Biomedical subjects

M Quik

Publications and source records attributed to M Quik.

At least 73 records · Page 4Linked to original sources

PubMed1988

Neuronal nicotinic alpha-bungarotoxin sites.

At the vertebrate neuromuscular junction and in the electroplax of eel and electric fish, the nicotinic alpha-bungarotoxin site and the nicotinic receptor involved in synaptic transmission are very tightly coupled and, indeed, appear to be the same molecular component. On the other hand, the nature of the relationship between the nicotinic receptor mediating synaptic events and the nicotinic alpha-bungarotoxin binding site in nervous tissue has been a matter of controversy over the last few years. Experimental studies have been accumulating which suggest that in many neuronal tissues these two components are distinct molecular entities with their own unique regulation. However, it also appears that in other nervous tissues, possibly in species lower on the evolutionary scale, the toxin binding site is part of the nicotinic receptor. An evaluation of all available evidence would point to the conclusion that, in neuronal tissues, the nicotinic acetylcholine receptor involved in synaptic events and the nicotinic alpha-bungarotoxin site can exist both in a tightly coupled form and one in which the two sites are mutually distinct. The possible physiological significance of the nicotinic alpha-bungarotoxin site is discussed in light of current experimental data. Evidence is available which may imply that the alpha-toxin site, whether it is present as a distinct entity or in association with the nicotinic acetylcholine receptor, is involved in trophic or growth related activities, as well as in other cellular functions. The possibility of an endogenous ligand for the nicotinic alpha-bungarotoxin site is also discussed.

Acetylcholine↗

PubMed1988

Regulation of alpha-bungarotoxin sites in chromaffin cells in culture by nicotinic receptor ligands, K+, and cAMP.

Previous work had shown that incubation with the nicotinic antagonist d-tubocurarine resulted in a marked increase in alpha-bungarotoxin (alpha-BGT) binding in adrenal medullary chromaffin cells in culture; the possible molecular mechanisms involved in up-regulating the alpha-BGT sites were investigated. To determine whether changes in the extracellular K+ concentration could influence the number of toxin binding sites, the chromaffin cells were incubated in the presence of 2-50 mM K+; this resulted in an increase in alpha-BGT binding similar to that observed with the nicotinic antagonist. This enhanced binding was maximal with 20 mM K+ and was not due simply to a generalized ion effect, inasmuch as incubation of the cells with a concentration of Na+ of equivalent osmolarity did not alter alpha-BGT binding. Carbachol and the agonist nicotine completely prevented the K+-induced increase in the binding sites. In contrast to the marked up-regulation of the nicotinic alpha-BGT sites by K+, this agent did not increase the acetylcholine-induced release of [3H]noradrenaline from chromaffin cells in culture, further supporting the contention that the nicotinic alpha-BGT site and the functional nicotinic receptor are distinct. The increases in toxin binding due to K+ and d-tubocurarine were partially additive, suggesting that d-tubocurarine and K+ may share a common pathway, but only to a small degree. The calcium channel agonist BAY K 8644 and antagonist D600 had no effect on alpha-BGT binding either alone or in the presence of K+ or d-tubocurarine. On the other hand, forskolin, an activator of adenylate cyclase, and dibutyryl cAMP, an analog of cAMP, partially prevented the K+ and the d-tubocurarine-induced increases in toxin binding. These results suggest an involvement of cAMP in both the nicotinic antagonist-induced and K+-induced up-regulation of the sites. The observation that several mechanisms exist for the fine regulation of the nicotinic alpha-BGT binding sites in adrenal chromaffin cells could imply that this nicotinic receptor population plays a role in this tissue.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1987

Dopamine D2 receptor binding in adrenal medulla: characterization using [3H]spiperone.

