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D Goldman

Publications and source records attributed to D Goldman.

At least 289 records · Page 16Linked to original sources
PubMed1987

Functional expression of two neuronal nicotinic acetylcholine receptors from cDNA clones identifies a gene family.

A family of genes coding for proteins homologous to the alpha subunit of the muscle nicotinic acetylcholine receptor has been identified in the rat genome. These genes are transcribed in the central and peripheral nervous systems in areas known to contain functional nicotinic receptors. In this paper, we demonstrate that three of these genes, which we call alpha 3, alpha 4, and beta 2, encode proteins that form functional nicotinic acetylcholine receptors when expressed in Xenopus oocytes. Oocytes expressing either alpha 3 or alpha 4 protein in combination with the beta 2 protein produced a strong response to acetylcholine. Oocytes expressing only the alpha 4 protein gave a weak response to acetylcholine. These receptors are activated by acetylcholine and nicotine and are blocked by Bungarus toxin 3.1. They are not blocked by alpha-bungarotoxin, which blocks the muscle nicotinic acetylcholine receptor. Thus, the receptors formed by the alpha 3, alpha 4, and beta 2 subunits are pharmacologically similar to the ganglionic-type neuronal nicotinic acetylcholine receptor. These results indicate that the alpha 3, alpha 4, and beta 2 genes encode functional nicotinic acetylcholine receptor subunits that are expressed in the brain and peripheral nervous system.

Amino Acid Sequence↗
PubMed1987

Genetic variants of C2 muscle cells that are defective in synthesis of the alpha-subunit of the acetylcholine receptor.

We have analyzed two genetic variants of C2 muscle cells that have reduced levels of binding activity for alpha-bungarotoxin and have found that both synthesize only low levels of the alpha-subunit of the acetylcholine receptor. In both variants the uptake of 22Na in response to carbachol is diminished in proportion to the reduction in toxin-binding activity. In addition, the kinetic and sedimentation properties of the residual toxin-binding activity in both is indistinguishable from that seen in wild-type cells. Immunoblotting experiments on extracts of the variants using subunit-specific antibodies to alpha- and beta-subunits of the acetylcholine receptor demonstrated that the beta-subunit was present, but failed to detect alpha-subunit. In both variants, the amount of alpha-subunit accumulated after a 5-min period of labeling with [35S]methionine was reduced by over 90%, leading to the conclusion that the alpha-subunit is synthesized at greatly reduced rates. Northern blot and S1 nuclease analysis showed no differences between the alpha-subunit mRNA in wild-type and variant cells.

Animals↗
PubMed1987

Sex-differences in ethanol sensitivity and alcohol and aldehyde dehydrogenase activities in the Syrian hamster.

Because male Syrian hamsters demonstrate greater preference for ethanol than female hamsters, we compared them with regard to ethanol sensitivity and hepatic alcohol and aldehyde dehydrogenase activities. Male hamsters were slower to recover righting response and had lower blood alcohol levels upon recovery than did females. Hepatic alcohol dehydrogenase activity was approximately twice as high in females as males, but gender differences were not found for either cytosolic or non-cytosolic aldehyde dehydrogenase activities. The results suggest that the reduced ethanol sensitivity of female hamsters is due to more rapid metabolism. However, the finding that female hamsters have higher blood alcohol concentrations upon recovery also suggests the possibility of reduced CNS sensitivity.

Alcohol Dehydrogenase↗
PubMed1986

Isolation and sequence of cDNA clones coding for the precursor to the gamma subunit of mouse muscle nicotinic acetylcholine receptor.

