segment M2 but not segment M 1 of the nicotinic acetylcholine receptor α‐subunit

et al. 1989

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The membrane bound acetylcholine receptor from Torpedo marmorata was photolabeled by the noncompetitive channel blocker [~H]chlorpromazine under equilibrium conditions in the presence of the agonist carbamoylcholine. The radioactivity incorporated into the AChR subunits was reduced by addition of phencyclidine, a specific ligand for the high-affinity site for noncompetitive blockers. The ~t-subunit was purified and digested with trypsin and/or CNBr and the resulting fragments fractionated by HPLC. Sequence analysis resulted in the identification of Ser-248 as a major residue labeled by pH]chlorpromazine in a phencyclidine-sensitive manner. This residue is located in the hydrophobic and putative transmembrane segment M2 of the ~t-subunit, a region homologous to that containing the chlorpromazine-labeled Ser-262 in the Nchain [1] and the Ser-254 and Leu-257 in the//-chain [2]. Extended sequence analysis of the hydrophobic segment M 1 further showed that no labeling.occurred in this region.

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“…They can be blocked by nickel or cadmium [28,29]. Since these channels could have a cationic selectivity, we have tried various blockers known to affect the cationic channel of the nicotinic receptor (QX222, (+)-tubocurarine; review [30]) or that of photoreceptors (diltiazem, see [31]). None of these blockers affected Ca 2+ influx in macrophages.…”

Section: 3mentioning

confidence: 99%

Biphasic increase in intracellular calcium induced by platelet‐activating factor in macrophages

Randriamampita¹,

Trautmann²

1989

FEBS Letters

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In single mouse macrophages stimulated by platelet-activating factor (PAF), the intracellular calcium concentration (Cai) monitored with fura-2 at room temperature presents a biphasic increase, including a transient and a more sustained component. After pulse administration of PAF, the first phase lasts for a few seconds and reaches a peak value of 0.5-1 /~M Ca z+ at high PAF concentration. The amplitude of this peak is independent of extracellular Ca z÷ concentration, suggesting that the initial Ca 2÷ transient is due to the release of Ca 2÷ from intracellular stores. The second phase of the response lasts for several minutes; its maximum amplitude is reached 1-2 min after the brief initial PAF stimulation. This phase, suppressed in zero external Ca 2÷ and increased in 10 mM Ca z+, is probably due to influx of Ca 2÷ through the plasma membrane. This secondary Ca 2÷ increase is blocked by 10-50/tM lanthanum. At low PAF concentration, the initial Ca z+ transient is not followed by a second phase, showing that the initial rises of Ca 2÷ and of its activator (presumably inositol trisphosphate) are not sufficient to trigger the second phase of Ca 2÷ increase.

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Acetylcholine receptor kinetics

Cited by 206 publications

References 108 publications

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

The noncompetitive blocker [³H]chlorpromazine labels segment M2 but not segment M 1 of the nicotinic acetylcholine receptor α‐subunit

Biphasic increase in intracellular calcium induced by platelet‐activating factor in macrophages

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Acetylcholine receptor kinetics

Cited by 206 publications

References 108 publications

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

Diffusion and binding constants for acetylcholine derived from the falling phase of miniature endplate currents.

The noncompetitive blocker [3H]chlorpromazine labels segment M2 but not segment M 1 of the nicotinic acetylcholine receptor α‐subunit

Biphasic increase in intracellular calcium induced by platelet‐activating factor in macrophages

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The noncompetitive blocker [³H]chlorpromazine labels segment M2 but not segment M 1 of the nicotinic acetylcholine receptor α‐subunit