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H Betz

Publications and source records attributed to H Betz.

At least 253 records · Page 14Linked to original sources
PubMed1984

Monoclonal antibodies and peptide mapping reveal structural similarities between the subunits of the glycine receptor of rat spinal cord.

The glycine receptor of rat spinal cord is an oligomeric membrane glycoprotein of molecular mass 250,000 daltons that contains three polypeptides of 48,000, 58,000, and 93,000 daltons. Monoclonal antibodies (mAbs) were prepared against the affinity-purified glycine receptor protein by using 125I-labeled receptor preparations for the detection of positive hybrids. From nine monoclonal antibodies obtained, six recognized denatured receptor polypeptides blotted to nitrocellulose paper. Two of these antibodies bound to more than one glycine receptor polypeptide: mAb GlyR 4a stained the 48,000- and 58,000-dalton polypeptides, and mAb GlyR 7a stained the 48,000- and 93,000-dalton polypeptides. Common antigenic determinants thus are shared by the different subunits of the glycine receptor. Complementary results were obtained by peptide mapping of 125I-labeled glycine receptor polypeptides with various proteases. A set of peptide fragments of the same apparent molecular mass was produced from the different glycine receptor polypeptides by using V8 protease, chymotrypsin, and elastase. These data suggest that the subunits of the glycine receptor have significant homology within their primary structure and may have evolved from a common ancestor receptor polypeptide.

Amino Acid Sequence↗
PubMed1984

Monoclonal antibodies against the alpha-bungarotoxin-binding protein of chick optic lobe.

The nicotinic acetylcholine (ACh) receptor probe alpha-bungarotoxin (alpha-Butx) binds with high affinity to a membrane protein of the vertebrate central nervous system. To characterize further this putative neuronal ACh receptor, we have prepared monoclonal antibodies (mAbs) against the alpha-Butx-binding protein of chick optic lobe. Mice were immunized with affinity-purified protein preparations which were estimated to be 10 to 20% pure. Spleen cells from an immunized mouse were fused with the mouse myeloma cell line X63-Ag 8.653. From this fusion, 14 stable hybridoma lines were isolated which produce mAbs against the chick neuronal alpha-Butx-binding protein. Most of the antibodies inhibited alpha-Butx-binding to membrane fractions and/or detergent extracts of chick optic lobe. Some of the mAbs cross-reacted with the alpha-Butx-binding protein of the rat pheochromocytoma cell line PC12. However, none of the mAbs bound to a significant extent to the nicotinic ACh receptor of chick skeletal muscle or of Torpedo californica electric organ. All antibodies specifically isolated a polypeptide of Mr = 57,000 (+/- 2,000) from radiolabeled neuronal protein preparations. The present data show that these mAbs constitute useful tools for the further molecular and functional characterization of the neuronal alpha-Butx-binding protein.

Animals↗
PubMed1984

[Cerebral angiography in transitory global amnesia].

33 patients with transient global amnesia were examined by cerebral angiography. The presence of arterial lesions correlated with neurological symptoms, as well as with amnesia. In 55% of cases no pathological alterations were found, so that the pathogenesis of the transient global amnesia remained unclear. For clinical purposes, cerebral angiography is recommended only in those cases which show other neurological symptoms besides the transient global amnesia.

Adult↗
PubMed1983

Photoaffinity-labelling of the glycine receptor of rat spinal cord.

The irreversible incorporation upon ultraviolet illumination of the glycine receptor antagonist, [3H]strychnine, into synaptic membrane fractions of rat spinal cord has been investigated. The specificity of this photoaffinity-labelling reaction for the glycine receptor was demonstrated by the following results: (a) the Kd value (9.7 nM) of the glycine-displaceable irreversible incorporation of [3H]strychnine was similar to the previously reported Kd of [3H]strychnine binding to the glycine receptor; (b) pre-illumination of the membranes with unlabelled strychnine led to a corresponding reduction in the number, but not the affinity, of reversible glycine-displaceable [3H]strychnine binding sites; (c) the ultraviolet light-induced incorporation into the membranes of [3H]strychnine was inhibited by different glycine receptor agonists; other neurotransmitter substances had little or no effect. Also, [3H]strychnine alone was shown to be stable upon illumination with ultraviolet light; this suggests that photocrosslinking of [3H]strychnine may require energy transfer from specific groups of its high-affinity receptor binding site. Upon sodium dodecyl sulphate/polyacrylamide gel electrophoresis a single labelled polypeptide with a relative molecular mass of 48000 was revealed from spinal cord membranes photoaffinity-labelled with [3H]strychnine. Spinal cord membranes photoaffinity-labelled with the gamma-aminobutyric acid receptor ligand [3H]flunitrazepam, however, gave a single polypeptide with a relative molecular mass of 5- 0000. Treatment of membranes, labelled with [3H]strychnine, by endoglycosidase H did not alter the relative molecular mass of the 48000-Mr labelled polypeptide. Trypsin treatment, on the other hand, successively produced major fragments of relative molecular masses of 42000 and 37000. Also, even after extensive treatment with trypsin or chymotrypsin, greater than or equal to 90% of the radioactivity incorporated into the labelled membranes remained membrane-associated. It is concluded that the strychnine binding site of the glycine receptor is located on a protease-inaccessible, i.e. probably hydrophobic domain of the 48000-Mr subunit.