cDNA libraries have been constructed in plasmid (pBR322) and bacteriophage lambda gammagt10) vectors with poly (A+) RNA isolated from the nonfusing mouse muscle cell line BC3H-1. The libraries were screened with a restriction fragment derived from a genomic clone coding for a human acetylcholine receptor gamma subunit. Several clones were obtained whose cDNA inserts possessed nucleotide and deduced amino acid sequence homology with acetylcholine receptor gamma subunits from Torpedo californica, chick, calf, and human. One isolate, lambda BMG419, has 88 nucleotides of 5'-untranslated sequence, an open reading frame of 1,557 nucleotides coding for the precursor to the mouse acetylcholine receptor gamma subunit, and 144 nucleotides of 3'-untranslated sequence. Alignment of the lambda BMG419-deduced amino acid sequence with homologs from other species predicts a precursor peptide of 519 amino acids and a mature protein of 497 amino acids, with nonglycosylated molecular weights of 58,744 and 56,424 daltons, respectively. Comparison of the deduced amino acid sequence of the mouse gamma subunit with Torpedo, chick, calf, and human sequences showed overall homologies of 54%, 67%, 90%, and 90%, respectively; however, significantly higher homologies were found in several putative functional domains. Radiolabeled lambda BMG419 has been used to identify homologous RNA species, one of approximately 2 kb and one of about 3.5 kb, in poly (A+) RNA prepared from BC3H-1 cells and denervated mouse limb muscle. gamma Subunit-coding RNA species are considerably more abundant in denervated than in innervated muscle, suggesting that neural regulation of the abundance of the gamma subunit is exerted through regulation of the amount of its mRNA.

Animals↗
PubMed1986

Fourteen genetically variant proteins of mouse brain: discovery of two new variants and chromosomal mapping of four loci.

With the description here of variant proteins A13 (pI 5.9, MW 62 kd) and A14 (pI 5.3, MW 26 kd), 14 polypeptides of mouse brain visualized by two-dimensional electrophoresis (2DE) exhibit genetic variation in isoelectric point. Using 22 B X D recombinant inbred strains, we map four of these loci and show that a fifth is independent of known loci. A pI 5.6, 81-kd protein of mouse brain mitochondria designated A1 is demonstrated to be an independent locus closely linked to LY-2 and LVP-1 on mouse chromosome 6. A pI 5.6, 28-kd genetically variant brain polypeptide designated A12 maps to chromosome 1 and shows identity with the known mouse locus LTW-4. The locus for A8 is not closely linked to any previously mapped locus. However, the locus for the newly described variant A13 shows 3 of 18 recombinants with the DNA polymorphism RN7S-2 and 2 of 18 recombinants with HC (hemolytic complement) and is thus probably located proximally to HC near the centromere of chromosome 2. Genetic and biochemical evidence is presented for the identification of A14 as ALP-1 (apolipoprotein 1), mapping to chromosome 9. In addition to these 13 genetically variant polypeptides, the positions of 12 other polypeptides which have been identified on 2DE gels of mouse brain are given.

Animals↗
PubMed1986

Use of chromosomally mapped and identified mouse brain proteins for behavioral genetic analysis of alcoholism.

A logical first place to look in order to identify loci determining behavioral differences between inbred and certain outbred strains of mice is among the proteins expressed in brain. Fourteen mouse brain proteins have been demonstrated to be genetically variant, four of these have been chromosomally mapped and an additional twelve have been identified and can be simultaneously screened by two dimensional electrophoresis. Certain genetic differences in behavior relevant to alcohol consumption and the effects of alcohol occur between inbred, recombinant inbred and selectively outbred strains. Two genetic correlations are reported, one between an isoelectric point variant of A7 (a 71 kd, pI 5.4 abundant protein) and resistance to signs of ethanol withdrawal and the other between A12 (a 28 kd, pI 5.6 protein) and ethanol intake. Though tentative, these findings illustrate the power of this approach for behavioral genetic analysis and may allow the biochemical genetic bases of these traits to be understood.

Alcoholism↗
PubMed1986

Mapping of brain areas expressing RNA homologous to two different acetylcholine receptor alpha-subunit cDNAs.

We have used an in situ RNA X RNA hybridization technique to determine, in the central nervous systems of the mouse and rat, the distribution of RNA homologous to cDNA clones encoding the alpha subunit of a putative neural nicotinic acetylcholine receptor and the alpha subunit of the muscle nicotinic acetylcholine receptor. Hybridization of the neural alpha-subunit probe was strongest in the medial habenula but was also detected consistently in the compact part of the substantia nigra and ventral tegmental area, in the neocortex, and in certain parts of the thalamus and hypothalamus. The in situ hybridization technique makes it possible to compile a map of brain regions containing cell bodies expressing RNA coding for a specific receptor type and subsequently to apply the techniques of molecular biology to study these brain receptors.

