共查询到20条相似文献,搜索用时 15 毫秒
1.
Ageing of potato tuber discs markedly increases the rate of phosphate uptake. This increase is partially prevented by the presence of indoleacetic acid (IAA: 50 μ M ) in the ageing medium. 32 P distribution among the various phosphorylated fractions (P1 , organic soluble phosphate, acid-insoluble phosphate) was carried out after 24 h of ageing in the presence of IAA. An equal inhibition of the labelling rates of each of the different fractions is induced by the hormone. No important effect on respiration and ATP content was observed. Moreover, IAA neither changes the total phospholipid content nor the relative distribution of 32 P between the components. These results support the hypothesis that IAA acts specifically on the development of uptake mechanisms during the ageing period. 相似文献
2.
Abstract: Proton nuclear magnetic resonance (1 H NMR) spectroscopy was used to study metabolites of the brain cortex ex vivo. The superfused brain cortex preparation was judged to be metabolically viable on the basis of the 31 P NMR spectrum (intracellular pH of 7.23 ± 0.03 and phosphocreatine/ ATP ratio of 1.21 ± 0.09). Using'H NMR a group of previously unidentified signals was detectable at 0.94, 1.22, and 1.40 ppm with a water-suppressed spin-echo sequence. These signals had shorter spin-spin relaxation times (51-54 ms) than TV-acetylaspartate and lactate (84-93 ms) and also smaller saturation factors, an indication of shorter spin-lattice relaxation times than the latter two low-molecular-weight metabolites. The unidentified signals also displayed homo-nuclear coupling to other spins in the methine region of the spectrum. Acid extraction of the brain slices or cortex from animals that were killed yielded a mixture of proteins that exhibited NMR properties matching the 1 H NMR signals in the brain cortex. The molecular mass of these thermoresistant, "mobile' proteins, which contained proline plus hydroxy-proline (9-16% of all amino acids), ranged between 8 and 40 kDa. These "new' assigμMents of1 H NMR-detectable compounds may influence interpretation of NMR data of some metabolites, as their signals are in the vicinity of the -CH3 1 H NMR peaks of lactate and alanine. 相似文献
3.
Abstract: The cellular localization of two Ca2+ -dependent protein phosphorylation systems was investigated using the kainic acid lesioning technique for the selective destruction of neurons. In one of these systems, a crude synaptosomal (P2 ) fraction was preincubated with 32 Pj for 30 min; the phosphorylation of several proteins was increased during a short subsequent incubation with veratridine plus Ca2+ . In the second system, crude synaptosomal membranes isolated from the P2 fraction were incubated with [γ-32 P]ATP; in this system, the phosphorylation of several proteins was increased in the presence of a "calcium-dependent regulator" plus Ca2+ . Kainic acid lesioning greatly reduced the amount of Ca-+ -dependent protein phosphorylation in both systems. The results indicate a predominantly neuronal localization for both Ca2+ -dependent protein phosphorylation systems. 相似文献
4.
Irreversible blockade of sigma-1 receptors by haloperidol and its metabolites in guinea pig brain and SH-SY5Y human neuroblastoma cells 总被引:3,自引:0,他引:3
We evaluated the effect of haloperidol (HP) and its metabolites on [3 H](+)-pentazocine binding to σ1 receptors in SH-SY5Y human neuroblastoma cells and guinea pig brain P1 , P2 and P3 subcellular fractions. Three days after a single i.p. injection in guinea pigs of HP (but not of other σ1 antagonists or (−)-sulpiride), [3 H](+)-pentazocine binding to brain membranes was markedly decreased. Recovery of σ1 receptor density to steady state after HP-induced inactivation required more than 30 days. HP-metabolite II (reduced HP, 4-(4-chlorophenyl)-α-(4-fluorophenyl)-4-hydroxy-1-piperidinebutanol), but not HP-metabolite I (4-(4-chlorophenyl)-4-hydroxypiperidine), irreversibly blocked σ1 receptors in guinea pig brain homogenate and P2 fraction in vitro . We found similar results in SH-SY5Y cells, which suggests that this process may also take place in humans. HP irreversibly inactivated σ1 receptors when it was incubated with brain homogenate and SH-SY5Y cells, but not when incubated with P2 fraction membranes, which suggests that HP is metabolized to inactivate σ1 receptors. Menadione, an inhibitor of the ketone reductase activity that leads to the production of HP-metabolite II, completely prevented HP-induced inactivation of σ1 receptors in brain homogenates. These results suggest that HP may irreversibly inactivate σ1 receptors in guinea pig and human cells, probably after metabolism to reduced HP. 相似文献
5.
