ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb00899.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11809.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:The activity of UDP‐galactose:ceramide galactosyl transferase (CGalT) has been studied in isolated oligodendroglia from bovine brain white matter and myelinating rat brain. The specific activity and activity per mg DNA are 4‐ and 10‐fold higher in rat oligodendroglia compared with neuronal perikarya from rat brain, and is higher in oligodendroglia from myelinating rat brain compared with bovine oligodendroglia. In membranes isolated from oligodendroglia, the specific activity decreased in the order endoplasmic reticulum>plasma membrane>m

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11810.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Extracts of the nematodeCaenorhabditis eleganscontain five molecular forms of acetylcholinesterase (AChE) activity that can be separated by a combination of selective solubilization, velocity sedimentation, and ion‐exchange chromatography. These are called form IA (5.2s), form IB (4.9.s), form II (6.7s), form III (11.3s), and form IV (13.0s). All except form III are present in significant amounts in rapidly prepared extracts and are probably native; form III is probably derived autolytically from form IV. Most of forms IA and IB can be solubilized by repeated extractions without detergent, whereas forms II, III, and IV require detergent for effective solubilization and may therefore be membrane‐bound. High salt concentrations are not required for, and do not aid in, the solubilization of these forms. For all forms, molecular weights and frictional ratios have been estimated by a combination of gel permeation chromatography and velocity sedimentations in both H2O and D2O. The molecular weight estimates range from 83,000 to 357,000 and only form II shows extensive asymmetry. The separated forms have been characterized with respect to substrate affinity, substrate specificity, inhibitor sensitivity, thermal inactivation, and detergent sensitivity. Judging by these properties,C. elegansis like other invertebrates in that none of its cholinesterase forms resembles either the “true” or the “pseudo” cholinesterase of vertebrates. However, internal comparison of theC. elegansforms clearly distinguishes forms IA, III, and IV as a group from forms IB and II; the former are therefore designated “class A” forms, the latter “class B” forms. Genetic evidence indicates that separate genes control class A and class B forms, and that these two classes overlap functionally. Several factors, including kinetic properties, molecular asymmetry, molecular size, and solubility, all suggest that a molecular model of the multiple cholinesterase forms observed in vertebrate electric organs probably does not apply inC. elegans. Potential functional roles and subunit structures of the multiple AChE forms within eachC. elegans

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11811.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Irreversible photolabeling by [3H]flunitraze‐pam of four proteins with apparent molecular weights 51,000 (P51), 53,000 (P53), 55,000 (P55), and 59,000 (P59) was investigated in various rat brain regions by SDS‐polyac‐rylamide gel electrophoresis, fluorography, and quantitative determination of radioactivity bound to proteins. On maximal labeling of these proteins, only 15–25% of [3H]flunitrazepam reversibly bound to membranes becomes irreversibly attached to proteins. Results presented indicate that for every [3H]flunitrazepam molecule irreversibly bound to membranes, three molecules dissociate from reversible benzodiazepine binding sites. This seems to indicate that these proteins are either closely associated or identical with reversible benzodiazepine binding sites, and supports the hypothesis that four benzodiazepine binding sites are associated with one benzodiazepine receptor. When irreversible labeling profiles of proteins P51, P53, P55, and P59were compared in different brain regions, it was found that labeling of individual proteins varied independently, supporting previous evidence that these proteins are associated with distinct benzodiazepine re

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11812.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Solubilization of rat brain α1‐adrenoceptors was performed by treatment with 6 mMCHAPS (3–[(3‐cholamidopropyl)dimethylammonio] ‐ 1 ‐ propanesulfon‐ate). The α1‐adrenoceptor antagonist [3H]prazosin was shown to bind reversibly and specifically to the soluble extract obtained after centrifugation at 150,000 ±gfor 1 h. Separation of the soluble [3H]prazosin‐bound complexes was performed by the polyethylene glycol precipitation technique followed by filtration. A Scatchard plot of the concentration‐dependent binding curve showed only one class of binding sites, with a high affinity for [3H]prazosin. Affinity of the solubilized receptors for the ligand increased as the CHAPS concentration in the assay medium decreased; the number of binding sites remained unchanged (=70 fmol/mg protein). This corresponds to a 30% recovery of original membrane sites. The solubilized receptors presented the same characteristics of specificity and stereospecificity as membrane α1‐adrenoceptors. Moreover, 150 mMNaCl was found to modulate the affinity of epinephrine for the [3H]prazosin‐bound soluble complex, as previously described for membrane preparations. Thus, CHAPS appears to be a suitable detergent for solubilizing rat brain α1‐adrenoceptors and preserving