The possibility that dopamine may function as a neuromodulator or neurotransmitter in the adrenal gland, and not merely serve as a precursor to the catecholamines, has been suggested. If this hypothesis is correct, receptors for dopamine should be identifiable in the adrenal. The present work demonstrates the existence of a high-affinity receptor in adrenal medulla using [3H]spiperone as the radioligand to label the receptors. [3H]Spiperone bound rapidly, reversibly, and with high affinity to bovine adrenal medullary membranes. Scatchard analysis yielded a Kd of 0.09 nM and a Bmax of 51 fmol/mg protein. In competition binding experiments, dopaminergic antagonists were at least 100 times more potent in displacing [3H]spiperone from its binding sites than adrenergic or serotonergic receptor antagonists. Similarly, agonists at the dopamine receptor more readily competed for [3H]spiperone binding than other receptor agonist drugs tested. Furthermore, D2 selective antagonists and agonists were much more potent than D1 receptor ligands. These results suggest that [3H]spiperone may bind to a high-affinity D2 dopamine receptor in adrenal medulla.

Adrenal Medulla↗

PubMed1987

Target organ destruction enhances recovery of choline acetyltransferase activity in adult rat sympathetic ganglia after denervation.

We studied the effect of destruction of the adrenergic neuronal population on the recovery of preganglionic choline acetyltransferase activity in adult rat sympathetic ganglia. To produce a partial destruction of the adrenergic system, rats were injected with guanethidine for 4 weeks; the preganglionic nerve to the superior cervical ganglion was then crushed and the guanethidine injections were continued for an additional 3 days to 6 weeks. To determine that the drug was effective, tyrosine hydroxylase activity was assessed; enzymic activity was reduced by 76% or more after guanethidine administration. In addition, electron microscopy studies showed that the number of principal cell-synaptic contacts and vesicle-containing varicosities were decreased by 90% after guanethidine administration. Those measures indicated the drug effectively destroyed the postsynaptic adrenergic neurons. In contrast, crushing the preganglionic nerve in animals not treated with guanethidine did not change tyrosine hydroxylase activity, suggesting minimal nonspecific damage to the ganglion as a result of the lesion. Choline acetyltransferase activity was measured as an index of presynaptic cholinergic integrity. After crush of the preganglionic nerve, there was a gradual recovery of ganglionic choline acetyltransferase activity in the saline-injected rats from 5% of control 3 days after the crush to 49% of control after 6 weeks. On the other hand, in the ganglia of rats administered guanethidine, there was a much enhanced recovery of choline acetyltransferase activity after the nerve crush compared with saline-injected animals; in the guanethidine-injected rats, the ganglionic choline acetyltransferase activity 3 days and 6 weeks after the nerve crush was 15 and 96%, respectively, compared with the uncrushed side. These results demonstrate after destruction of the adrenergic target tissue, recovery of presynaptic choline acetyltransferase activity in the adult rat sympathetic ganglion can still occur after denervation; however, the mechanism(s) that controls the regeneration is altered, so that enzymic activity is enhanced.

Animals↗

PubMed1987

Evidence for an involvement of membrane lipids in the control of neuronal nicotinic receptor function using bungarotoxin II-S1.

Previous work has shown that a toxin fraction, bungarotoxin (BGT) II-S1, isolated from Bungarus multicinctus venom could inhibit nicotinic receptor-mediated function. Experimental evidence suggested that this effect of the toxin might be due to a direct interaction of the toxin at the acetylcholine binding site and/or to its phospholipase activity. The toxin's enzymic activity has been further characterized; it has phospholipase activity of the A2 type with a Vmax of 12 pmol/min/ng protein and a Km of 300 microM. Phospholipases can produce their effects on a tissue through a variety of mechanisms including the disruption of important lipid protein bonds or the production of free fatty acids which interact with the tissue. To test for this latter possibility, various concentrations of fatty acid-free bovine serum albumin were added to the incubation medium. Fatty acid-free bovine serum albumin partially reversed the inhibition of carbachol-stimulated 1-[1,2-3H(N)]amino-4-guanidobutane ([3H]agmatine) uptake (used as a measure of ion flux) into the ganglion produced by BGT II-S1 (1.0 microM). In an attempt to determine which fatty acids might be responsible for this effect, various fatty acids were added to the incubation medium and their effect on nicotinic receptor-mediated [3H]agmatine uptake determined. Arachidonic acid decreased amine uptake by approximately 50% over the control carbachol-stimulated uptake; linoleic and oleic acid, on the other hand, did not significantly affect the response. This observation could imply that arachidonic acid is the fatty acid produced by the action of BGT II-S1 on the tissue to mediate the toxin's inhibitory effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Action Potentials↗

PubMed1987

Characterization and localization of phospholipase A2 activity in sympathetic ganglia.