Animals↗
PubMed1986

Nonpyramidal motor activation produced by stimulation of the cerebellum, direct or transcranial: a cerebellar evoked potential.

There is a need to monitor the functional status of the motor pathways well enough to predict the state of that function during operations and in injured or diseased patients. We previously reported that a motor evoked potential (MEP) can be produced by direct or transcranial stimulation of the motor cortex in both cats and humans. This signal descends through both the dorsolateral and ventral spinal cord and is primarily localized in the pyramidal tracts, producing a peripheral nerve signal and an electromyogram (EMG) response. It is more sensitive to injury than the somatosensory evoked potential (SEP). We report here that one can stimulate the cerebellar cortex, either directly or transcranially, and produce a descending signal in the spinal cord that has different characteristics from the MEP. The cerebellar evoked potential (CEP), located in the dorsolateral and the ventral cord, has an earlier latency and a faster conduction velocity than the MEP. It is predominantly ipsilateral with some contralateral components and also produces EMG responses. In the peripheral nerves, the CEP often produces a pattern of several waves that is different from the one or two predominant contralateral waves of the MEP. The CEP is not diminished by pyramidotomy. It arises from two sites on the cerebellar cortex, medial and lateral. The pathways activated may be the vestibulospinal, rubrospinal, reticulospinal, and fastigiospinal systems. This test seems to offer a monitor of selected motor pathways in the spinal cord largely separate from and complementary to the MEP. The ventral pathways activated probably include those demonstrated to be most essential to basic ambulation after spinal cord injury in primates. Also of importance, one type of evoked potential can facilitate another, which provides additional diagnostic tests. The CEP should be of investigative and clinical value.

Animals↗
PubMed1985

Genetic brain polypeptide variants in inbred mice and in mouse strains with high and low sensitivity to alcohol.

Twelve genetically determined brain polypeptide charge variants were identified by comparing cerebellar vermis of 7 inbred mouse strains and of mice selectively bred from 8 strains closely related to these 7 ancestral strains and one other for acute behavioral sensitivity to the sedative effects of ethanol. The selectively bred ethanol-sensitive (LS, long sleep) and insensitive (SS, short sleep) mice exhibited different allelic variants at 6 of these 12 gene loci expressed in the cerebellum. Variant polypeptide A1 (81 kdalton, pI 5.6) was shown to be associated with the membrane of synaptosomal mitochondria and to exhibit a basic variant in SS mice that is determined by a dominant allele. Other variant polypeptides showed codominant inheritance in F1 crosses. However, the phenotype of no single one of these brain polypeptides consistently correlated with the ethanol behavioral sensitivity of the 7 inbred mouse strains nor of 8 recombinant inbred (B X D, C57BL X DBA) strains. This finding supports the hypothesis that a substantial amount of inbreeding, leading to random fixation of alleles independent of selection for ethanol sensitivity, occurred during the breeding of the SS and LS mice. The present findings of a lack of a strong association between sleep time and a brain polypeptide variant do not preclude the existence of a major gene effect contributing to variation in acute sensitivity to ethanol but are consistent with reports that multiple loci are responsible for the difference in ethanol sensitivity between SS and LS mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Homology between rat liver RNA populations during development, regeneration, and neoplasia.

To investigate the degree of homology which may exist between rat liver RNA populations during development, regeneration, and neoplasia, we hybridized polyadenylated RNAs from (a) normal adult, (b) 24-h regenerating, (c) 20-day fetal livers, and (d) the transplantable Morris hepatoma 5123tc to homologous and heterologous complementary DNAs and to cDNAs enriched for sequences preferentially transcribed in either adult or fetal liver. We also compared the in vitro translation products of these RNAs. Analyses of normal adult, regenerating, and fetal liver RNA populations and their translation products show that the overall pattern of gene expression during liver regeneration differs little from that of normal adult rat liver and that mature hepatocytes do not appear to revert to an "immature" state upon reentering the cell cycle. Comparisons between fetal, normal adult, and tumor RNA populations revealed that RNA populations from fetal liver and the 5123tc tumor lack sequences normally expressed in the mature adult liver. However, the tumor does not "reexpress" sequences which are preferentially expressed in fetal livers.

