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H Betz

Publications and source records attributed to H Betz.

At least 199 records · Page 11Linked to original sources
PubMed1989

Developmental accumulation and heterogeneity of myelin basic protein transcripts in the chick visual system.

By selective cloning of gene products newly expressed in the developing chick optic tectum, we have isolated cDNA clones encoding myelin basic protein (MBP). Three different transcripts of the corresponding single copy gene of 2.7 kb, 2.0 kb, and 1.1 kb were observed on Northern blots. In the developing optic lobe, the most abundant RNA (2.7 kb) was detected first at the 14th day in ovo. Nuclease S1 protection experiments revealed further heterogeneity of MBP transcripts, which result from alternative splicing of segments corresponding to the fifth and sixth exon of the homologous murine MBP gene. From these data, at least three different forms of MBP are predicted to exist in the chicken central nervous system.

Animals↗
PubMed1989

Functional expression in Xenopus oocytes of the strychnine binding 48 kd subunit of the glycine receptor.

The inhibitory postsynaptic glycine receptor (GlyR) of rat spinal cord is an oligomeric transmembrane protein which forms an agonist-gated anion channel. Expression in Xenopus oocytes of its mol. wt 48,000 subunit generated glycine-gated chloride channels which were analysed by voltage clamp. The agonist and antagonist response properties as well as the desensitization characteristics of these 48 kd subunit receptors resembled GlyRs expressed from spinal cord poly(A)+ RNA. These data indicate that the 48 kd subunit is capable of assembling into a functional receptor homo-oligomer which displays the pharmacology characteristic of the spinal cord GlyR.

Animals↗
PubMed1989

Neuraxin, a novel putative structural protein of the rat central nervous system that is immunologically related to microtubule-associated protein 5.

During screening of a rat spinal cord lambda gt11 cDNA library with poly- and monoclonal antibodies against the postsynaptic glycine receptor a cDNA was isolated which covers an open reading frame encoding a protein of calculated mol. wt 94 kd. Sequence analysis identified a novel type of neuron-specific protein (named neuraxin) which is characterized by an unusual amino acid composition, 12 central heptadecarepeats and putative protein and/or membrane interaction sites. The gene encoding neuraxin appears to be unique in the haploid rat genome and conserved in higher vertebrates. Northern blot and in situ hybridization revealed neuraxin mRNA to be expressed throughout the rodent central nervous system (CNS). In spinal cord, neuraxin transcripts were abundant in motoneurons which also expressed glycine receptor subunit mRNA. A bacterial fusion protein containing approximately 90% of the neuraxin sequence was found to specifically bind tubulin. Polyclonal neuraxin antibodies cross-reacted with microtubule-associated protein 5 (MAP5), and a monoclonal antibody against MAP5 recognized the neuraxin fusion construct. Based on these data we suggest that neuraxin is related to MAP5 and may be implicated in neuronal membrane-microtubule interactions.

Amino Acid Sequence↗
PubMed1989

Central nicotinic acetylcholine receptors in the chicken and Drosophila CNS: biochemical and molecular biology approaches.

Putative neuronal nicotinic acetylcholine receptors (nAChRs) were investigated using biochemical and molecular biology approaches. In the chick visual system, a nicotinic cholinergic binding site was localized on a polypeptide of Mr 57,000 using the potent antagonist, alpha-bungarotoxin (alpha-Btx). Ion flux experiments indicate that this membrane protein is different from the toxin-insensitive nAChR present in vertebrate neurons. Crosshybridization with a Torpedo nAChR cDNA probe allowed isolation of a cholinergic receptor cDNA (ARD) from the Drosophila CNS. Analysis of the corresponding gene, its transcripts and the ARD protein are presented here.

Animals↗
PubMed1989

Expression of synaptophysin in the rat pheochromocytoma cell line PC12.

Expression of the integral membrane protein of small synaptic vesicles, synaptophysin, was investigated in the pheochromocytoma cell line PC12 using a quantiative dot immunoassay. Specific synaptophysin contents of the cultures varied with cell density, high levels being observed in densely seeded dishes and/or after some days of subculturing. Northern blot analysis revealed these cell density-related changes in synaptophysin protein contents to result partly from corresponding alterations in mRNA levels. Treatment with nerve growth factor (NGF), but not with various other effectors of intracellular messenger systems, inhibited both synaptophysin and DNA accumulation in the cultures. These data indicate that synaptophysin expression is high in densely proliferating PC12 cells and uncoupled from process formation and neuronotypic differentiation induced by NGF.