Superior cervical ganglion phospholipase A2 activity was characterized using 1-palmitoyl-2-[1-14C]arachidonoyl-sn-glycero-3-phosphocholine as a substrate. The enzyme activity exhibited linearity with interval of incubation and tissue concentration; there appeared to be two pH optima of the enzyme, at pH 6.0 and 9.0. A Lineweaver-Burk plot of the reciprocal of activity versus substrate concentration yielded an apparent Km of 0.53 mM and a Vmax of 5.3 nmol/h/mg of protein. The enzyme exhibited a partial Ca2+ dependence; in the absence of Ca2+ and presence of EGTA, activity was reduced by 40%. The phospholipase A2 activity was heat sensitive and was completely inactivated after treatment at 100 degrees C for 30 min. For determination of whether the enzyme had a preference for hydrolysis of specific fatty acid substituents in the 2 position of phosphatidylcholine, several different substrates were tested. The order of preference for hydrolysis by the ganglionic enzyme was 1-palmitoyl-2-[1-14C]arachidonoyl-sn-glycero-3-phosphocholine = 1-palmitoyl-2-[1-14C]linoleoyl-sn-glycero-3-phosphocholine greater than 1-palmitoyl-2-[1-14C]palmitoyl-sn-glycero-3-phosphocholine. For determination of the localization of the phospholipase A2 enzyme in sympathetic ganglia, two approaches were used. Guanethidine, which results in destruction of adrenergic cell bodies in sympathetic ganglia, was administered to rats; an approximately 50% decline in phospholipase A2 activity was observed after this treatment. In other experiments, the preganglionic nerve to the ganglion was sectioned in rats; after 2 weeks of denervation, no significant change in ganglionic phospholipase A2 activity was seen.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1987

Marked up-regulation of the beta-bungarotoxin site in adrenal chromaffin cells by specific nicotinic antagonists.

The effect of nicotinic antagonists was studied on various parameters of adrenal medullary chromaffin cells in culture. Incubation of the cells in culture with d-tubocurarine or mecamylamine for 0.5-6 days resulted in up to an 8-fold increase in the binding of alpha-bungarotoxin (alpha-BGT) to the cells; other ganglionic blockers, on the other hand, such as hexamethonium and dihydro-beta-erythroidine, had no effect. This enhanced binding was due to an increase in the number of alpha-BGT sites with little change in affinity of the ligand for the receptor. The nature of the increase in the number of toxin-binding sites in chromaffin cells observed after treatment with d-tubocurarine or mecamylamine was subsequently investigated. A direct interaction of the drug with the alpha-BGT recognition site was not required for the increase in receptor number to occur; mecamylamine did not affect binding of alpha-BGT to chromaffin cells in culture in competition binding experiments, although d-tubocurarine did compete with alpha-BGT for binding to its recognition site. The reversal of the antagonist-induced increase in the alpha-BGT-binding sites by nicotine and carbachol suggested it was mediated through an interaction at an acetylcholine receptor recognition site. The depolarizing agent veratridine greatly attenuated the increase in the number of toxin-binding sites in response to antagonists; this effect of veratridine could be reversed by tetrodotoxin. These latter findings indicate that neuronal excitability can influence the observed increase in the number of alpha-BGT sites after exposure of the cultures to nicotinic antagonists. The antagonist-induced increase in the alpha-BGT sites in the cells was not associated with an increased functional responsiveness of the cells to acetylcholine. The present results demonstrate that the number of alpha-BGT-binding sites in adrenal medullary chromaffin cells can increase dramatically in response to some, but not other, nicotinic antagonists by an interaction at a nicotinic acetylcholine recognition site. The differential effect of antagonists at the nicotinic-like alpha-BGT site and the functional nicotinic receptor suggests these two parameters are distinct. The unusually large alteration in receptor number may be related to the unique localization of the adrenal medulla and could infer that these receptors have a role in this tissue.