Animals↗
PubMed1985

Isolation of a clone coding for the alpha-subunit of a mouse acetylcholine receptor.

The mouse cell line BC3H-I synthesizes an acetylcholine receptor (AChR) with the pharmacological properties of a muscle nicotinic cholinergic receptor. We have purified mRNA from this cell line and used the size-fractionated poly(A)+RNA to produce a cDNA library of approximately 50,000 clones. The library was screened with a subclone containing genomic sequences coding for the putative acetylcholine-binding site of the alpha-subunit of chicken AChR. We obtained a plasmid, pMAR alpha 15, with a 1,717-base pair insert. The insert cDNA has 26 nucleotides at the 5'-end which code for a portion of the signal peptide followed by a single open reading frame of 1,311 nucleotides which code for a protein of 49,896 daltons. The insert has 377 bases of 3'-untranslated sequence with 3 polyadenylation sites. Radiolabeled plasmid DNA has been used to identify homologous RNA species of about 2 kilobases in Northern blot analyses of poly(A)+ selected RNA from BC3H-I cells. A similar size mRNA is seen in innervated mouse diaphragm and leg muscle, and both mouse and rat brain. Comparisons of the deduced amino acid sequence of the mouse AChR alpha-subunit with Torpedo marmorata, T. californica, chicken, human, and calf sequences show overall homologies of 80%, 80%, 86%, 96%, and 95%, respectively. More detailed analyses reveal a non-random distribution of amino acid substitutions in several structural domains. Based on the absolute conservation of cysteine residues, a new model for the arrangement of the disulfide bonds in the extracellular portion of the alpha-subunit is proposed.

Amino Acid Sequence↗
PubMed1985

Twenty-seven protein polymorphisms by two-dimensional electrophoresis of serum, erythrocytes, and fibroblasts in two pedigrees.

Twenty-seven independent polymorphic loci were detected by two-dimensional electrophoresis (2DE) of serum, erythrocytes, and fibroblasts in two large families and analyzed for linkage to classical genetic markers. We detected seven serum, four erythrocyte, and 17 fibroblast protein loci that exhibited charge variation in these two families and in a sample of unrelated individuals. The genetic basis of protein variants was confirmed by quantitative gene-dosage dependence and by conformance to Mendelian transmission in the two families, except for four rare variants for which transmission analysis was not possible. Linkage analysis demonstrated that each of the variants represent products of independent loci, with the exception of erythrocyte locus (RBC4), which we also detected in fibroblasts (NC27). Two allozyme polymorphisms, glyoxalase-1 (GLO1) and phosphoglucomutase-3 (PGM3) were specifically identified here based on genotypic concordance and molecular mass. Unknown fibroblast protein (NC22) may be linked to apolipoprotein E (lod score = 2.8 at theta m = theta f = 0), while a serum protein locus (SER1) may be linked to alpha-haptoglobin (lod score = 2.54 at theta m = .20, theta f = .01). Six of seven polymorphic serum loci were previously located on two-dimensional gels: alpha-1 antitrypsin (PI), Gc-globulin (GC), alpha-2 HS glycoprotein (HSGA), alpha-haptoglobin (HP), and two apolipoproteins (APOE and APOA4). Six of 17 polymorphisms detected in fibroblasts were positionally identical to polymorphic loci seen in lymphocytes. These studies indicate a minimum level of average protein charge heterozygosity of approximately 2.2% for the most predominant human cellular proteins and of 5.6% for the most predominant proteins of serum.

Alleles↗
PubMed1985

Muscle denervation increases the levels of two mRNAs coding for the acetylcholine receptor alpha-subunit.

The mRNA coding for the alpha-subunit of the acetylcholine receptor was studied in mouse leg and rat diaphragm muscle. We find that denervation of rat diaphragm results in a 7-fold increase in mRNA coding for the alpha-subunit, whereas denervation of mouse leg muscle results in approximately a 50-fold increase in alpha-subunit-specific mRNA. The relationship of the mRNAs purified from innervated and denervated muscle was investigated by SI nuclease mapping. Two mRNA species were found in both innervated and denervated muscle which differ in their 3'-untranslated region. The levels of both these mRNA species increase upon denervation of mouse leg muscle.

Animals↗