ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07288.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07289.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:Incubation of cultured bovine adrenal medullary cells withp‐chloromercuribenzoate (50–500 μM), a sulfhy‐dryl‐reacting agent, caused an increase in the secretion of catecholamines.p‐Chloromercuriphenyl sulfonate, ap‐chlo‐romercuribenzoate analogue that poorly penetrates the cell membrane, caused a similar increase in catecholamine secretion. In both cases, catecholamine secretion was dependent on extracellular Ca2+. Furthermore,p‐chloromercuribenzoate caused both45Ca2+influx into the cells and an increase in the intracellular free Ca2+concentration. The increases in catecholamine secretion and45Ca2+influx behaved similarly in relation to p‐chloromercuribenzoate concentration. The time courses of the increased secretion,45Ca2+influx, and intracellular free Ca2+concentration byp‐chloromercuri‐benzoate were also quite similar. The stimulation of cate‐holamine secretion byp‐chloromercuribenzoate was reversed by washing the cells with dithiothreitol‐containing medium, but not by dithiothreitol‐free medium. When the cells were treated withp‐chloromercuribenzoate, dopamineβ‐hydroxylase, an enzyme present in the chromaffin granules along with catecholamines, was also released. However,p‐chloromercuribenzoate did not cause release of phenyletha‐nolamine‐JV‐methyltransferase, an enzyme present in the cytoplasm. These results indicate that catecholamine secretion due top‐chloromercur

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07290.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:The interaction of the nonselective opioid ligand [3H]bremazocine and of thek‐opioid [3H]U69593 with thek‐receptor was investigated in guinea‐pig cortical membranes. Each radioligand bound to a single population of high‐affinity sites, although [3H]U69593 apparently recognised only 70% of those sites labelled by [3H]bremazocine. Naloxone and the selective ligands U69593 and PD117302 exhibited full inhibition of the binding of both radioligands. Kinetic analysis demonstrated biphasic rates of association and dissociation for both [3H]bremazocine and [3H]U69593. Detailed analysis of the binding of [3H]U69593 revealed that the fast rate of association was dependent on radioligand concentration, in contrast to the slow rate, which was independent of ligand concentration. Guanylyl‐5′‐imidodiphosphate (GppNHp) inhibited binding of [3H]U69593; saturation analysis demonstrated that the inhibitory effects of GppNHp resulted in a decrease in affinity without any significant change in binding capacity. GppNHp attenuated the formation of the slow component of [3H]U69593 binding, while accelerating the fast component. The data are consistent with the formation of a high‐affinity complex between thek‐receptor and a guanine nucleotide binding protein. Guanine nucleotides promote the dissociation of this ternary complex and the stabilisation of a lower‐affinity st

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07291.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:Mitochondrially bound hexokinase (ATP‐D‐hexose‐6‐phosphotransferase; EC 2.7.1.1) was dissociatively extracted from normal rat brains and intracerebral and subcutaneous implants of the 36B‐10 glioma. At least 70% of the total hexokinase enzyme activity in normal and glioma tissue was associated with the mitochondrial fraction. Purification of the crude tissue extracts by ion‐exchange and affinity chromatography followed by analysis with sodium dodecyl sul‐fate‐polyacrylamide gel electrophoresis revealed a successive purification of the enzyme to homogeneity with a molecular size of 98 kilodaltons. Enzyme kinetics with glucose or 2‐deoxyglucose (2‐DG) as the substrate were measured spec‐trophotometrically by coupling the appropriate reactions to either NADPH or NAD+formation. TheKmof hexokinase with glucose as the substrate in the intracerebral glioma (0.138m.M) and subcutaneous glioma (0.183mM) tissues was 2.1 ‐2.7‐fold higher than that observed in normal brain tissue (0.067mM) (p<0.001). No significant differences were observed in theKmfor hexokinase with 2‐DG as the substrate in the glioma and normal brain tissue. The phosphorylation ratio for normal brain was 0.320 and was increased in the intracerebral glioma to 0.694 and in the subcutaneous glioma to 0.519. The ratios of deoxyglucose and glucose volumes of distribution in normal brain and intracerebral glioma tissues were 1.70 and 1.85, respectively. The lumped constants calculated directly from the phosphorylation ratios and the volumes of distribution of deoxyglucose and glucose were 0.517 in normal brain and 1.168 in intracerebral glioma. Our results indicate the lumped constant is increased 2.26‐fold in intracerebral glio

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07292.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:The expression of τ mRNA and of the corresponding encoded protein variants was studied during postnatal development in two brain regions differing in their timing of differentiation: the cerebral neocortex and the cerebellum, (a) The expression of r mRNA was different in the two regions. Maximal contents were found at early stages in the cerebral neocortex, with a 10‐fold decrease at later stages. In the cerebellum, two peaks of τ mRNA were observed soon after birth and in adulthood, with minimal values at postnatal day 6. (b) The expression of total τ proteins was similar to that of their encoding mRNAs in the cerebral neocortex, i.e., high concentrations after birth and low contents at later stages. In contrast, two peaks of τ proteins were observed in the cerebellum: the first perinatally and the second with a maximum at postnatal day 15. (c) Both in the cerebral neocortex md especially in the cerebellum, increasing concentrationsof mature τ variants were expressed at late developmental stages, i.e., when total τ protein contents were decreased. In conclusion, the fluctuations in expression of τ and of its encoding mRNA seen in the cerebellum seem to reflect differences in the timing of differentiation of the various cell types, i.e., the macroneurons and the interneurons, present in this brain region. The adult τ variants appear in both the neocortex and the cerebellum only at late developmental stages, i.e., when most of the circuitry has been established, although these two regions markedly differ in their timing of differ