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11813.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Acetylcholinesterase (acetylcholine acetylhy‐drolase, EC 3.1.1.7) activity and muscarinic receptor binding of homogenates from several brain structures were inhibited by β‐carbolines. The inhibition was of the noncompetitive type in the case of the enzyme and of the mixed type in the case of the receptor binding. This effect was most strongly manifested by pyridoindoles(harmane, norharmane), i.e., carbolines containing an aromatic C ring than by the corresponding piperidoindoles (tetrahy‐droharmane, tetrahydronorharmane), i.e., those with a reduced C ring. The activity of choline acetyltransferase (acetyl‐CoAxholineO‐acetyltransferase, EC 2.3.1.6) was not altered. These data are further evidence of the interactions between indoleamine derivatives and the cholinergic system. The results are discussed in terms of their possible biological si

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11814.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Prolyl endopeptidase cleaves peptide bonds on the carboxyl side of proline residues within a peptide chain. The enzyme readily degrades a number of neuropeptides including substance P, neurotensin, thyrotropin‐releasing hormone, and luteinizing hormone‐releasing hormone. The finding that the enzyme is inhibited by benzyloxycarbonyl‐prolyl‐proline, with aKiof 50 μM, prompted the synthesis of benzyloxycar‐bonyl‐prolyl‐prolinal as a potential transition state analog inhibitor. Rabbit brain prolyl endopeptidase was purified to homogeneity for these studies. The aldehyde was found to be a remarkably potent inhibitor of prolyl endopeptidase with aKiof 14 nM. ThisKiis more than 3000 times lower than that of the corresponding acid or alcohol. By analogy with other transition state inhibitors, it can be assumed that binding of the prolinal residue to the S1subsite and the formation of a hemiacetal with the active serine of the enzyme greatly contribute to the potency of inhibition. The specificity of the inhibitor is indicated by the finding that a variety of proteases were not affected at concentrations 150 times greater than theKifor prolyl endopeptidase. The data indicate that benzyloxycarbonyl‐prolyl‐prolinal is a specific and potent inhibitor of prolyl endopeptidase and that consequently it should be of value inin vivostudies on the physiological

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11815.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:The oxidation of differentially labelled glucose, pyruvate and glutamate in brain slices from rats aged 20 days to 26 months has been studied and the partition of the glucose used into the glycolytic‐tricarboxylic acid cycle pathway, the pentose phosphate pathway and the glutamate‐GABA shunt has been calculated. Over the time range 4 to 26 months, there is an approximately 20% decrease in the production of CO2via the glycolytic‐tricarboxylic acid cycle route, as there is in the rate of glucose phosphorylation. The glutamate‐GABA pathway falls by about 50% over this same time span. The broad activity of the pentose phosphate pathway falls rapidly and cannot be detected in the brains of rats aged 18 months or more, whereas the fully stimulated pathway, i.e. in the presence of the artificial electron acceptor phen‐azine methosulphate, declines only marginally over this period, falling sharply only after 23 months. The pentose phosphate pathway is stimulated by the presence of 5‐hy‐droxytryptamine and this stimulation appears to incr

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11816.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:An analytical procedure for the subcellular fractionation of rat brain cortex is presented; it consists of a two‐step procedure involving a differential centrifuga‐tion using the five‐fraction scheme and an isopycnic cen‐trifugation in continuous sucrose gradients. All fractions obtained were analyzed for their content of various constituents, such as receptor binding, uptake, and several marker enzymes. Special attention was paid to the subcellular distribution of the serotonin S2receptors; they were mainly recovered in the microsomal P fraction, but a significant amount was also associated with the mito‐chondrial (M and L) fractions. After equilibration in density gradients, serotonin S2receptors revealed two peaks, which were similarly affected after treatment with ami‐triptyline and/or yohimbine. There is no evidence to suggest that serotonin S2receptors are associated with nerve endings containing the neurotransmitter serotonin. Although three main profiles, a microsomal, a mitochondrial, and a mixed one, clearly appear from the differential centrifugation, subgroups of these main profiles were also found. For instance, the microsomal distribution patterns of serotonin S2receptors and 5′‐nucleoti‐dase are very similar, but differ from that of UDP‐galactosyltransferase. Similarly, the mitochondrial profiles of cytochrome oxidase and 5‐HT (serotonin) uptake are different. An analytical approach for brain fractionation, when performed with appropriate measurements (cytochrome oxidase, amine uptake, 5′‐nucleotidase, and receptor binding), is rapid and clearly differentiates pre‐an

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb11817.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY