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Biomedical subjects

A T Eldefrawi

Publications and source records attributed to A T Eldefrawi.

At least 55 records · Page 3Linked to original sources
PubMed1985

Comparison of the actions of carbamate anticholinesterases on the nicotinic acetylcholine receptor.

Neostigmine (Neo), pyridostigmine (Pyr), and physostigmine (Phy) at low concentrations inhibited acetylcholine (ACh) esterase, thereby indirectly potentiating ACh enhancement of [3H]perhydrohistrionicotoxin (H12-HTX) binding to the channel sites of the nicotinic ACh receptor of Torpedo membranes. However, at higher concentrations, they inhibited ACh action due to their direct binding to the ACh receptor. They displaced binding of [3H]ACh and 125I-alpha-bungarotoxin (alpha-BGT) to the receptor sites with the following order of decreasing potency: Neo greater than Phy greater than Pyr. Furthermore, Neo and Pyr potentiated [3H] H12-HTX binding to the receptor's channel sites. Preincubation of ACh receptors with any of the three carbamates reduced the rate of binding of 125I-alpha-BGT and increased the potency of carbamylcholine in inhibiting 125I-alpha-BGT binding, suggesting that the three carbamates act as partial agonists and potentiate receptor desensitization. Although none of the three carbamates inhibited [3H]H12-HTX binding to the receptor's closed channel conformation, only Phy was a potent inhibitor of [3H]H12-HTX binding to the carbamylcholine-activated conformation. The potency of Phy was not due to the absence of positive charge since Phy methiodide acted similarly. The data suggest that the major action of the three carbamates at nicotinic cholinergic synapses is inhibition of ACh-esterase. Their interactions with the nicotinic ACh receptor are with its "receptor" as well as allosteric "channel" sites, but they differ in their effects. Neo and Pyr act mainly as partial agonists, while Phy is mostly an inhibitor of the channel in the activated receptor conformation.

Acetylcholine↗
PubMed1984

Distinction between high-affinity [3H]phencyclidine binding sites and muscarinic receptors in guinea-pig ileum muscle.

[3H]Phencyclidine ([3H]PCP) binding was studied in guinea-pig ileum muscle membranes. Specific binding of [3H]PCP was time dependent, reversible and saturable, with an equilibrium dissociation constant of 154 nM and maximum binding of 12.9 pmol/mg of protein at pH 9. Its pH dependency suggests that the unionized PCP is the pharmacologically active form. The binding site was on a protein which was sensitive to heat, proteolytic enzymes and the carboxylic group reagent dicyclohexylcarbodiimide, but insensitive to phospholipase A and C, concanavalin A, dithiothreitol and N-ethylmaleimide. Specific [3H]PCP binding was displaced effectively by several PCP analogs and Ca++ channel antagonists including verapamil, to which these sites had a high affinity. These high-affinity PCP-binding sites were found at a much higher concentration in the same membrane preparation than muscarinic receptor sites identified by their specific binding of [3H]quinuclidinyl benzilate. PCP bound to both sites, but with a lower affinity to the muscarinic receptor sites. The PCP and muscarinic receptor sites differed in their sensitivities to pH and drug specifities .

Animals↗
PubMed1984

Interactions of phencyclidine with crayfish muscle membranes. Sensitivity to calcium channel antagonists and other drugs.

[3H]Phencyclidine ( [3H]PCP) bound to crayfish abdominal muscle membranes at pH 7.4 with two affinities (Kd of 0.96 nM for 0.38 pmole/mg of protein, and 18.9 nM for 7.6 pmoles/mg of protein). Binding affinities increased at higher pH, suggesting that binding may be due mostly to the un-ionized form of [3H]PCP. This high-affinity [3H]PCP binding was sensitive to the actions of trypsin, protease, and dicyclohexylcarbodiimide, but insensitive to phospholipase A, concanavalin A,N-ethylmaleimide, and dithiothreitol. Calcium channel antagonists were most potent in inhibiting the high-affinity [3H]PCP binding with the following descending order of potencies: bepridil greater than nicardipine = diltiazem = verapamil greater than cinnarizine greater than (+)-D-600 greater than (-)-D-600 greater than 4-NO2-nifedipine greater than 2-NO2-nifedipine. The binding was also highly sensitive to several PCP analogues, antipsychotics, piperocaine , and tilorone, and moderately sensitive to d-tubocurarine, atropine, imipramine, nortryptyline , and tetracaine. Although verapamil and nifedipine inhibited the action potential of crayfish muscle, PCP did not and actually prolonged slightly the falling phase of the action potential. Although it is unlikely that the [3H]PCP binding protein in crayfish muscles is a Ca2+ channel, it is possible that it may be a K+ channel.

Action Potentials↗
PubMed1983

Neurotransmitter receptors as targets for pesticides.