Adrenal Medulla↗

PubMed1986

Contrasting effect of a brain supernatant extract on neuronal vs neuromuscular alpha-bungarotoxin receptors.

The effects of a rat brain supernatant extract and a partially purified supernatant preparation from bovine brain were determined on the binding of [125I]alpha-bungarotoxin (alpha-BGT) to muscle membranes, as well as to membranes prepared from brain. In agreement with previous work, the supernatant preparations inhibited alpha-BGT binding to brain membranes in a dose-dependent fashion, (Brain Research, 245 (1982) 57-67); however, no significant effect of either of the preparations was observed on the binding of the toxin to muscle membranes. As well, the supernatant preparations did not affect binding of radiolabelled alpha-BGT to muscle cells in culture in competition binding experiments. The effect of long-term incubation of cells in culture with the supernatant preparations was subsequently determined. These studies showed that the binding of [125I]alpha-BGT increased markedly (300%) in the presence of a crude rat brain supernatant preparation, while incubation of the muscle cells in the presence of the partially purified bovine supernatant extract had no significant effect on radiolabelled toxin binding. In contrast, both the rat and bovine brain supernatant preparations significantly decreased (up to 65%) radiolabelled toxin binding to a cultured neuronal cell population, adrenal medullary chromaffin cells. These results suggest that an endogenous factor(s), present in brain extracts, differentially regulates the neuronal as compared to the neuromuscular nicotinic alpha-bungarotoxin binding sites.

Animals↗

PubMed1986

Modulation of the nicotinic alpha-bungarotoxin site in chromaffin cells in culture by a factor(s) endogenous to neuronal tissue.

An endogenous factor(s) which affects the in vitro binding of (alpha-BGT) to rat brain membranes has previously been found in brain supernatant. This fraction, as well as a partially purified preparation of this material from bovine brain, is here shown to affect the binding of alpha-BGT to chromaffin cell membranes. To study possible long term effects, the supernatant extract was added to adrenal medullary chromaffin cells in culture. The cells were incubated for several days and at the end of this time, the medium bathing the cells, which contained the endogenous factor(s), was removed and alpha-BGT binding to the cells measured. Binding to control cultures had shown that alpha-BGT bound to the chromaffin cells in a saturable manner, with high affinity (Kd = 1.5 nM) and the specificity of a nicotinic receptor ligand. After incubation of the cells with supernatant factor, a marked decline in the number of alpha-BGT binding sites was observed with no change in affinity. This does not appear to be due to a detrimental effect on the cells as cell number did not appear to be decreased in the cultures preincubated with the supernatant extract and the DNA and protein content were similar in the control and treated cultures. The possibility that there was some non-specific detrimental effect to the chromaffin cell membrane was considered; however, the stimulated release of noradrenaline from the cells was not affected by treatment of the cultures in the presence of the supernatant fractions. In addition, tyrosine hydroxylase activity was significantly increased in the treated cultures. D-Tubo-curarine, an antagonist at the acetylcholine receptor, caused an increase in alpha-BGT binding after 7 days of treatment, while the agonist nicotine and choline had no effect. These results suggest that in brain supernatant there may exist an endogenous factor(s), which may function in the regulation of the nicotinic-like alpha-BGT receptors in neuronal cell.

Adrenal Medulla↗

PubMed1985

Binding characteristics of the bungarotoxin fraction II-S1 to rat brain membranes.

Bungarotoxin fraction II-S1 (designated BGT II-S1), isolated from the venom of Bungarus multicinctus, appears to affect nicotinic transmission in rat sympathetic ganglia through its phospholipase activity. On the other hand, the present investigation suggests that other modes of interaction of this toxin with nervous tissue may also exist as, in a rat brain membrane preparation, binding of this toxin to a specific binding site can be demonstrated. In a buffer containing calcium, binding of [125I]BGT II-S1 saturated with a Bmax of approximately 16 fmol/mg protein and a Kd of 5 nM. This site did not appear to be directly linked to the nicotinic acetylcholine recognition site as the binding was not displaced by nicotinic agents; however, alpha-bungarotoxin, which interacts with a nicotinic-like site in neural tissue, did affect the binding and, conversely, BGT II-S1 inhibited the binding of [125I]alpha-bungarotoxin.