Chefer SI Pavlova OA Zhang Y Vaupel DB Kimes AS Horti AG Stein E Mukhin AG 《Journal of neurochemistry》2008,104(2):306-315
A novel radioligand, 6-chloro-3-((2-( S )-azetidinyl)methoxy)-5-(2-fluoropyridin-4-yl)pyridine (NIDA522131), for imaging extrathalamic nicotinic acetylcholine receptors (nAChRs) was characterized in vitro and in vivo using positron emission tomography. The Kd and T1/2 of dissociation of NIDA522131 binding measured at 37°C in vitro were 4.9 ± 0.4 pmol/L and 81 ± 5 min, respectively. The patterns of radioactivity distribution in monkey brain in vivo was similar to that of 2-[18 F]fluoro-3-(2( S )-azetidinylmethoxy)pyridine (2FA), a radioligand that has been successfully used in humans, and matched the α4 β2 * nAChRs distribution. Comparison between [18 F]NIDA522131 and 2FA demonstrated better in vivo binding properties of the new radioligand and substantially greater radioactivity accumulation in brain. Consistent with [18 F]NIDA522131 elevated affinity for nAChRs and its increased lipophilicity, both, the total and non-displaceable distribution volumes were substantially higher than those of 2FA. Estimated binding potential values in different brain regions, characterizing the specificity of receptor binding, were 3–4 fold higher for [18 F]NIDA522131 than those of 2FA. Pharmacological evaluation in mice demonstrated a toxicity that was comparable to 2FA and is in agreement with a 2300 fold higher affinity at α4 β2 * versus α3 β4 * nAChRs. These results suggest that [18 F]NIDA522131 is a promising positron emission tomography radioligand for studying extrathalamic nAChR in humans. 相似文献
6.
Souad Hilmi Michel Fournier Henri Valeins Jean Claude Gandar Jacques Bonnet 《Journal of neurochemistry》1995,64(2):902-907
Abstract: Myelin membrane prepared from mouse sciatic nerve possesses both kinase and substrates to incorporate [32 P]PO4 3− from [γ-32 P]ATP into protein constituents. Among these, P0 glycoprotein is the major phosphorylated species. To identify the phosphorylated sites, P0 protein was in vitro phosphorylated, purified, and cleaved by CNBr. Two 32 P-phosphopeptides were isolated by HPLC. The exact localization of the sequences around the phosphorylated sites was determined. The comparison with rat P0 sequence revealed, besides a Lys172 to Arg substitution, that in the first peptide, two serine residues (Ser176 and Ser181 ) were phosphorylated, Ser176 appearing to be modified subsequently to Ser181 . In the second peptide, Ser197 , Ser199 , and Ser204 were phosphorylated. All these serines are clustered in the C-terminal region of P0 protein. This in vitro study served as the basis for the identification of the in vivo phosphorylation sites of the C terminal region of P0 . We found that, in vivo, Ser181 and Ser176 are not phosphorylated, whereas Ser197 , Ser199 , Ser204 , Ser208 , and Ser214 are modified to various extents. Our results strongly suggest that the phosphorylation of these serine residues alters the secondary structure of this domain. Such a structural perturbation could play an important role in myelin compaction at the dense line level. 相似文献
7.