Nicotinic and muscarinic acetylcholine (ACh) receptors have been identified biochemically by means of their specific binding of [3H] alpha-bungarotoxin ([3H]alpha-BGT) and [3H]quinuclidinyl benzilate, respectively. There are some differences in the drug specificities, and sensitivities to active group reagents, of these receptors in insects when compared to those in vertebrates. Also, insect brain contains more nicotinic than muscarinic receptors, while the reverse is found in mammalian brain. Insect brain contains a third kind of putative ACh-receptor that is relatively soluble and is both nicotinic and muscarinic in its pharmacology but does not bind alpha-BGT. Toxic nicotine and analogs bind to it with high affinities. Several organophosphorus and carbamate insecticides and nereistoxin bind with high affinities to the nicotinic ACh-receptor of the electric organ of Torpedo. A few chlorinated hydrocarbon insecticides and derivatives interact with Torpedo nicotinic ACh-receptors, not at their 'receptor' sites but at their allosteric or 'channel' sites (which are identified by their specific binding of [3H]perhydrohistrionicotoxin). A few also bind to mammalian brain muscarinic receptors. The most potent on both receptors is the acaricide chlorobenzilate. Pyrethrins and synthetic pyrethroids also bind with high affinities to the channel sites of the Torpedo nicotinic ACh-receptor, though not to its receptor sites. Another group that binds to ACh-receptors is the organic and inorganic mercury compounds, which interact with both the Torpedo nicotinic and rat brain muscarinic receptors. Thus, neurotransmitter receptors act as molecular targets, primary or secondary for different pesticides.

Acetylcholinesterase↗
PubMed1982

Allethrin interactions with the nicotinic acetylcholine receptor channel.

Interactions of the synthetic pyrethroid allethrin with the nicotinic acetylcholine (ACh) receptor/channel were studied in membranes from Torpedo electric organ. Allethrin did not inhibit binding of [3H]ACh to the receptor sites, but inhibited noncompetitively binding of [3H]perhydrohistrionicotoxin ([3H]H12-HTX) to the ionic channel sites in a dose-dependent manner. The inhibition constant (Ki) of [3H]H12-HTX binding in absence of receptor agonist was 30 micro M, while in presence of 100 micro M carbamylcholine it was 4 micro M. This inhibitory effect of allethrin had a negative temperature coefficient. The high affinity binding of allethrin to the channel sites of the nicotinic ACh-receptor may be indicative of a postsynaptic site of action for pyrethroids, in addition to their known action on the sodium channel.

Acetylcholine↗
PubMed1982

Structure-activity relationships of amantadine. I. Interaction of the N-alkyl analogues with the ionic channels of the nicotinic acetylcholine receptor and electrically excitable membrane.

In this study the effects of amantadine (1-adamantanamine) and its N-alkyl-substituted analogues [N-methyl- (NMA), N-ethyl- (NEA), N-propyl- (NPA), N-butyl- (NBA), and N,N-diethyl-amantidine (NNDEA)] were investigated on ionic channels of the electrically excitable membrane and of the nicotinic acetylcholine (ACh) receptors in frog sartorius muscles and on the binding of perhydrohistrionicotoxin (H12-HTX) to isolated membranes of the electric organ of the electric ray Torpedo. Amantadine and each analogue blocked the indirectly elicited twitch, but NPA, NBA, and NNDEA also potentiated the directly elicited twitch. The order of potency in inhibiting the indirect twitch was: NEA = NPA = NNDEA (10 microM) greater than NMA (15 microM) greater than NBA (40 microM) much greater than amantadine (130 microM). Neither amantadine nor its N-alkyl analogues affected miniature end-plate potential frequency or resting membrane potential but decreased miniature end-plate potential amplitude. Each compound prolonged the directly elicited action potential but did not alter delayed rectification. All of the compounds induced a concentration-dependent depression of the peak end-plate current (EPC) amplitude at negative membrane potentials and induced nonlinearity in the response at membrane potentials more negative than -40 mV. The order of potency in inhibiting the EPC (at -90 mV) was NNDEA (less than 0.5 microM) greater than NPA (less than 1.0 microM) greater than NBA (less than 2.0 microM) greater than NEA (19 microM) greater than NMA (42 microM) greater than amantadine (64 microM). Only NPA, NBA, and NNDEA depressed the peak EPC amplitude at positive membrane potentials as well. The shortening of the time constant of EPC decay by all compounds was concentration-dependent. At the higher concentrations examined, the slope of the relationship between the time constant of decay and membrane potential was reversed for all compounds. Only NPA induced a double-exponential decay of the EPC at positive membrane potentials. Neither amantadine nor its N-alkyl analogues inhibited the binding of [3H]ACh to its receptor in Torpedo electroplax but they inhibited the binding of [3H]H12-HTX binding to the ionic channel sites of the ACh receptor. The Ki for inhibition of [3H]H12-HTX binding was NEA = NNDEA (15 microM) greater than NMA (30 microM) greater than NPA = NBA (40 microM) greater than amantadine (60 microM). A gross correlation exists between their ability to block the indirect muscle twitch, miniature end-plate potential amplitude, peak EPC amplitude and the binding of [3H]H12-HTX. But, no correlation was found between these potencies and their antiviral activity. It is suggested that these compounds may interact with the ionic channel of the ACh receptor in its open and closed conformation.

