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P Seeburg

Publications and source records attributed to P Seeburg.

13 recordsLinked to original sources
PubMed1993

Molecular distinction of three N-methyl-D-aspartate-receptor subtypes in situ and developmental receptor maturation demonstrated with the photoaffinity ligand 125I-labeled CGP 55802A.

Activation of N-methyl-D-aspartate (NMDA) receptors is essential for synaptic plasticity in the central nervous system and contributes to neuronal death under various pathological conditions. Although several subunits have been cloned, the structure of NMDA receptors in situ is unresolved. By using a photoreactive antagonist with nanomolar affinity to the NMDA-binding site, three types of receptors were differentiated by their pattern of photoaffinity-labeled subunits. In adult brain, a protein of 175-kDa was photoreactive that displayed a profile of ligand binding and autoradiographical distribution corresponding to NMDA receptors. In contrast, in early postnatal brain, proteins of both 175 kDa and 115 kDa were photolabeled. This labeling pattern is switched to that of adult brain around postnatal day 10, pointing to a structural maturation of NMDA receptors. A third type of receptor could be identified in cerebellar granule cell cultures, where NMDA receptors mediate trophic effects and photolabeling was exclusively targeted to a 115-kDa protein. To identify the proteins labeled in situ, recombinant receptors were subjected to photolabeling. When the NR1 subunit was coexpressed with either the NR2A, NR2B, or NR2C subunit, only the combination of NR1/NR2A was photoreactive. Both the NR1 and NR2A subunits were photolabeled, corresponding in size to the proteins labeled in situ. However, the lack of subunit-selectivity in photolabeling the NR1/NR2A combination suggests the presence of additional receptor components in situ to explain the subunit-selective photoreactivity in adult brain (175 kDa) and in cerebellar granule cells (115 kDa). The subunit combination NR1/NR2A by itself appears insufficient to describe a major population of NMDA receptors, in particular, in adult brain.

Affinity Labels↗
PubMed1993

Comparison of in vitro binding properties of a series of dopamine antagonists and agonists for cloned human dopamine D2S and D2L receptors and for D2 receptors in rat striatal and mesolimbic tissues, using [125I] 2'-iodospiperone.

We investigated the ligand binding properties in vitro of two splice variants of the cloned human dopamine D2 receptor (the 443 and 414 amino acids long forms called D2L and D2S, respectively), expressed in 293 human kidney cells, in comparison with those of the dopamine D2 receptors in rat striatum, nucleus accumbens and tuberculum olfactorium. The new radioligand, [125I]2'-iodospiperone, showed a similar high binding affinity (KD:0.056-0.122 nM) for cloned human D2S and D2L receptors and for the D2 receptors in the three rat brain areas. Binding affinities of 25 dopamine antagonists and of 10 dopamine agonists belonging to different chemical classes were measured. The IC50 values of the antagonists were virtually identical in the five preparations: spiperone was the most potent compound (pIC50 approximately 9.9), remoxipride the least potent one (pIC50 approximately 5.7). The agonists showed similar IC50 values for the cloned human D2S and D2L receptors but their affinity for rat brain D2 receptors was 2- to 5-fold higher. Dopamine showed shallow inhibition curves, the high affinity binding was 10-fold lower for the cloned human D2 receptors than for the rat brain D2 receptors. Addition of stable guanosine-5'-triphosphate (GTP) analogues shifted the D2 receptors in the rat brain tissues to the "low" affinity state, the low affinity binding of dopamine was equal to the affinity for the cloned human receptor. None of the dopamine antagonists or agonists could differentiate between the two splice forms of the cloned human D2 receptors or between the D2 receptors in rat striatal and mesolimbic tissues. The lower apparent affinity of some agonists and of dopamine in the absence of stable GTP analogues suggests a less appropriate receptor G-protein coupling for the cloned human D2 receptors expressed in the 293 human kidney cells. Unexpectedly, guanosine-5'-O-(3-thiotriphosphate) (GTP-gamma-S) reduced the [125I]2'-iodospiperone binding to the D2 receptors by 20-35% in the rat brain tissues and the cloned human D2L receptor, and by 75% to the cloned human D2S receptor. The inhibition in the last case could be prevented partly by submicromolar concentrations of dopamine. The GTP-gamma-S effect is suggested to be due to reduction of disulphide bonds in the receptor. Recent molecular modelling studies indicated an important role of the disulphide bridge between Cys107 at the start of transmembrane domain three and Cys182 in the third extracellular loop, for the binding of dopamine to the D2 receptor.