A method for oxygenating and mixing suspensions of turbot Psetta maxima red blood cells (RBC) was tested in 31 P nuclear magnetic resonance (NMR) spectroscopy. In normoxia, the levels of inorganic phosphate (Pi) and nucleoside triphosphates (NTP) were stable up to 140 min and intracellular pH (pHi) was maintained and decreased oxygen partial pressure ( P O 2 ) from 30 to 15 and 600 Pa produced a significant fall in the intensity of NTP resonance, balanced by an increase in the Pi signal. Treatment of RBC with 0· 5 M isoproterenol during hypoxia exposure did not affect the pattern of changes in NTP or pHi induced by hypoxia and the effect was manifest only on Pi levels. 相似文献
8.
Phosphorylation and Fucosylation of Myelin Protein In Vitro by Sciatic Nerve from Developing Rats 总被引:11,自引:8,他引:3
Abstract: Proteins of the paniculate fraction of sciatic nerve of rats ranging from 1 to 55 days of age were analyzed by polyacrylamide gel electrophoresis. The major myelin protein, P0 , could not be detected at 1 day of age, but by 10 days it comprised from 15 to 20% of the particulate protein, the same proportion as in adult rats. Growth of nerve continued throughout the period studied. Rat sciatic nerves were incubated with [32 P]orthophosphate or [3 H]fucose. Particulate matter proteins from sciatic nerve (and in certain cases proteins of myelin purified from sciatic nerve) were separated by polyacrylamide disc gel electrophoresis and the distribution of protein and of radioactivity along the gels was determined. [32 P]Phosphate appeared to label all myelin proteins. Labeling with fucose was more specific; myelin basic proteins were not fucosylated. A developmental study showed that sciatic nerves from 2-day-old rats could incorporate radioactive fucose and [32 P]-phosphate into several proteins at the P0 region of polyacrylamide gels. Specific radioactivity of [3 H]fucose in P0 protein was highest in preparations from 5-day-old rats and declined by 80% over the next 5 days as it was diluted by accumulating myelin. The specific radioactivity of incorporated [32 P] phosphate was high at the early age points and declined as a result of the accumulation of compact myelin. The results indicate an association of fucosylation and/or phosphorylation with some step in the formation of myelin. 相似文献
9.
†Stefan Bluml ‡ Eli Zuckerman †Jeannie Tan † Brian D. Ross 《Journal of neurochemistry》1998,71(4):1564-1576
Abstract: Quantitative proton and quantitative proton-decoupled 31 P magnetic resonance spectroscopy (MRS) of the brain was performed in 16 patients with liver disease (10 with and six without chronic hepatic encephalopathy) and four patients with hyponatremia, as well as 20 age-matched normal subjects. Patients with hepatic encephalopathy were distinguished from controls by significant reduction in levels of cerebral nucleoside triphosphate (2.45 ± 0.20 vs. 2.91 ± 0.21 mmol/kg of brain; p < 0.0003), inorganic phosphate ( p < 0.03), and phosphocreatine ( p < 0.04). In addition of increased levels of cerebral glutamate plus glutamine and decreased concentrations of myo -inositol, patients with hepatic encephalopathy showed a reduction of total visible choline and of glycerophosphoryl-choline (0.67 ± 0.13 vs. 0.92 ± 0.20 mmol/kg of brain in controls; p < 0.005) in 1 H MRS, and of glycerophosphoryl-ethanolamine (0.40 ± 0.12 vs. 0.68 ± 0.12 mmol/kg of brain in controls; p < 0.0003) in proton-decoupled 31 P MRS. Of the reduction of "total choline," 61% was accounted for by glycerophosphorylcholine, a cerebral osmolyte. Similar metabolic abnormalities were seen in hyponatremic patients. The results are consistent with disturbances of cerebral osmoregulation and energy metabolism in patients with chronic hepatic encephalopathy. 相似文献
10.