Acetophenones↗

PubMed1985

Inhibition of nicotinic receptor mediated ion fluxes in rat sympathetic ganglia by BGT II-S1 a potent phospholipase.

The mechanism of action of the bungarotoxin fraction II-S1 (BGT II-S1), which copurifies with alpha-bungarotoxin (alpha-BGT) and inhibits nicotinic transmission, has been further characterized. BGT II-S1 (1 microM) inhibited the carbachol (100 microM) or nicotine (50 microM) stimulated uptake of [3H]agmatine into rat sympathetic ganglia by 73% and 52%, respectively. These responses were inhibited 90% by D-tubocurarine (100 microM), but unaffected by alpha-BGT (1 microM) or atropine (10 microM), suggesting that BGT II-S1 affects nicotinic function at a postsynaptic site. Binding of physiologically active [125I]BGT II-S1 could be demonstrated to intact sympathetic ganglia; however, the binding could not be displaced by nicotinic agents, suggesting that BGT II-S1 is not interacting at the receptor. Because some neurotoxins produce their effect at the synapse through a phospholytic action, the phospholipase activity of BGT II-S1 was determined. The results demonstrate that BGT II-S1 is a very potent calcium dependent phospholipase. In addition, conditions which abolished the toxin's phospholytic activity prevented its effects on nicotinic transmission and on nicotinic receptor mediated ion fluxes. These include irreversible inhibition of enzymic activity by treatment of BGT II-S1 with p-bromophenacylbromide, as well as reversible inhibition of the phospholipase by substitution of Ba2+ or Sr2+ ions for Ca2+ ions in the physiological medium. Thus, in rat sympathetic ganglia, BGT II-S1 blocks the nicotinic receptor mediated movement of ions across the membrane. This is probably not due to a direct interaction at the nicotinic acetylcholine recognition site; rather, it may be an ion channel associated effect which is mediated by alterations in the phospholipid environment of the receptor complex or of the membrane. Although BGT II-S1 also has presynaptic actions, in a cultured system of postsynaptic cells, it could prove a useful tool to study the role of phospholipids in neuronal nicotinic receptor regulation.

Acetophenones↗

PubMed1984

Effect of chemical destruction of adrenergic neurones on some cholinergic mechanisms in adult rat sympathetic ganglia.

Rats were treated for 2-6 weeks with guanethidine after which their superior cervical ganglia were removed. Ganglionic tyrosine hydroxylase and alpha-bungarotoxin binding sites were reduced by the guanethidine treatment indicating adrenergic cell body destruction. Choline acetyltransferase activity and acetylcholine content of ganglia were not clearly changed by the guanethidine treatment, indicating that the drug does not destroy presynaptic terminals and that these presynaptic indicators do not adapt markedly to postsynaptic loss. The cholinesterase in the ganglia was reduced by guanethidine treatment, but such ganglia retained their ability to accumulate surplus acetylcholine when they were incubated with physostigmine. This is interpreted as indicating surplus acetylcholine accumulation is a presynaptic phenomenon. Choline uptake by resting ganglia was not reduced as a result of guanethidine treatment nor was it affected by preganglionic denervation. This is interpreted as indicating that during rest, choline uptake is into supporting cells or intraganglionic cells rather than cholinergic nerve terminals or adrenergic cell bodies.

Acetylcholine↗

PubMed1983

Selective destruction of the serotonergic fibers of the fornix-fimbria and cingulum bundle increases 5-HT1 but not 5-HT2 receptors in rat midbrain.

Selected and localized lesions of serotonergic (5-HT) neurons were made by microinjection of 5,7-dihydroxytryptamine (5,7-DHT), after pretreatment with desipramine, into the cingulum bundle and fornix-fimbria; these are the major serotonergic hippocampal inputs from the median raphe nucleus. Two weeks after the lesion, the binding of [3H]5-HT (5-HT1 receptor) was determined in the hippocampus which receives the afferent terminals and, in addition, in the septum/hypothalamus and midbrain from where the fibers originate. Scatchard analysis showed there was no significant change in binding parameters in the hippocampus; however, a significant increase was observed in the Bmax in the midbrain (38%) with no change in the KD. The caudate which receives 5-HT inputs via other 5-HT tracts was not affected by the lesion. The changes in 5-HT1 receptor number or affinity were not observed 6 days or 5 weeks after the lesion. The binding of the ligands [3H]spiroperidol and [3H]ketanserin to the 5-HT2 receptor population was also determined in the same brain areas; no changes in receptor binding occurred two weeks after the lesion. These experiments demonstrate that a selective lesion of the serotonergic system can increase 5-HT1 receptors in the midbrain, which contains the serotonin cell bodies. In addition, as 5-HT2 binding is not altered, this further supports the hypothesis that 5-HT1 and 5-HT2 receptors are distinct populations of receptors.

5,7-Dihydroxytryptamine↗

PubMed1982

Presence of an endogenous factor which inhibits binding of alpha-bungarotoxin 2.2 to its receptor.

Cerebral cortical membranes and supernatant from rat were prepared by centrifugation of tissue homogenates at 45,000 g for 10 min. The supernatant fraction thus obtained was found to significantly inhibit alpha-bungarotoxin binding to the membrane preparation. After a 3 min incubation period, the supernatant inhibited toxin binding by approximately 65%, while the inhibition declined to about 40% after 30 min of incubation, presumably due to the slow reversibility of alpha-bungarotoxin binding. The choice of buffer was found to be an important determinant of the degree of inhibition observed, with 10 mM Tris pH 7.4 providing the most effective condition. This inhibition of toxin binding to cortical membranes by the 45,000 g supernatant was shown not to be due to adsorption of the radiolabeled compound to soluble or residual particulate material in the supernatant fraction. Specificity of the supernatant for the alpha-bungarotoxin site was demonstrated; a supernatant fraction could be prepared which inhibited alpha-bungarotoxin binding by 50% but had no effect on [3H]spiroperidol (DA2 and 5-HT2), [3H]prazosin (alpha 1-adrenergic, [3H]5-hydroxytryptamine (5-HT1) and [3H]quinuclidinylbenzilate (muscarinic cholinergic) binding. The inhibition of toxin binding also occurred in several other CNS regions including hippocampus, brainstem, spinal cord and cerebellum with an 80 to 90% inhibition of binding occurring in the latter two regions. In addition, the 45,000 g cortical supernatant completely prevented the binding of alpha-bungarotoxin to extrajunctional neuromuscular receptors and inhibited the binding to junctional receptors by 50%. Supernatants prepared from heart, liver and kidney or bovine serum albumin, at a concentration similar to the supernatant fraction, did not alter radiolabeled toxin binding to cortical membranes, while supernatant prepared from striated muscle tissue was effective. These results suggest there may be an endogenous ligand for the alpha-bungarotoxin 2.2 binding site in tissues which receive nicotinic cholinergic innervation.

Animals↗

PubMed1982

The alpha-bungarotoxin site and its relation to the cholinergic and nerve growth factor mediated increases in tyrosine hydroxylase activity in cultures of sympathetic ganglia and chromaffin cells.

alpha-Bungarotoxin has been proposed to interact with a membrane site in neuronal tissue which has the characteristics of a nicotinic acetylcholine receptor and also a trophic receptor. A nerve cell function which is affected by both nicotinic stimulation and nerve growth factor is the induction of tyrosine hydroxylase. For this reason, alpha-bungarotoxin was tested on the carbachol-induced increase and nerve growth factor-mediated increase in tyrosine hydroxylase activity in two preparations of neuronal origin, organ cultures of rat superior cervical ganglia and cultured bovine adrenal medullary cells. The results demonstrate that the alpha-bungarotoxin site is not involved in tyrosine hydroxylase induction.

Animals↗

PubMed1982

Blockade of transmission in rat sympathetic ganglia by a toxin which co-purifies with alpha-bungarotoxin.

Bungarus multicinctus venom was fractionated into its toxin components using ion-exchange chromatography on CM-Sephadex. According to previous reports, rechromatography of fraction II on a CM cellulose column yields chemically homogenous alpha-bungarotoxin (II2) of molecular weight 9000. However, in our hands, using the identical purification procedure, two discrete proteins of molecular weight 9000 and 15,000 were obtained as demonstrated by SDS gel electrophoresis. Subsequent fractionation of this alpha-bungarotoxin fraction (II2) was achieved on Sephadex G-50. The 9000 weight component (labelled II-S2) was identical to alpha-bungarotoxin; at a concentration of 1 microgram/ml it blocked transmission at the neuromuscular junction but did not block nicotinic responses in rat sympathetic ganglia. Very different properties were exhibited by II-SI, the 15,000 molecular weight component; it inhibited ganglionic transmission but was ineffective at the neuromuscular junction at the same concentration (1 microgram/ml). BGT II-S1 was equipotent in blocking the ganglionic action potential in the presence or absence of eserine; thus, it is not acting as an acetylcholinesterase by increasing acetylcholine breakdown. In the presence of toxin, [3H]choline incorporation into ganglionic acetylcholine during preganglionic stimulation was not altered, suggesting that the toxin did not block transmission by a presynaptic mechanism. Thus, the site of action of the toxin appears to be postsynaptic although it did not affect depolarization of the ganglia induced by carbachol.

Acetylcholine↗

PubMed1981

A critical evaluation of the use of toxins from Dendroaspis viridis to block nicotinic responses at central and ganglionic synapses.

Previous work by other investigators has shown that toxins prepared from Dendroaspis viridis venom block cholinergic transmission at the neuromuscular junction, as well as nicotinic transmission in frog spinal cord and in snail neurons. This suggested that these ligands may be useful for studying nicotinic receptors in the central nervous system. Thus, Dendroaspis viridis venom was fractionated into its toxin components. Only one of the fractions possessed activity as assessed by: (1) inhibition of alpha-bungarotoxin (alpha-BGT) binding at the neuromuscular junction (25% at 50 microgram toxin/ml) or (2) inhibition of the ventral root--dorsal root potential (VR--DRP), a nicotinic response in frog spinal cord. However, in the spinal cord preparation, in addition ot this blockade of the nicotinic pathway, convulsant activity and an increase in the amplitude of other root potentials was observed. Binding experiments using [125I]dendrotoxin demonstrated that the labeled compound bound to central nervous tissue such as brain or spinal cord; this was not displaced by nicotine (10(-4) M) or D-tubocurarine (10 (-4) M), a nicotinic antagonist, indicating either non-specific binding or binding to a non-nicotinic receptor. These results thus suggest that toxins from Dendroaspis viridis venom are not suitable ligands for central nicotinic receptors. In addition, as experiments also demonstrated that the dendrotoxins did not block cholinergic transmission in frog sympathetic ganglia, it contraindicates their use at ganglionic nicotinic receptors.

Acetylcholine↗

PubMed1981

Convulsant and possible anticholinergic actions of dendrotoxin in the amphibian spinal cord.

1 Dendrotoxin (DTOX)6, 6a and 5,6-1, fractions of the venom isolated from the green mamba (Dendroaspis viridis) promoted both spontaneous and stimulus-coupled rhythmic activity and antagonized the cholinergically mediated ventral root-dorsal root potential (VR-DRP) of frog spinal cord. The different time course and reversibility of these two effects indicates that the toxin has two entirely separate sites of action on the frog spinal cord. 2 Since DTOX 6 neither blocked nor enhanced responses of ventral and dorsal roots to glutamate, gamma-aminobutyric acid (GABA), beta-alanine, glycine or aspartate, it is unlikely that its convulsant action resulted from an alteration of the postsynaptic actions of inhibitory or excitatory amino acids. 3 An alteration in the threshold for action potential generation could perhaps contribute to the convulsant action of DTOX 6, although other mechanisms such as blockade of the release of unspecified inhibitory substances cannot be excluded. 4 In addition to the lack of effect on amino acid responses, DTOX failed to block the polysynaptic DR-VRP or DR-DRP pathways, which are mediated at least in part by amino acid neurotransmitters. Although this would be consistent with a specific action of DTOX at the cholinergic synapse of the VR-DRP pathway, this site of action has not yet been demonstrated unequivocally. Other possible mechanisms whereby DTOX could block VR-DRP are discussed.

Animals↗