Animals↗
PubMed1992

Glutamate receptor expression in the rat retina.

The expression of five genes (GluR A; B; C; D; GluR 5) encoding functional subunits of glutamate receptors was investigated in the rat retina using in situ hybridization with oligonucleotide probes. All five genes are expressed in the retina. All probes label cell bodies in the ganglion cell layer as well as somata in the inner third of the inner nuclear layer (INL), where the amacrine cells are located. In addition GluR 5, B and D, and to a lesser extent also GluR A are found in the middle and outer part of the INL, where bipolar and horizontal cells reside. Different subsets of retinal neurons may thus use glutamate receptors of different subunit composition.

Animals↗
PubMed1990

Molecular cloning of a novel G protein-coupled receptor that may belong to the neuropeptide receptor family.

A novel cDNA encoding a putative G protein-coupled receptor was isolated from a rat forebrain cDNA library by homology screening. Sequence comparison demonstrates that the encoded polypeptide containing seven transmembrane domains shows highest homology with tachykinin receptors, particularly in the transmembrane domains and the first intracellular loop. The mRNA for this new receptor seems specifically expressed in brain and is prominently localized in specific nuclei of the rat brain, including thalamus, cerebral cortex and hippocampus. We speculate that a neuropeptide may be the natural ligand of this novel G protein-coupled receptor.

Amino Acid Sequence↗
PubMed1990

Differential expression of inhibin alpha and beta A subunit genes in rat and mouse ovarian follicles during pregnancy.

Relative levels of rat ovarian alpha inhibin (alpha I) and beta A inhibin (beta AI) mRNAs were measured during pregnancy by dot-blot hybridization of ovarian poly(A+) RNA. Follicular patterns of alpha I and beta AI expression in contralateral ovaries from the same rats were also studied by hybridization histochemistry. Oligodeoxynucleotide probes specific for porcine alpha I and beta AI were synthesized, 32P end-labelled and used as hybridization probes on dot-blots of ovarian RNA and frozen sections of ovarian tissue from pregnant rats. During pregnancy, levels of alpha I and beta AI mRNAs remained fairly constant from day 7 after mating until parturition and then fell within 16 h post partum. In all ovaries observed, expression of inhibin genes was located in granulosa cells of healthy antral follicles. In general, the strongest signals for alpha I and beta AI mRNAs were obtained in large follicles, with weaker signals in smaller follicles. Follicular patterns of alpha I and beta AI expression during pregnancy were often dissimilar when alpha I and beta AI were compared over a range of follicles. Considerable alpha I mRNA was detectable in some follicles in which beta AI was reduced or undetectable, despite strong signals for both alpha I and beta AI in an adjacent follicle. Essentially, alpha I mRNA levels were relatively consistent between groups of follicles, whereas beta AI levels varied considerably. beta AI mRNA was never observed in a follicle in the absence of alpha I mRNA, indicating that activin production in any follicle occurs in the presence of alpha I mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Potassium channels expressed from rat brain cDNA have delayed rectifier properties.

Injection into Xenopus oocytes of RNA synthesized in vitro using the rat brain cDNA RCK1 as a template or nuclear injection of the cDNA results in the expression of functional potassium channels. These channels exhibit properties similar to those of the non-inactivating delayed rectifier channel found in mammalian neurons and other excitable cells.

4-Aminopyridine↗
PubMed1984

Complete nucleotide sequence of the glutamate dehydrogenase gene from Escherichia coli K-12.