Ronald J. T. Corbett Abbot R. Laptook Greg Tollefsbol Ben Kim 《Journal of neurochemistry》1995,64(3):1224-1230
Abstract: The goal of this study was to evaluate the potential of using the difference between the 1 H NMR frequencies of water and N -acetylaspartic acid (NAA) to measure brain temperature noninvasively. All water-suppressed and non-water-suppressed 1 H NMR spectra were obtained at a field strength of 4.7 T using a surface coil. Experiments performed on model solutions revealed a decrease in the difference between NMR frequencies for NAA and water as a linear function of increasing temperature from 14 to 45°C. Changing pH in the range 5.5–7.6 produced no discernible trends for concurrent changes in the slope and intercept of the linear relationship. There were minor changes in slope and intercept for solutions containing 80 or 100 mg of protein/ml versus no protein, but these changes were not considered to be of sufficient magnitude to deter the use of this approach to measure brain temperature. The protein content of swine cerebral cortex was found to remain constant from newborn to 1 month old (78 ± 12 mg/g; n = 41). Therefore, data collected for the model solution containing 80 mg of protein/ml were used as a calibration curve to calculate brain temperature in eight swine during control, hypothermia, ischemia, postischemia, or death, over a temperature range of 23–40°C. A plot of 61 temperatures determined from 1 H NMR versus temperatures measured from an optical fiber probe sensor implanted 1 cm into the cerebral cortex showed excellent linear agreement (slope = 1.00 ± 0.03, r 2 = 0.96). We conclude that 1 H NMR spectroscopy presents a practical means of making noninvasive measurements of brain temperature with an accuracy of better than ± 1°C. 相似文献
11.
Abstract: P0 glycoprotein, the major protein of PNS myelin, contains approximately 1 mol of covalently bound long-chain fatty acids. To determine the chemical nature of the fatty acid-protein linkage, P0 was labeled in rat sciatic nerve slices with [3 H]palmitic acid and subsequently treated with various reagents. The protein-bound palmi-tate was released by incubation with the reducing agents dithiothreitol and 2-mercaptoethanol, and with 1 M hydrox-ylamine at pH 7.5. In addition, P0 was deacylated by treatment with 10 m M NaBH4 with the concomitant production of [3 H]hexadecanol, indicating that the fatty acid is bound in a thioester linkage. This conclusion was supported further by the fact that deacylation with hydroxylamine generated free thiol groups, which were titrated with [14 C]-iodoacetamide. To identify the cysteine residue involved in the thioester linkage, [14 C]carboxyamidomethylated P0 was digested with trypsin and the resulting peptides analyzed by reversed-phase HPLC. Identification of the radioactive protein fragments by amino acid analysis and amino-terminal peptide sequencing revealed that Cys153 in rat P0 glycoprotein is the acylation site. The acylated cysteine is located at the junction of the putative transmem-brane and cytoplasmic domains. This residue is also present in the P0 glycoprotein of other species, including human, bovine, mice, and chicken. 相似文献
12.
Allen R. Rhoads Rabin Parui Ngoc-Diep Vu Robert Cadogan Paul D. Wagner 《Journal of neurochemistry》1993,61(5):1657-1666
Abstract— Secretion of catecholamines by rat PC12 cells is strongly stimulated by extracellular ATP via a P2 -type pur-inergic receptor. ATP-induced norepinephrine release was inhibited 80% when extracellular Ca2+ was absent. Only four nucleotides, ATP, ATPγS, benzoylbenzoyl ATP (BzATP), and 2-methylthio-ATP, gave substantial stimulation of norepinephrine release from PC12 cells. ATP-induced secretion was inhibited by Mg2+ , and this inhibition was overcome by the addition of excess ATP suggesting that ATP4- was the active ligand. ATP-induced secretion of catecholamine release was enhanced by treatment of cells with pertussis toxin or 12- O -tetradecanoylphorbol 13-acetate. The stimulatory effects of 12- O -tetradecanoyl-phorbol 13-acetate and pertussis toxin on norepinephrine release were additive. After brief exposure of intact cells to the photoaffinity analog, [α-32 P]BzATP, two major proteins of 44 and 50 kDa and a minor protein of 97 kDa were labeled. An excess of ATP-γS and BzATP but not GTP blocked labeling of the proteins by [32 P]BzATP. Labeling of the 50-kDa protein was more sensitive to competition by 2-methylthio-ATP than the other labeled proteins, suggesting that the 50-kDa protein represents the P2 receptor responsible for ATP-stimulated secretion in these cells. 相似文献
13.