Animals↗
PubMed1989

Neuronal acetylcholine receptors in Drosophila: mature and immature transcripts of the ard gene in the developing central nervous system.

The ARD protein is a Drosophila homolog of vertebrate nicotinic acetylcholine receptor (AChR) polypeptides. Here, an analysis of transcripts of the corresponding ard gene is presented. In situ hybridization experiments revealed ard gene expression in nervous tissue only. During development, ard transcripts are prevalent in late embryos, pupae, and newly eclosed flies. Both the spatial and the temporal pattern of ard gene expression is consistent with the ARD protein being part of a neuronal AChR that is produced in large amounts during major periods of neuronal differentiation. In situ hybridization with an intron-specific probe indicated codistribution of immature and mature ard RNAs in pupae and adult flies. In addition to the mature 3.2 kb RNA species, two large immature transcripts are found in newly eclosed flies but not in embryos, suggesting a developmentally regulated processing of ard RNA.

Animals↗
PubMed1989

Functional chloride channels by mammalian cell expression of rat glycine receptor subunit.

Cultured human cells were transfected with cloned rat glycine receptor (GlyR) 48 kd subunit cDNA. In these cells glycine elicited large chloride currents (up to 1.5 nA), which were blocked by nanomolar concentrations of strychnine. However, no corresponding high-affinity binding of [3H]strychnine was detected in membrane preparations of the transfected cells. Analysis by monoclonal antibodies specific for the 48 kd subunit revealed high expression levels of this membrane protein. After solubilization, the 48 kd subunit behaved as a macromolecular complex when analyzed by sucrose density centrifugation. Approximately 50% of the solubilized complex bound specifically to a 2-aminostrychnine affinity column, indicating the existence of low-affinity antagonist binding sites on most of the expressed GlyR protein. Thus, the 48 kd strychnine binding subunit efficiently assembles into high molecular weight complexes, resembling the native spinal cord GlyR. However, formation of functional receptor channels of high affinity for strychnine occurs with low efficiency.

Amino Acid Sequence↗
PubMed1989

Primary cultures of mouse spinal cord express the neonatal isoform of the inhibitory glycine receptor.

Expression of the inhibitory glycine receptor complex was investigated in primary cultures of fetal mouse spinal cord using sensitive immunomethods. In these cells, glycine receptor is predominantly of the neonatal isoform characterized by a low affinity for the antagonist strychnine. It contains a ligand binding subunit that differs from that of the adult receptor in antigenic epitopes and apparent molecular weight. Whereas in vivo the neonatal receptor isoform is completely replaced by the adult isoform within 3 weeks after birth, this exchange of subtypes is not seen in culture. The increased expression of the cytoplasmic glycine receptor-associated polypeptide of 93 kd occurring after birth is also seen under culture conditions. Purification of glycine receptor from cultures yielded polypeptides of 49 kd and 93 kd, suggesting that the membrane-spanning core of the neonatal receptor may be a homooligomer composed of 49 kd subunits. About half of the 49 kd subunit is cleaved by trypsinization of the cultures, indicating a predominant cell surface localization of the receptor. Pulse-labeling experiments revealed the 49 kd subunit to be a metabolically stable glycoprotein (half-life approximately 2 days). After its synthesis, a transition time of 30-45 min is required for acquisition of a strychnine binding conformation.

Animals↗
PubMed1989

Sensitive immunoassay shows selective association of peripheral and integral membrane proteins of the inhibitory glycine receptor complex.