Affinity Labels↗
PubMed1983

Identification by cross-linking of a beta-bungarotoxin binding polypeptide in chick brain membranes.

beta-Bungarotoxin (beta-BTX) is a snake venom neurotoxin which inhibits neurotransmitter release from different types of nerve terminals. To identify presynaptic membrane components potentially important in neurosecretion, 125I-labeled beta-BTX (mol. wt. 21 000) was cross-linked to a high-affinity binding site in synaptic membrane fractions of chick brain using the photoactivable cross-linker N-succinimidyl-6(4'-azido-2'-nitrophenylamino)-hexanoate. Electrophoretic analysis of the cross-linked membrane proteins under both reducing and non-reducing conditions revealed a single [125I]beta-BTX-polypeptide adduct of apparent mol. wt. 116 000 (+/- 2000). The labeling of this band was prevented under conditions previously shown to inhibit the binding of [125I]beta-BTX to its high-affinity binding site. It is concluded that the cross-linking procedure identified a polypeptide of the presynaptic binding site for beta-BTX, and that this polypeptide has a mol. wt. of 95 000.

Animals↗
PubMed1983

External carotid steal and lateral medullary infarction--is there a pathogenetic connection?

The case of a patient with occlusion of the external carotid artery is reported; he suffered from vertebrobasilar insufficiency with Wallenberg's syndrome. Arteriography showed stealing of intracranial blood from the vertebral artery via the occipital artery, and thromboendarterectomy was successfully performed. Since no localised processes could be demonstrated could the lateral medullary infarction be due to the haemodynamic changes? In cases of external carotid steal thromboendarterectomy might be the appropriate treatment.

Carotid Artery Thrombosis↗
PubMed1983

Regulation of alpha-bungarotoxin receptor accumulation in chick retina cultures: effects of membrane depolarization, cyclic nucleotide derivatives, and Ca2+.

The regulation of a putative neuronal nicotinic acetylcholine receptor, the alpha-bungarotoxin (alpha-Btx)-binding protein, was investigated in primary cultures of chick embryo retina. Depolarization of the cells by veratrum alkaloids, or by a high K+ concentration of the culture medium, reduced the accumulation of cell surface alpha-Btx receptors in these cultures. In contrast, the addition of a blocker of voltage-dependent Na+ channels, tetrodotoxin, increased alpha-Btx receptor accumulation in differentiated retina cultures. Derivatives of cyclic adenosine 3':5'-monophosphate also increased the number of alpha-Btx receptor sites, whereas dibutyryl cyclic guanosine 3':5'-monophosphate, but not 8-bromo-cyclic guanosine 3':5'-monophosphate, had the opposite effect. The blocker of voltage-dependent Ca2+ channels, D600, and media containing a reduced Ca2+ concentration also increased alpha-Btx receptor levels. All of these different culture conditions altered the rate of receptor loss after blocking protein synthesis by cycloheximide. These results show that the synthesis of the neuronal alpha-Btx receptor is regulated by membrane depolarization, cyclic nucleotides, and Ca2+ in a fashion analogous to the regulation of muscle acetylcholine receptor.

Animals↗
PubMed1982

Interaction of histrionicotoxin with the putative nicotinic acetylcholine receptor of the chick visual system.