A 2.3-kb PstI- ClaI chromosomal DNA segment, carrying the complete coding region of the glutamate dehydrogenase (GDH) structural gene from Escherichia coli K-12, has been sequenced. The complete amino acid sequence (447 residues) of the GDH monomer has been deduced, and comparisons are made with reported amino acid sequences of GDH from other organisms.

Amino Acid Sequence↗
PubMed1983

Nucleotide sequence of the promoter and amino-terminal coding region of the glutamate dehydrogenase structural gene of Escherichia coli.

A 610-bp DNA fragment carrying the promoter and amino-terminal coding regions of the glutamate dehydrogenase (GDH) structural gene from Escherichia coli has been sequenced. The amino-terminal sequence of the enzyme was also determined to help localize the transcriptional and translational signals for this gene. Three possible promoters and a CRP binding site were identified by concensus criteria. The sequence of 102 amino acids at the amino terminus of the enzyme is compared with the amino acid sequence from other GDH enzymes.

Animals↗
PubMed1983

Primary structure of the human proenkephalin gene.

The nucleotide sequence of a 6.8-kb region of human DNA containing the proenkephalin gene and flanking regions is reported. The gene consists of four exons separated by three introns and spans approximately 5.3 kb of DNA. Location and identification of several repetitive DNA sequences within and flanking the gene are also described. The distribution of CpG dinucleotides as well as the extent of CpG methylation at several restriction sites within and surrounding the gene is also presented. The structural organization of the human proenkephalin gene exhibits striking similarities to the organization of the human pro-opiomelanocortin (POMC) gene. Nucleotide sequence homologies suggest that two opioid peptide precursors, proenkephalin and prodynorphin, may have arisen by duplication from a common ancestral gene.

5-Methylcytosine↗
PubMed1982

Molecular cloning establishes proenkephalin as precursor of enkephalin-containing peptides.

Molecular cloning and DNA sequencing have yielded considerable structural information about proenkephalin. All previously characterized intermediate peptides of the enkephalin pathways in bovine adrenal medulla have now been aligned into an unambiguous primary structure. Two basic amino acid residues serve as processing signals for release of each of the different components.

Adrenal Medulla↗
PubMed1981

Expression of two human growth hormone genes in monkey cells infected by simian virus 40 recombinants.

We have constructed simian virus 40 recombinants carrying two different human growth hormone (hGH) genes. Monkey kidney cells infected with these recombinants synthesize, process, and secrete hGH. The product of gene 1, which has coding sequences identical to those of a cloned hGH complementary DNA, is indistinguishable from pituitary hGH by several criteria. The product of gene 2, which is predicted to encode a variant protein, is less immunoreactive than pituitary hGH but binds efficiently to hGH cell surface receptors. These results show that gene 2 has the potential to be expressed into a previously unidentified form of hGH. They also demonstrate that it is possible to produce a mature hormone by gene transfer in eukaryotic cells and indicate the utility of the simian virus 40-monkey cell system for producing and characterizing secreted animal cell proteins.

Animals↗
PubMedDate not supplied

Localization of GABAA receptors in the rat retina.

Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mammalian retina. The present paper describes the localization of GABAA receptors in the rat retina as revealed by in situ hybridization and immunocytochemistry. In situ hybridization with probes against various alpha subunits revealed a marked differential expression pattern. The alpha 1 subunit gene is expressed mainly in the bipolar and horizontal cell layer, the alpha 2 gene in the amacrine and ganglion cell layer, and the alpha 4 gene in a subpopulation of amacrine cells. beta subunit mRNA is present diffusely throughout the entire inner nuclear layer and in the ganglion cell layer. The monoclonal antibody bd 17 (against beta 2/beta 3 subunits) stained subpopulations of GABAergic and glycinergic amacrine cells as well as some ganglion cells and bipolar cells. Immunoreactivity was not restricted to synaptic input sites. In the outer plexiform layer bipolar cell dendrites were immunoreactive; in the inner plexiform layer mainly amacrine and ganglion cell processes were labeled, and bipolar cell axons appeared unstained. The results demonstrate a strong heterogeneity of GABAA receptors in the retina.

Animals↗