摘要:

Abstract:Suriclone (RP 31,264), like zopiclone (RP 27, 267), belongs to the family of cyclopyrrolones and is chemically entirely different from the benzodiazepines (BZDs). However, it possesses a pharmacological profile close to that of the BZDs and proved to be useful in therapeutics as an anxiolytic agent. In the present paper it is shown that suriclone possesses a high affinity for flunitrazepam binding sites and that tritiated suriclone binds specifically with high affinity in rat hippocampus (Kd= 0.44 ± 0.03 nM) and rat cerebellum (Kd= 0.53 ± 0.12 nM. Further, suriclone binding sites are recognized by BZDs or zopiclone, similarly in the two regions. The affinities of four BZD derivatives–nitrazepam, flunitrazepam, diazepam, and chlordiazepoxide–are similar for suriclone and flunitrazepam binding sites. Suriclone binding sites are, like flunitrazepam sites, protected from thermal inactivation by γ‐aminobutyric acid (GABA) (10 μM), but only flunitrazepam binding is enhanced by GABA. It could be postulated from this that suriclone interacts with a subpopulation of receptors that might be modulated differently from flunitrazepam binding sites. Our results indicate that suriclone could be a new probe for investigating the so‐called BZ

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08023.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Nerve ending particles (synaptosomes) were prepared from pieces of rat and human brain and from brain homogenate that had been frozen and thawed under a variety of conditions. Their purity, as judged by electron microscopy, and performance in terms of a number of metabolic and functional parameters [accumulation of tissue potassium, respiration, release of transmitter amino acids, and the responses on these indices to depolarisation by veratrine (VX)] were compared with those of fresh tissue‐derived synaptosomes. It was found that rapid freezing and/or slow thawing severely impaired the subsequent performance of incubated synaptosomes. In contrast, synaptosomes from tissue frozen slowly and thawed rapidly showed relatively good retention of morphology and metabolic performance. It was better to use whole (1‐5 g) pieces of tissue than tissue homogenate: the synaptosome fraction from frozen tissue pieces contained 80% of the proportion of identified synaptosomes found in the fresh tissue synaptosome fraction, its respiratory rate was 65%, and its tissue potassium content 70% of that of fresh controls. Moreover, it responded to VX or potassium stimulation by showing increased respiratory rate, decreased tissue potassium, and increased release of neurotransmitter amino acids, to an extent that was comparable to that of fresh tissue fractions. Thus, preparations from frozen rat and human brain were shown to be metabolically and functionally active, and can be used for a variety of neurotransmitter‐related st

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08024.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Rat brain slices, prelabeled with [3H]noradrenaline, were superfused and exposed to K+depolarization (10‐120 mMK+) or to veratrine (1‐25 μM). In the absence of extracellular Ca2+veratrine, in contrast to K+‐depolarization, caused a substantial release of [3H]noradrenaline, which was completely blocked by tetrodotoxin (0.3 μM). The Ca2+antagonist Cd2+(50 μM), which strongly reduced K+‐induced release in the presence of 1.2 mMCa2+, did not affect release induced by veratrine in the absence of extracellular Ca2+. Ruthenium red (10 μM), known to inhibit Ca2+‐entry into mitochondria, enhanced veratrine‐induced [3H]noradrenaline release. Compared with K+depolarization in the presence of 1.2 mMCa2+, veratrine in the absence of Ca2+caused a somewhat delayed release of [3H]noradrenaline. Further, in contrast to the fractional release of [3H]noradrenaline induced by continuous K+depolarization in the presence of 1.2 mMCa2+, that induced by prolonged veratrine stimulation in the absence of Ca2+appeared to be more sustained. The data strongly suggest that veratrine‐induced [3H]noradrenaline release in the absence of extracellular Ca2+is brought about by a mobilization of Ca2+from intracellular stores, e.g., mitochondria, subsequent to a strongly increased intracellular Na+concentration. This provides a model for establishing the site of action of drugs that alter the stimulussecretion coupling process in central noradrenergi

