A Kinetic Model for the Action of Xylocaine on Receptors for Acetylcholine

Steinbach, Alan

doi:10.1085/jgp.52.1.162

Cited by 88 publications

(40 citation statements)

References 29 publications

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“…The site to which local anaesthetics (quinacrine included) bind on the receptor-rich membrane fragments is distinct from the receptor site for agonists [19]. It is probably functionally related to and possibly located on the ion-translocating device, or ionophore [28,29]. The failure to recover both the effect of local anesthetics on the affinity increase caused by agonists and the slow response monitored by quinacrine may explain the variable success of the attempts to 'reconstitute' the permeability response to cholinergic agonists.…”

Section: Discussionmentioning

confidence: 99%

Recovery of Some Functional Properties of the Detergent-Extracted Cholinergic Receptor Protein from Torpedo marmorata after Reintegration into a Membrane Environment

Briley¹,

Changeux

1978

Eur J Biochem

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The change of affinity of the acetylcholine receptor for agonists and the influence of local anaesthetics has been studied in detail in receptor-rich membranes. These properties are changed after solubilisation by ionic detergents.A method for reproducibly reintegrating the receptor protein into a lipid environment is described. Reintegration of the receptor results in partial recovery of the binding and fluorescence properties of the membrane-bound receptor protein. In particular, the slow affinity change caused by agonists can be recovered but not the effect of local anaesthetics on this change. The fluorescence response to cholinergic ligands of the reintegrated receptor protein labelled with quinacrine does not appear identical to that found with the native receptor-rich membranes. It is suggested that the failure to recover the sensitivity to local anaesthetics is at the origin of the difficulties to regain functional reconstitution.Many attempts to reconstitute a functional 'excitable' membrane from preparations of purified nicotinic receptor protein from fish electric organ have been reported in the literature (for review see [l] and [2]). Crude detergent extracts of membrane preparations, rich in acetylcholine receptor protein [3], receptor protein purified by affinity chromatography in the presence of detergent [4,5] and even proteolipid fractions prepared directly from the electric organ [6,7] have been used and in several instances recovery of a permeability response to cholinergic agonists was found [3,4,8] but the reproducibility of these results is often questionable [1,2].As a first step to understanding the several parameters involved, our approach has been to study 'the binding properties of the receptor protein for cholinergic ligands in the membrane-bound and detergentsolubilised states and reintegrated into a membrane environment. Conditions have been previously defined under which a reversible transition between highaffinity and low-affinity states of the receptor protein takes place by varying the concentration of detergent [9]. Subsequently, it was shown that such transitions between affinity states exist in 'native' receptor-rich membrane fragments [lo] and are also regulated by cholinergic agonists. In the course of these studies it was also noticed that, in the same membrane fragments, local anaesthetics stabilize, at equilibrium, the receptor in its high-affinity state.In this paper we describe the reintegration of the receptor protein from Torpedo miirmorata electric organ into a lipid environment and demonstrate that, under these conditions, the agonist-binding characteristics of the membrane-bound receptor and the agonist-dependent affinity change can be recovered. The characteristic effect of local anaesthetics on this transition, however, cannot be demonstrated with the reintegrated receptor protein. The failure to recover the sensitivity to local anaesthetics might be at the origin of the difficulty to regain the full permeability response. MATERIALS AND METHODS Preparation ...

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Section: Discussionmentioning

confidence: 99%

Recovery of Some Functional Properties of the Detergent-Extracted Cholinergic Receptor Protein from Torpedo marmorata after Reintegration into a Membrane Environment

Briley¹,

Changeux

1978

Eur J Biochem

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“…However this effect is probably accounted for by the prolongation of the terminal action potential (Adams, 1974a;Thomson & Turkanis, 1973). The observation that the main effect develops only following the onset of receptor activation recalls Steinbach's (1968b) …”

Section: Methodsmentioning

confidence: 97%

Drug blockade of open end‐plate channels.

Adams¹

1976

The Journal of Physiology

316

258

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4. Conductance changes produced by bath applied agonists were depressed by thiopentone, the effect becoming greater the higher the agonist concentration. This effect, and also the observation that the concentration of thiopentone required to depress the bath agonist response is much greater than the apparent dissociation constant for binding to active receptor-channel complexes calculated from kinetic measurements, suggest that the selectivity for binding to open receptor-channel complexes is very high.5. Methyprylone, which is structurally similar to the barbiturates, is only a weak antagonist and shows no interpulse interaction. It was predicted that methyprylone should produce fast and slow components in the e.p.c. decay, and this prediction was verified.6. In the presence of barbiturates large iontophoretic carbachol applications produce conductance changes which show fast and slow components. Under these conditions the effects of carbachol prepulses become complex. However the effects are qualitatively consistent with the notion that different components of the response are contributed by channels located at various distances from the iontophoretic pipette tip.

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“…Use dependence of non-competitive blocking agents is one of the characteristics of open channel blockade and is seen with local anaesthetic block of the nicotinic acetylcholine receptor (Steinbach, 1968;Skok, 1990) and with quaternary ammonium ion block of the delayed rectifier potassium channel (Hille, 1967;Armstrong, 1975;Stanfield, 1983). For different reasons both of these phenomena are voltage dependent.…”

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confidence: 99%

Multiple mechanisms of picrotoxin block of GABA‐induced currents in rat hippocampal neurons.

Yoon¹,

Covey²,

Rothman³

1993

The Journal of Physiology

131

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SUMMARY1. WAe have examined the effect of picrotoxin on GABA-induced currents in dissociated rat hippocampal neurons. In addition, we used the putative picrotoxin receptor antagonist, ox-isopropyl-Lx-methyl-y-butyrolactone (cIMGBL), and the picrotoxin agonist, /?-ethyl-/I-methyl-y-butyrolactone (,/EMGBL) to explore the mechanisms of picrotoxin's interaction with the GABA-Cl-receptor-ionophore complex.2. The picrotoxin block of GABA current was use dependent, suggesting that the site of picrotoxin block is exposed by the conformational change initiated by GABA binding to the receptor.3. The alkyl-substituted butyrolactone antagonist, aIMGBL, selectively blocked the use-dependent mechanism of picrotoxin effect. After the apparent complete inhibition of the use-dependent effect, there was a residual picrotoxin effect that was independent of the time or concentration of GABA application. This indicates that the picrotoxin block of the GABA current is mediated by two different mechanisms. xIMGBL influences just one of these mechanisms.4. The picrotoxin receptor agonist, 8JEMGBL, exclusively blocked the GABA current in a use-dependent manner. Consistent with a use-dependent mechanism, the rate of onset of block increased with GABA concentration. Surprisingly, the fraction of GABA current block decreased with increasing GABA concentration.5. These results suggest that the relationship of picrotoxin and y-butyrolactones with the GABA-Cl-receptor-ionophore is quite complex. They are consistent with at least two possible models of agonist-antagonist interactions. Both cases require different antagonist affinities for the various kinetic states of the GABA-Clreceptor-ionophore. However, there is no need to require that either picrotoxin or

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A Kinetic Model for the Action of Xylocaine on Receptors for Acetylcholine

Cited by 88 publications

References 29 publications

Recovery of Some Functional Properties of the Detergent-Extracted Cholinergic Receptor Protein from Torpedo marmorata after Reintegration into a Membrane Environment

Recovery of Some Functional Properties of the Detergent-Extracted Cholinergic Receptor Protein from Torpedo marmorata after Reintegration into a Membrane Environment

Drug blockade of open end‐plate channels.

Multiple mechanisms of picrotoxin block of GABA‐induced currents in rat hippocampal neurons.

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