Nonequivalence of alpha-bungarotoxin receptors and acetylcholine receptors in chick sympathetic neurons.

Carbonetto, Salvatore; Fambrough, Douglas M.; Muller, Kenneth J.

doi:10.1073/pnas.75.2.1016

Cited by 123 publications

(45 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Earlier experiments showing specific and saturable binding sites for ␣-bungarotoxin could not find that the toxin affected some functions in several synapses (27)(28)(29)(30)(31)(32). However, more recent studies have provided data implicating ␣ 7 receptors in some neural functions.…”

Section: Discussionmentioning

confidence: 99%

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers

López

Montiel

Herrero

et al. 1998

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

105

View full text Add to dashboard Cite

show abstract

Section: Discussionmentioning

confidence: 99%

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers

López

Montiel

Herrero

et al. 1998

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

105

View full text Add to dashboard Cite

show abstract

“…If they bind specifically to the AcCho receptor, they will be useful agents for studying the distribution and regulation of this membrane component. a-Bungarotoxin [Bgt 2.2 (1)1, a small protein toxin isolated from the venom of the elapid snake Bungarus multicinctus, binds tightly and specifically to nicotinic acetylcholine (AcCho) re (2,3), rat superior cervical ganglia (4), and the pheochromocytoma cell line PC12 (5). The (6).…”

mentioning

confidence: 99%

Inhibition of neuronal acetylcholine sensitivity by α-toxins from Bungarus multicinctus venom

Ravdin

Berg

1979

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

146

View full text Add to dashboard Cite

Bungarus multicinctus venom contains several a-toxins in addition to the widelv used a-bungarotoxin (Bgt 2.2). We have found that two of the a-toxins (Bgt 3.1 and 3.3) inhibit neuronal acetylcholine (AcCho) sensitivity when tested on ciliary ganglion neurons in cell culture. Over 90% of the AcCho sensitivity recorded in response to iontophoretic application of AcCho was blocked when the neurons were incubated with either of the toxins at 10-7 M for I hr at 370C. component in the cell homogenate different from the component that bound Bgt 2.2 (6). It has been reported that high concentrations of Bgt 2.2 (10-6 M) can reversibly block nicotinic transmission through the ciliary ganglion (7). An earlier study found no effect of Bgt 2.2 on transmission through the ganglion (8).B. multicinctus venom contains several a-toxins that block AcCho receptors in skeletal muscle (1, 9) in addition to the widely used Bgt 2.2. We examined the effects of these toxins on ciliary ganglion neurons in cell culture to determine if they could also block neuronal AcCho receptors. Two of the a-toxins (Bgt 3.1 and ,3.3) were very effective in blocking sensitivity of the neurons to iontophoretically applied AcCho. Studies with rhodamine-labeled Bgt 2.2 demonstrated that the neurons also had a high-affinity binding site for Bgt 2.2, but concentrations of Bgt 2.2 adequate to saturate the site did not inhibit AcCho sensitivity. MATERIALS AND METHODSPreparation of Cell Cultures. Ciliary ganglion neuronmyotube cultures were prepared and maintained as described (10). Briefly, ciliary ganglia from 8-day-old chicken embryos were dissociated and plated at a density of about 104 neurons on 5-day-old myotube cultures (35-mm dishes) prepared from embryonic chicken pectoral muscle. The cultures were grown for 1-2 weeks in culture medium containing Eagle's minimal essential medium supplemented with 10% (vol/vol) horse serum, 5% (vol/vol) chicken embryo extract, 50 units of penicillin per ml, and 50 ,ug of streptomycin per ml.For experiments with fluorescence microscopy, glass-bottomed culture dishes were prepared by drilling an 18-mm hole in a 35-mm plastic culture dish and attaching a 25-mm Vanlab glass coverslip with Sylgard. The surface was coated with collagen and sterilized by UV irradiation. Cells were then plated and maintained in the cultures as described above.Purification of a-Toxins. a-Toxins from B. multicinctus venom were purified by ion-exchange chromatography as described (1), with the following modifications. The venom (20O mg in 6 ml) was applied to a column (40 X 2.5 cm) of carboxymethyl-Sephadex C-50 equilibrated with 50 mM ammonium acetate (pH 5.0). The column was eluted at 22 ml/hr with a 500-ml linear gradient of 50 mM ammonium acetate (pH 5.0) to 1.0 M ammonium acetate (pH 6.9). Fractions for peaks II and III (1) were pooled separately, lyophilized, and redissolved in 0.1 M ammonium acetate (pH 6.9). Further fractionation was achieved by chromatography on Whatman CM32 carboxymethyl-cellulose. Peak II was applied to a colu...

show abstract

“…Patrick and Stallcup [8], and Carbonetto et al [9], have reported that detergent-solubilized a-bungarotoxin receptors from rat and chick sympathetic neurons have a sedimentation coefficient greater than that of the nicotinic acetylcholine receptor from skeletal muscle (9.5-10s). This was also found with the retinal a-bungarotoxin receptor.…”

Section: Biochemical and Immunological Properties Of The A-bungarotoxmentioning

confidence: 99%

“…Sedimentation of detergent-solubilized receptors labelled with radioactive a-toxin was performed in 5 -20 sucrose gradients as described previously [25], except that the receptor-toxin interaction was stabilized by crosslinking with 0.1 % (wiv) glutaraldehyde according to [9] prior to centrifugation.…”

Section: Muterials and Generul M E T L M Kmentioning

confidence: 99%

Characterization of the α‐Bungarotoxin Receptor in Chick‐Embryo Retina

Betz

1981

European Journal of Biochemistry

View full text Add to dashboard Cite

Primary cultures and membranes fractions from chick embryo retina bind iodinated a-bungarotoxin, a highly selective ligand for nicotinic acetylcholine receptors fromskeletal muscle and fishelectric organ. The binding issaturable with anequilibrium dissociationconstant (Kd) of0.75 i 0.09nM. The pseudo-first-order rate constant(k + , )for binding at 37°C is 1.76 x lo5 M-' s-', the dissociation rate constant (k,) at 37,'C is 1.15 x s-'. Nicotinic cholinergic ligands and local anaesthetics inhibit a-bungarotoxin binding. In the case of carbamoylcholine, the inhibition of binding depends on the time of exposure to this cholinergic agonist. a-Bungarotoxin has no effect on the carbamoylcholine-induced stimulation in sodium permeability of cultured retinal neurons. On sucrose density gradients containing Triton X-I 00, the toxin binding site sediments with an apparent sedimentation coefficient of 10.5 -11 S. Detergent-solubilized a-bungarotoxin receptor crossreacts with antisera raised against nicotinic acetylcholine receptors from Torpedo murmoruta. These results are interpreted as indicating that a-bungarotoxin binds to retinal nicotinic acetylcholine receptors without affecting cholinergic receptor function.During the past decade, the nicotinic acetylcholine receptor from fish electric organ and skeletal muscle has been extensively purified and characterized in several laboratories (for reviews see [I, 21). This progress has become possible mainly through the use of a-toxins from the venoms of various elapid snake species which are highly selective ligands for this protein.One of these toxins, a-bungarotoxin from Bungurus multirinctus, has been most commonly used and shown to block the function of the muscle and fish acetylcholine receptor by binding to the cholinergic ligand recognition site of the receptor protein [I -31.Binding sites for a-bungarotoxin of nicotinic specificity are also widely distributed in the peripheral and central nervous systems of both vertebrates and invertebrates (for review see [4]). The nature of these toxin binding sites, though otten referred to as neuronal nicotinic acetylcholine receptors, remains, however, unsolved since their identification as cholinergic receptors relies almost exclusively on pharmacological binding experiments. Both electrophysiological and sodiumflux studies on spinal cord and sympathetic neurons have shown that a-bungarotoxin is unable to antagonize the activation of neuronal nicotinic acetylcholine receptors [5 -101. As an important argument against the identity of the neuronal a-bungarotoxin binding protein with a nicotinic acetylcholine receptor, Patrick and Stallcup [7] reported that antibodies to the cholinergic receptor from Electrophorus electricus failed to recognize the detergent-solubilized a-bungarotoxin binding site of PC12, a rat phaeochromocytoma cell line. The same antibodies, however, were effective in blocking the function of the nicotinic acetylcholine receptor present on these PC12 cells. Controversial observations suggesting a crossreactivity...

show abstract

Nonequivalence of alpha-bungarotoxin receptors and acetylcholine receptors in chick sympathetic neurons.

Cited by 123 publications

References 28 publications

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers

Inhibition of neuronal acetylcholine sensitivity by α-toxins from Bungarus multicinctus venom

Characterization of the α‐Bungarotoxin Receptor in Chick‐Embryo Retina

Contact Info

Product

Resources

About

Nonequivalence of alpha-bungarotoxin receptors and acetylcholine receptors in chick sympathetic neurons.

Cited by 123 publications

References 28 publications

Unmasking the functions of the chromaffin cell α 7 nicotinic receptor by using short pulses of acetylcholine and selective blockers

Unmasking the functions of the chromaffin cell α 7 nicotinic receptor by using short pulses of acetylcholine and selective blockers

Inhibition of neuronal acetylcholine sensitivity by α-toxins from Bungarus multicinctus venom

Characterization of the α‐Bungarotoxin Receptor in Chick‐Embryo Retina

Contact Info

Product

Resources

About

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers

Unmasking the functions of the chromaffin cell α ₇ nicotinic receptor by using short pulses of acetylcholine and selective blockers