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07293.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:A panel of monoclonal antibodies (mAbs) was developed to identify polypeptides sorted in subtypes of brain coated vesicles (CVs) and to separate these by immunoprecipitation. The corresponding antigen of some of the mAbs elicited by CV components was present also in synaptosomal plasma membrane, synaptic vesicles, or microsomes. On im‐munoblots the mAbs reacted with constitutive brain CV proteins, with cargo molecules, and with a novel CV component that interacts with the actin cytoskeleton. Analysis of radio‐iodinated brain CVs immunoprecipitated with a tubulin antibody revealed that all brain CVs contained tubulin. The mAb A‐7C11 recognized a 40‐kilodalton (kDa) polypeptide on the clathrin coat and immunoprecipitated one‐quarter of the total brain CVs. The mAb S‐l1 D9 reacted with a 44‐kDa antigen and immunoprecipitated 25% of the CVs. This antigen (44 kDa) was present in synaptic vesicles and synaptosomal membrane as well. Moreover, this mAb (S‐11 D9) reacted with a polypeptide of 56 kDa detected only in synaptosomal membrane. A mAb (C‐10B2) that reacted with one of the clathrin light chains (LCb) immunoprecipitated 90% of the brain CVs. One of the mAbs immunoprecipitated a CV subtype that displayed a reversed ratio of the clathrin LCs (LCa>LCb). Each of the mAbs yielded different im‐munofluorescent staining patterns of vesicles in culture cell types that included nerve growth factor‐differentiated PC 12 cells, neuroblastoma cells, and Madin Darby bovine kidney cells. The data suggest that in brain tissue there is a heterogeneous population of CVs with different polypeptide compositions and subcellular distributions and that each of these subtypes performs a differen

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07294.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:In homogenates of rat cerebral neocortex prostaglandin D2(PGD2) was found to be quantitatively the main PG biosynthesized by a cytosolic PGD synthetase from en‐dogenously released arachidonic acid. Amounts of 628 ng/g wet weight were found after 30‐min incubation periods compared with basal levels of 2.3 ng/g wet weight. In human cerebral cortex, whether obtained at biopsy or postmortem, only small amounts of PGD2(4.5–11.7 ng/g wet weight/30 min) were formed. Furthermore, PGD2, added to homogenates of human biopsy temporal cortex, was converted efficiently into 9α,11β‐PGF2by a NADPH‐dependent 11‐ke‐toreductase as has been reported in other human tissues (liver and lung). PGF2αwas determined directly as the fl‐butylbo‐ronate derivative. It became clear that 9α,11β‐PGF2was formed in considerably greater amounts than PGF2αand that other metabolites are also formed. These results can account for the low amounts of PGD2found in incubations of human brain tissue. The rat brain does not contain 11‐ketoreductase activity. The present results indicate that the 9α, 11β‐PGF2must be considered along with other eicosanoids in pathoph

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07295.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:Glutamate (10–100 μM) reversibly depolarizes guinea‐pig cerebral cortical synaptosomes. This does not appear to be because of a conventional autoreceptor. Neither kainate at 1mM, 100 μMn‐methyl‐D‐aspartate (NMDA), 100 μML‐2‐amino‐4‐phosphonobutanoate (APB), nor 100 μMquisqualate affects the Ca2+‐dependent release of glutamate from suboptimally depolarized synaptosomes. However, kainate, quisqualate, and the quisqualate agonists β‐N‐ox‐alylamino‐L‐alanine and α‐amino‐3‐hydroxy‐5‐methylisox‐azole propionate cause a slow Ca2+‐independent release of glutamate from polarized synaptosomes. However, unlike kainate, quisqualate does not inhibit the acidic amino acid carrier. APB, NMDA, and the NMDA receptor‐mediated neurotoxin β‐N‐methylamino‐L‐alanine do not influence Ca2+‐independent release at 100 μM. The depolarization of the plasma membrane by glutamate can be mimicked by D‐aspartate, can be blocked by the transport inhibitor dihydro‐kainate, and is accompanied by the net uptake of acidic amino acids. L‐Glutamate or D‐aspartate at 100 μMincreases the cytoplasmic free Ca2+concentration. D‐aspartate at 100 μMcauses a Ca2+‐dependent release of endogenous glutamate, superimposed on the Ca2+‐independent heteroexchange with glutamate through the acidic amino acid carrier. The results suggest that the glutam

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07296.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY

摘要:

Abstract:The effect of intermittent light stimulation (ILS) on the distribution of thiamine derivatives in three brain areas (occipital, motor, and premotor) was compared in photosensitive and nonphotosensitive baboons. ILS induces paroxysmal discharges in the motor and premotor areas of photosensitive animals only. In baboons submitted to ILS, thiamine triphosphate (TTP) decreases in both photosensitive and nonphotosensitive animals; thiamine monophosphate (TMP) increases in photosensitive animals, which present ILS‐induced paroxysmal discharges, whereas it is unaffected in nonphotosensitive animals. The variations are the most significant in the occipital (visual) cortex. A consumption of TTP may result from electrical activity induced by light stimulation in the occipital area. No correlation between ILS‐induced paroxysmal activity and a decrease in TTP contents was found. However, photosensitive animals are affected differently from nonphotosensitive animals, as their content of TMP in the cerebral cortex increases on stimulation. However, as long as the exact role of thiamine compounds in relation to membrane excitability in the nervous system remains unknown, it is impossible to conclude whether the differences observed in the metabolism of thiamine compounds are the cause or the consequence of the photosensitivity in the baboonPapio pa

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1989.tb07297.x

出版商:Blackwell Publishing Ltd    

年代:1989

数据来源: WILEY