Action Potentials↗
PubMed1982

Interactions of d-tubocurarine with the nicotinic acetylcholine receptor/channel molecule.

The interactions of d-tubocurarine (d-TC) with the ionic channel of the nicotinic acetylcholine receptor were studied by biochemical methods in Torpedo electric organ membranes and by electrophysiological methods on frog sciatic nerve-sartorius muscle preparation. Torpedo membranes were treated with alpha-bungarotoxin to inhibit the acetylcholine receptor sites, then binding of [3H]perhydrohistrionicotoxin to the ionic channel sites was studied and found to be inhibited by d-TC. At 37 degrees C, the Ki of d-TC was 10 microM, and at 22 degrees C it was 100 microM. The affinity of d-TC for the ionic channel sites relative to that of perhydrohistrionicotoxin was constant at temperatures from 2-20 degrees C, but increased at higher temperatures up to 37 degrees C. The peak endplate current amplitude was depressed with 1 to 2 microM d-TC in a voltage-dependent manner, with considerable departure from linearity at 10 and 30 degrees C. The effect of d-TC on spontaneous miniature endplate currents was similar and slightly more potent. The time constant of endplate current decay was decreased by d-TC (1 and 2 microM) at temperatures of 10, 15 and 30 degrees C. The channel lifetime was reduced by d-TC, but channel conductance was unaffected. It is suggested that d-TC interacts with both the acetylcholine receptor sites as well as its ionic channel sites in closed and open conformations.

Amphibian Venoms↗
PubMed1982

Nicotinic acetylcholine receptor desensitization studied by [3H]perhydrohistrionicotoxin binding.

Desensitization of the nicotinic acetylcholine (ACh) receptor of Torpedo electric organs was studied by monitoring activated [3H]perhydrohistrionicotoxin binding to the channel sites of the receptor. Receptor desensitization was produced by incubating Torpedo membranes that had been pretreated with diisopropylfluorophosphate with ACh and was quenched by the addition of AChesterase. Desensitization caused concentration- and time-dependent decrease in the binding of [3H]perhydrohistrionicotoxin to carbamylcholine-activated receptors and an increase in the apparent affinity of carbamylcholine to the receptor. d-Tubocurarine and concanavalin A antagonized the desensitizing effects of ACh. The kinetics of this desensitization showed an extremely rapid temperature- and concentration-dependent component which was complete by the earliest measurement (i.e., 6 sec) and a much slower component (T1/2 = 55 sec) which reached a plateau after 120 sec. The rate of this slow component was temperature- and concentration-independent. Recovery from desensitization was temperature dependent, but independent of the ACh concentration used for desensitization. It also showed biphasic kinetics: half the receptors recovered fast (T1/2 = 36 sec), whereas the other was slow (T1/2 = 180 sec).

Acetylcholinesterase↗
PubMed1981

Regulation of [3H]perhydrohistrionicotoxin binding to Torpedo ocellata electroplax by effectors of the acetylcholine receptor.

The specific binding of [3H]perhydrohistrionicotoxin ([3H]H12-HTX) to the ionic channel of the nicotinic receptor in membranes from the electric organ of the electric ray Torpedo ocellata was studied by use of a rapid filter assay. The time course of the binding was monitored and the initial rate of binding (i.e. within 30 s) was found to be increased up to several hundredfold by the presence of several receptor agonists in a dose-dependent manner. Receptor antagonists blocked this agonist-increased initial rate of binding. The presence of receptor antagonists, with the exception of alpha-bungarotoxin, also increased the initial rate of [3H]H12-HTX binding, though to a much lesser degree than agonists. Preincubation of the membranes with carbamylcholine reduced the initial rate of binding in a time-dependent manner. This was proposed to be due to receptor desensitization. Thus, it is suggested that the time course of [3H]H12-HTX binding to the ionic channel sites is influenced by the conformational state of the receptor-channel complex such that receptor activation, inactivation, and desensitization appear to be reflected in [3H]H12-HTX binding.

Amphibian Venoms↗
PubMed1981

Recent advances in the molecular mechanisms of human and animal models of myasthenia gravis.

The receptor-channel molecule is a dynamic system which exists in multiple conformations and that is the way we should think of it when we study antibody interaction with the molecule. The results presented here suggest that some antibodies may affect receptor function by occupying sites other than the receptor site. Some of these sites may by exposed only in certain conformations, and occupation of some site by antibodies may effect conformational changes. These small but perhaps important differences in cholinergic channel properties of the myasthenic muscle from the normal one are revealed by studying the effect of myasthenic sera on drug interactions with the channel sites. The sera of myasthenics are able to react with certain channel conformations and are able to affect the interaction of channel antagonists such as H12HTX and QNB. The sera appear to act preferentially with the open conformation of the channel. As a consequence of such an effect, important conformational changes of the channel may fail to occur upon activation.

Acetylcholine↗