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08025.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Tricyclic antidepressant drugs inhibit [3H]imipramine binding to the rat brain cortex in a competitive manner, giving linear Hofstee plots and Hill coefficients of approximately 1.0. Serotonin, the only neurotransmitter to inhibit [3H]imipramine binding, does so in a complex manner, exhibiting a Hill coefficient of 0.40‐0.50. Nontricyclic inhibitors of serotonin uptake such as fluoxetine, paroxetine, norzimelidine, and citalopram inhibit [3H]imiprarnine binding in the same complex manner as serotonin. These results are interpreted as suggesting that [3H]imipramine binds to a site associated with the serotonin uptake system but different from either the substrate recognition site for serotonin or the site of action of the nontricyclic inhibitors of neuronal uptake of serotoni

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08026.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Addition of 10μM guanyl‐5′‐ylimidodiphos phate at 30° or 0° to guinea pig brain particulates instantaneously evoked nearly 50% inhibition of adenylate cyclase activity as determined after removal of the GTP analog by washing of the particulates. The inhibitory state, once formed, persisted for at least 60 min as long as the preparation was kept either in a medium devoid of the analog (0‐30°) or in its presence at 0°. During incubation at 30° in the presence of the analog, however, the inhibited or nontreated enzyme showed a gradual increase in enzyme activity. Both the inhibitory and the activating effects of the analog were saturable, with a half‐maximal concentration of about 1.0 μM, and were antagonized by simultaneous addition of GTP, GDP, and GMP (in decreasing order). The persistently inhibited enzyme enabled the detection of marked stimulation by norepinephrine and histamine, whereas these amines showed only marginal stimulation of the enzyme before treatment with the analog. Formation of such a persistent inhibitory state appears to be specific

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08027.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:The conventional assay for measuring cell surface antigens‐the quantitative absorption of antibody by tissue homogenates‐proved inadequate when used to determine the level of Thy‐1 glycoprotein in rat nerves and peripheral ganglia. In this paper we report that the binding of1251‐labelled Fab fragments of a monoclonal anti‐Thy‐1 antibody to cryostat sections is sufficiently sensitive to give consistent estimates of the Thy‐1 level on single samples of even small nerves. Observed levels of Thy‐1 were generally higher than had previously been thought, and in particular we found no nerves totally lacking the antigen. The lowest levels (6‐10 pmol/mg protein) were in peripheral nerves with a large motor component. Autonomic and sensory nerves had higher levels (15‐20 pmol/mg protein). The highest levels were on the optic nerve (34 pmol/mg protein), superior cervical sympathetic ganglion (40 pmol/mg protein), and the cerebellar vermis (46 pmoles/mg protein; the only brain region examined in this study). From a practical point of view, the cryostat assay has the advantage that measurements of Thy‐1 can be done on sections from the same series as is used for immunohisto

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08028.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Microvessels isolated from rat cerebral cortex consist mainly of capillaries (>85%). Fresh, intact microvessel preparations have been analyzed by radioligand binding techniques for muscarinic receptors. Scatchard analysis of specific quinuclidinyl benzilate (QNB) binding indicates that microvessels possess a large number of muscarinic sites (914 fmol/mg protein) of high affinity (Kd= 0.034 nM). The association and dissociation rate constants (0.37 min−1nM−1and 0.0067 min−1, respectively) yield an equilibriumKdof 0.018 nM. Displacement of [3H]QNB by muscarinic ligands and control substances is typical of muscarinic receptors. The results indicate that cerebral microvessels possess a large population of muscarinic rece