C. J. Chang H. Ishii H. Yamamoto T. Yamamoto M. Spatz 《Journal of neurochemistry》1993,60(4):1483-1490
Abstract: To expand on the nature of regional cerebral vulnerability to ischemia, the release of dopamine (DA) and dopaminergic (D1 and D2 ) receptors were investigated in Mongolian gerbils subjected to bilateral carotid artery occlusion (15 min) alone or with reflow (1–2 h). Extracellular cortical and striatal content of DA and its metabolites was measured by microdialysis using HPLC with electrochemical detection. The kinetic properties of D1 and/or D2 receptor binding sites were determined in cortical and striatal membranes with the use of radiolabeled ligands (125 I-SCH23982 and [3 H]YM-09151-2, respectively). The ischemic release of DA from the striatum was greater (400-fold over preischemic level) than that from the cortex (12-fold over preischemic content). The affinity for the D1 -receptor ligand was lower ( K D = 1.248 ± 0.047 n M ) after ischemia than that for sham controls ( K D = 0.928 ± 0.032 n M, p < 0.001). The number of binding sites for D2 receptors decreased in striatum ( B max = 428 ± 18.4 fmol/mg of protein) after ischemia compared with sham controls ( B max = 510 ± 25.2 fmol/mg of protein, p < 0.05). D1 or D2 binding sites were not changed either in the ischemic cortex or postischemic striatum and cortex. The findings strongly suggest that the ischemic release of DA from striatum is associated with early transient changes in D1 - and D2 -mediated DA neurotransmission. 相似文献
14.
The metabolism of glucose was studied in Lactococcus lactis subsp. lactis CNRZ 125 by 13 C NMR. The initial rate of glucose utilization was higher for exponential phase cells than for stationary phase cells [150 vs 85 nmol g (dry wt)-1 s-1 ]. 31 P NMR was used to determine changes in glycolytic phosphorylated intermediates (fructose-1,6-diphosphate, dihydroxyacetone phosphate and phosphoglycerate). The internal pHs of L. lactis subsp. lactis CNRZ 141 and CNRZ 125 were also measured by 31 P NMR as a function of the external pH during growth. When the external pH was 6·8, the internal pHs of strain CNRZ 141 and CNRZ 125 were similar, 7·4. After the external pH had decreased to 5·5, the internal pH of strain CNRZ 141 had declined by 0·6 unit, whereas that of strain CNRZ 125 had decreased by only 0·2 unit of pH. 相似文献
15.
Jussi-Pekka Usenius Pauli Vainio Juha Hernesniemi † Risto A. Kauppinen 《Journal of neurochemistry》1994,63(4):1538-1543
Abstract: We have studied 14 patients with different grades of astrocytomas using 1 H NMR spectroscopy in vivo. Typically, astrocytomas exhibited a low N -acetyl-aspartate peak, a prominent signal from choline group-containing compounds, and lactate in the 1 H NMR spectra in vivo. The uncorrected choline/creatine + phosphocreatine peak area ratios were higher in tumors than in normal brain tissue. Absolute concentration of choline-containing compounds (1.74 ± 0.09 mmol/L) in the normal brain tissue was not different in any grade of astrocytoma, but total creatine concentration in healthy brain (7.49 ± 0.30 mmol/L) was higher than that in grade IV astrocytomas (4.84 ± 0.89 mmol/L). Relaxation constants of choline-containing compounds did not differ in tumors from those determined in normal brain. Perchloric acid extracts of biopsy samples from 35 astrocytomas and 13 samples of normal temporal white matter were analyzed with 1 H NMR. Total concentration of choline-containing compounds did not differ between controls and any grade of astrocytoma when the quantification was done in vitro. It is interesting that phosphorylcholine concentration was about twofold greater in grade IV astrocytomas than in controls or other grades of astrocytomas. We conclude that high phosphorylcholine in grade IV astrocytomas may be an indicator of degree of malignancy. The proportional changes within the group of choline-containing compounds observed in vitro were not reflected in the NMR properties of choline signal in vivo. 相似文献
16.