The inhibitory glycine receptor of mammalian spinal cord is a ligand-gated chloride channel that, on affinity purification, contains two subunits of 48-kilodalton (kD) and 58-kD molecular mass in addition to an associated 93-kD protein. Ligand-binding 48-kD subunit and 93-kD protein were quantified in the CNS of the adult rat using a newly developed dot receptor assay (detection limit less than or equal to 1 fmol/assay) which employs monoclonal antibodies specific for glycine receptor polypeptides. The 93-kD protein was found to codistribute at a fixed stoichiometry with the 48-kD subunit throughout the CNS of the rat. Moreover, the 93-kD protein cofractionated with the ligand-binding subunit on solubilization and affinity chromatography or immunoprecipitation. However, both proteins were separated on sucrose gradient centrifugation of detergent extracts of spinal cord membranes in accord with earlier observations on purified receptor. These data prove that the 93-kD polypeptide is selectively associated with the membrane core of the strychnine-sensitive glycine receptor. The regional distribution of glycine receptor polypeptides was also determined in the CNS of the spastic rat mutant. In contrast to hereditary spasticity in mouse and cattle, no reduction of glycine receptors was found in the spastic rat.

Animals↗
PubMed1989

Localization of glycine and beta-adrenergic receptors in the rat brain.

The localization of glycine (GR) and adrenergic receptors (AdR) was examined in the rat brain using a monoclonal antibody against the affinity purified glycine receptor and a polyclonal antibody against purified beta2-AdR. GR were concentrated in the lower brainstem, whereas no immunoreactivity was observed in the diencephalon and forebrain except in a few diencephalic nuclei. The highest density was found in the cranial motor nuclei, reticular formation, parabrachial area, dorsal and ventral cochlear nuclei, and dorsal and ventral tegmental nuclei. On the other hand, AdR were widely but unevenly distributed in the rat brain. The present study further showed the presence of AdR in catecholaminergic terminals in the hypothalamus.

Animals↗
PubMed1989

[Molecular biology of the inhibitory synapse].

The amino acids glycine and gamma-aminobutyric acid are the major inhibitory neurotransmitter substances in the central nervous system of all vertebrates. Due to an increase of chloride ion conductance of the postsynaptic membrane these neurotransmitters inhibit the excitability of the nerve cell. During the last years the receptor proteins mediating the action of glycine and gamma-aminobutyric acid were characterized in more detail by biochemical and molecular biology techniques. The present data show that all neurotransmitter receptors which alter the ion permeability of the postsynaptic membrane are members of a large protein family which has evolved from a common precursor.

Animals↗
PubMed1988

Glycine receptor heterogeneity in rat spinal cord during postnatal development.

Two different isoforms of the inhibitory glycine receptor were identified during postnatal development of rat spinal cord. A neonatal form characterized by low strychnine binding affinity, altered antigenicity, and a ligand binding subunit differing in mol. wt (49 kd) from that of the adult receptor (48 kd) predominates at birth (70% of the total receptor protein). Separation from the adult form could be achieved by either use of a selective antibody or glycine gradient elution of 2-aminostrychnine affinity columns. Both isoforms co-purify with the mol. wt 93 kd peripheral membrane protein of the postsynaptic glycine receptor complex.

Animals↗
PubMed1988

The beta-bungarotoxin-binding protein from chick brain: binding sites for different neuronal K+ channel ligands co-fractionate upon partial purification.

beta-Bungarotoxin (beta-Butx) is a presynaptically active neurotoxin which blocks neuronal A-type K+ channels. Here, the efficient solubilisation and about 300-fold purification of the beta-Butx-binding protein from chick brain were achieved by detergent extraction at high ionic strength followed by chromatography on DEAE Affigel Blue, beta-Butx Affigel 102 and wheat germ agglutinin Sepharose. Binding of 125I-labelled beta-Butx to the purified protein was inhibited by two other K+ channel ligands, dendrotoxin I and mast cell-degranulating peptide. It is concluded that the beta-Butx-binding protein is a member of a family of voltage-gated K+ channels which exhibit varying affinities for different polypeptide ligands.

Animals↗
PubMed1988

Identification of synaptophysin as a hexameric channel protein of the synaptic vesicle membrane.

The quaternary structure and functional properties of synaptophysin, a major integral membrane protein of small presynaptic vesicles, were investigated. Cross-linking and sedimentation studies indicate that synaptophysin is a hexameric homo-oligomer, which in electron micrographs exhibits structural features common to channel-forming proteins. On reconstitution into planar lipid bilayers, purified synaptophysin displays voltage-sensitive channel activity with an average conductance of about 150 picosiemens. Because specific channels and fusion pores have been implicated in vesicular uptake and release of secretory compounds, synaptophysin may have a role in these processes.

Animals↗