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08029.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Ependymins β and γ (MW 32,000 and 26,000 daltons) are two secreted goldfish brain glycoproteins that exhibit a specifically enhanced turnover rate when the animals successfully acquire a new pattern of swimming behaviour. Both proteins are bound identically to concanavalin A and can be isolated from brain extracellular fluid and from brain cytoplasm by lectin affinity chromatography. Radioimmunoassay data, using purified125I‐ labeled ependymins and antisera directed against ependymin β or ependymin γ, show complete cross‐reactivity between the two proteins. It is demonstrated by Scatchardplot analysis that the antisera recognize identical immunological determinants in both proteins. The amino acid composition of the ependymins is similar, and several identical polypeptide fragments are obtained after limited proteolysis withStaphylococcus aureusprotease. The proteins are capable of forming complexes of the compositions γ2, βγ, and β2. A protease present in the extracellular fluid of goldfish brain promotes proteolysis of ependymin β to ependymin γ. The finding that ependymin γ is physiologically derived from ependymin β suggests the possibility that ependymin β might exert its biological function during consolidation of new behavioural patterns via smaller polyp

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08030.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Membranes of the secretory vesicles from bovine adrenal medulla were investigated for the presence of the endogenous protein phosphorylation activity. Seven phosphoprotein bands in the molecular weight range of 250,000 to 30,000 were observed by means of the sodium dodecyl sulphate electrophoresis and autoradiography. On the basis of the criteria of molecular weight, selective stimulation of the phosphorylation by cyclic AMP (as compared with cyclic GMP) and immunoprecipitation by specific antibodies, band 5 (molecular weight 60,300) was found to represent the phosphorylated form of the secretory vesicle‐bound tyrosine hydroxylase. The electrophoretic mobility, the stimulatory and inhibitory effects of cyclic AMP in presence of Mg2+and Zn,2+respectively, and immunoreactivity toward antibodies showed band 6 to contain two forms of the regulatory subunits of the type II cyclic AMP‐dependent protein kinase, distinguishable by their molecular weights (56,000 and 52,000, respectively). Phosphorylation of band 7 (molecular weight 29,800) was stimulated about 2 to 3 times by Ca2+and calmodulin in the concentration range of both agents believed to occur in the secretory tissues under physiological conditi

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08031.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY

摘要:

Abstract:Studies were undertaken to optimize the conditions for isolation andin vitrotranslation of poly(A)‐containing mRNA from human postmortem brain. The comparison of several methods for preparation of mRNA from frozen mouse brain indicated that although the yield of mRNA was increased using polysomes prepared in the presence of ribonucleoside vanadyl complexes and subsequently extracted with guanidinium thiocyanate, the translation products were indistinguishable from those synthesized by total cellular RNA directly extracted from tissue with guanidinium thiocyanate. The oligo d(T)‐cellulose‐purified poly(A)‐containing mRNA preparations were translatedin vitroin a rabbit reticulocyte lysate in the presence of L‐[35S]methionine. Messenger RNA from frozen mouse brain stimulated protein synthesis from 9‐ to 20‐fold over endogenous mRNA. Over 450 polypeptides were reproducibly synthesized and separated by two‐dimensional polyacrylamide gel electrophoresis (PAGE): size classes up to 130,000 daltons were present. Direct extraction of RNA from frozen human cerebral cortex and cerebellum with guanidinium thiocyanate followed by oligo d(T)‐cellulose chromatography yielded 1.8 μg/g and 2.0 μg/g, respectively, of poly(A)‐containing mRNA; this represents a two‐ to fourfold increase over our earlier results. In the rabbit reticulocyte translation system human brain mRNA stimulated protein synthesis nearly threefold over endogenous mRNA. Compared with earlier studies, the number of newly synthesized polypeptides was increased by 30%. Over 300 species were separated by two‐dimensional PAGE, and size classes up to 130,000 daltons were present, as compared to 70,000 in an earlier report. The polypeptides synthesized by human cerebral cortex and cerebellum were indistinguishable. However, several appeared to be uniquely human when compared with the products synthesized by mouse brain mRNA. The method described for the preparation of postmortem human brain mRNA eliminates the need to prepare polysomes, which are recovered in variable and low yield from the postmortem human brain. The procedure appears applicable to studies on the synthesis of moderately large human brain polypeptides and for investigations of brain protein polymorphism when relatively large numbers of products

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1983.tb08032.x

出版商:Blackwell Publishing Ltd    

年代:1983

数据来源: WILEY