The frog alkaloid histrionicotoxin inhibits neuromuscular transmission in a non-competitive fashion and is thought to bind to the ion channel region of the nicotinic acetylcholine receptor (nAChR) of skeletal muscle and fish electric organ. Here, the use of histrionicotoxin as a probe for the putative nAChR of the chick visual system was investigated. Histrionicotoxin inhibited the binding of [125I]alpha-bungarotoxin to synaptic membranes of optic lobe or of retina in a competitive manner (Ki = 6 +/- 3 microM). However, the number of displaceable membrane binding sites for the physiologically active derivative [3H]perhydrohistrionicotoxin was 20-100-fold higher than of [125I]alpha-bungarotoxin binding sites. Also, the binding of [3H]perhydrohistrionicotoxin to the membranes could be displaced by local anaesthetics and phencyclidine, but was insensitive to nicotinic-cholinergic ligands or to alpha-bungarotoxin. It is concluded that histrionicotoxins are not specific ligands for nAChR in the chick CNS.

Amphibian Venoms↗
PubMed1982

Beta-bungarotoxin-induced cell-death of neurons in chick retina.

The cytotoxicity of beta-bungarotoxin (beta-BTX), a snake venom neurotoxin with phospholipase A2 activity, for chick neurons was investigated using organ and monolayer cultures of retina. Beta-BTX led to a marked reduction in the total activities of choline acetyltransferase and glutamate decarboxylase of retina cultures at concentrations as low as 100 pM. The total activity of lactate dehydrogenase was, however, much less affected by beta-BTX. Also, the total activity of tyrosine hydroxylase of organ-cultured retina decreased only at 30-50 fold higher concentrations of the toxin. The total activity of the glial marker glutamine synthetase was not changed by beta-BTX. In contrast to this selectivity for neurons displayed by beta-BTX, non-neurotoxic phospholipases A2 from bee venom and porcine pancreas led to a simultaneous loss of both neuronal and glial marker enzymes. Light and electron microscopy of organ-cultured retina showed that only cells in the ganglion cell layer and the inner third of the amacrine cell layer degenerated after incubation with beta-BTX. In the toxin-sensitive cells, the Golgi apparatus and the endoplasmatic reticulum appeared the first subcellular structures to be affected. It is concluded that beta-BTX preferentially recognizes and/or destroys cholinergic and GABAergic cells in the amacrine and ganglion cell layers of the developing chick retina. This toxin may thus be a useful probe to investigate cell surface properties of cholinergic and GABAergic neurons in the chick central nervous system.

Animals↗
PubMed1982

Identification of polypeptides associated with a putative neuronal nicotinic acetylcholine receptor.

Polypeptides involved in the binding of the nicotinic acetylcholine receptor ligand alpha-bungarotoxin (Mr = 8,000) to neuronal membranes were identified by three independent methods: (i) 125I-alpha-bungarotoxin bound to membrane fractions or to monolayer cultures of chick retina was cross-linked to its binding site by using glutaraldehyde, or the photoactivatable bifunctional reagent N-succinimidyl-6-(4'-azido-2'-nitrophenylamino)hexanoate. Electrophoretic analysis of the cross-linked membrane proteins revealed 125I-alpha-bungarotoxin-polypeptide adducts of apparent Mr = 63,000, 43,000, and 33,000. (ii) Affinity purification of the alpha-bungarotoxin binding protein from detergent extracts of [35S]methionine-labeled retina cultures identified one major polypeptide with an Mr = 57,000. (iii) Indirect immunoprecipitation from detergent extracts of [35S]methionine-labeled rat pheochromocytoma cells (PC 12) gave evidence for a specific co-precipitation of alpha-bungarotoxin with three polypeptides (Mr = 57,000, 34,000, and 25,000). The data suggest that polypeptides of Mr - 57,000, 35,000, and 25,000 (+/- 3,000) are located at or close to the alpha-bungarotoxin binding domain of the putative neuronal nicotinic acetylcholine receptor.

Animals↗
PubMed1982

Binding of beta-bungarotoxin to synaptic membrane fractions of chick brain.

beta-Bungarotoxin (beta-BuTx) is a presynaptically active snake venom phospholipase A2 which, in the chick, is cytotoxic for certain subclasses of central and peripheral neurons. In order to elucidate whether the toxin's specificity is due to the existence of specific neuronal binding sites, the binding of 125I-labeled beta-BuTx to synaptic membrane fractions of chick brain was investigated. These experiments defined a single class of saturable high affinity binding sites (KD = 0.47 +/- 0.14 nM, k+1 = 4.3 x 10(6) M-1 s-1, k-1 = 1.08 x 10(-4) s-1) whose pharmacological properties correlated with that of the cytotoxic action of beta-BuTx on cholinergic neurons in chick retina cultures. The density of 125I-beta-BuTx binding sites in synaptic membrane fractions was low (50 fmol/mg of protein). Also specific toxin binding occurred only to membrane fractions of chick organs known to contain beta-BuTx-sensitive cells or nerve endings, i.e. brain, retina, and muscle, but not liver and heart. The binding of 125I-beta-BuTx to synaptic membrane fractions was dependent on Ca2+; Co2+ and Sr2+, but not Mg2+, could replace Ca2+ in the binding reaction. The membrane binding site for 125I-beta-BuTx was sensitive to heat and high concentrations of pronase, and thus most likely is a protein.

Animals↗
PubMed1982

Purification by affinity chromatography of the glycine receptor of rat spinal cord.

The glycine receptor of rat spinal cord was solubilized with the nonionic detergent Triton X-100 and subsequently purified by affinity chromatography on aminostrychnine-agarose and wheat germ agglutinin-Sepharose. An overall purification of 1950-fold was achieved. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and mercaptoethanol revealed three glycine receptor-associated polypeptides of Mr = 48,000, 58,000, and 93,000. [3H]Strychnine was incorporated irreversibly into the Mr = 48,000 polypeptide upon UV-illumination. The dissociation constant (KD) of [3H]strychnine binding to the purified glycine receptor was 9.3 +/- 0.6 nM. The glycine receptor agonists glycine, beta-alanine, and taurine inhibited the binding of [3H]strychnine to the purified receptor. Gel filtration and sedimentation in sucrose/H2O and sucrose/D2O gradients gave a Stokes radius of 7.7 nm, a partial specific volume of 0.780 +/- 0.005 ml/g and a sedimentation coefficient s20,w of 8.2 +/- 0.2 S for the purified glycine receptor. From these data, a molecular weight of 246,000 +/- 6,000 was calculated for the glycine receptor protein.

Affinity Labels↗
PubMed1982

Avermectin B1a inhibits the binding of strychnine to the glycine receptor of rat spinal cord.

The anthelmintic drug, avermectin B1a, has been reported to interfere with gamma-aminobutyric acid-mediated chloride conductance. Also, enhancement of diazepam binding to mammalian brain membranes by avermectin B1a has led to the suggestion that avermectin B1a interacts with the 'benzodiazepine receptor-gamma-aminobutyric acid receptor-chloride ionophore' complex. Here we report an interaction of avermectin B1a with the glycine receptor. The binding of the glycine receptor antagonist, strychnine, to both membranes and solubilized receptor from rat spinal cord, was inhibited by avermectin B1a with Ki values of 1.3 microM and 3.6 microM, and Hill coefficients of 0.46 and 0.62, respectively.

Animals↗
PubMed1982

Differential down-regulation by carbamylcholine of putative nicotinic and muscarinic acetylcholine receptor sites in chick retina cultures.

The effect of chronic exposure to the cholinergic against carbamycholine on the number of nicotinic and muscarinic cholinergic ligand binding sites in primary cultures of chick embryo retina has been investigated. Within 24 h, 1 mM carbamylcholine led to a 60-70% loss of specific binding sites for the muscarinic antagonist [3H] quinuclidinylbenzilate, whereas the binding of the nicotinic cholinergic ligand [125I] alpha-bungarotoxin remained almost unchanged under the same conditions. The reduction in muscarinic cholinergic ligand binding sites by carbamylcholine was blocked by atropine and scopolamine, but not affected by the inhibition of protein synthesis or the removal of extracellular calcium. These data indicate a different metabolic regulation by agonist occupancy of putative nicotinic and muscarinic acetylcholine receptors in the chick central nervous system.

Animals↗
PubMed1981

Solubilization of the glycine receptor from rat spinal cord.

Glycine receptors, as detected by glycine-displaceable [3H]strychnine binding, were solubilized from a membrane fraction of rat spinal cord by the non-ionic detergent Triton X-100. The solubilized material retained its high affinity for [3H]strychnine and exhibited the typical pharmacological properties of the membrane-bound glycine receptor. On sucrose density gradients, the solubilized receptor had a sedimentation coefficient of 8.3 +/- 0.4 S. Gel exclusion chromatography on Sepharose 6 B in the presence of phosphatidylcholine gave a Stokes radius of 7.3 +/- 0.3 nm.

Animals↗