Leng-Chu Shi Hoau-Yan Wang Joel Horwitz Eitan Friedman 《Journal of neurochemistry》1996,67(4):1478-1484
Abstract: Platelet-activating factor (PAF) may be a neuromodulator involved in neural cell differentiation, cerebral inflammation, and ischemia. The PAF receptor is a member of the G protein-coupled receptor superfamily. In the present study, we sought to define the specific G protein(s) that mediate PAF-stimulated phosphoinositide (PI) metabolism in an immortalized hippocampal cell line, HN33.11. PAF increased the production of 3 H-labeled inositol phosphates (IPs) with EC50 values of 1.2–1.5 n M . The effect of PAF on 3 H-IPs formation was completely blocked by the PAF antagonist BN 50739 at a concentration of 300 n M . Pertussis toxin pretreatment attenuated PAF-stimulated 3 H-IPs production by 20–30% ( p < 0.05). Consistent with a role for Gi1/2 in this response, antiserum against Gαi1/2 blocked the response to a similar degree. Pretreatment of permeabilized cells with Gαq/11 antiserum attenuated the response by 70% ( p < 0.05), suggesting a role for Gq/11 in mediating the PAF response in this cell line. Stimulation with PAF increased [α-32 P]-GTP binding to both Gαq and Gαi1/2 proteins. Moreover, specific [3 H]PAF binding sites coprecipitated with Gαq and Gαi1/2 proteins. The results suggest that PAF-stimulated PI metabolism in HN33.11 cells is mediated by both Gq and Gi1/2 proteins. 相似文献
17.
Richard E. Roberts Charles A. Marsden David A. Kendall 《Journal of neurochemistry》1997,69(3):1291-1299
Abstract: Recent in vivo microdialysis studies have demonstrated the presence of extracellular levels of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3 ] that can be increased in a concentration-dependent manner by muscarinic receptor activation. The aim of the present study was to determine whether extracellular levels of Ins(1,4,5)P3 could be measured in vitro. Despite rapid increases in internal Ins(1,4,5)P3 levels after stimulation with 1 m M carbachol, there was no change in external levels in both rat brain cortical slices and human neuroblastoma SH-SY5Y cells. Suprafusion of myo -[3 H]inositol-prelabelled hippocampal slices with 1 m M carbachol caused an increase in 3 H-inositol phosphates over basal levels in the perfusate after 10 min, reaching a peak (223 ± 56% of basal) 20 min after suprafusion with carbachol was started. This response to carbachol was potentiated in the presence of 30 m M K+ . Analysis of the individual 3 H-inositol phosphates in the perfusate revealed that levels of [3 H]inositol monophosphate, [3 H]inositol bisphosphate, [3 H]inositol trisphosphate, and [3 H]inositol tetrakisphosphate were all significantly increased. A similar increase in extracellular 3 H-inositol phosphates was demonstrated in SH-SY5Y cells incubated with 1 m M carbachol for 30 min. This response was again enhanced by 30 m M K+ , although the intracellular response was not potentiated. Possible roles for extracellular inositol phosphates are discussed. 相似文献
18.
Traba J Froschauer EM Wiesenberger G Satrústegui J Del Arco A 《Molecular microbiology》2008,69(3):570-585
Sal1p, a novel Ca2+ -dependent ATP-Mg/Pi carrier, is essential in yeast lacking all adenine nucleotide translocases. By targeting luciferase to the mitochondrial matrix to monitor mitochondrial ATP levels, we show in isolated mitochondria that both ATP-Mg and free ADP are taken up by Sal1p with a K m of 0.20 ± 0.03 mM and 0.28 ± 0.06 mM respectively. Nucleotide transport along Sal1p is strictly Ca2+ dependent. Ca2+ increases the V max with a S 0.5 of 15 μM, and no changes in the K m for ATP-Mg. Glucose sensing in yeast generates Ca2+ transients involving Ca2+ influx from the external medium. We find that carbon-deprived cells respond to glucose with an immediate increase in mitochondrial ATP levels which is not observed in the presence of EGTA or in Sal1p-deficient cells. Moreover, we now report that during normal aerobic growth on glucose, yeast mitochondria import ATP from the cytosol and hydrolyse it through H+ -ATP synthase. We identify two pathways for ATP uptake in mitochondria, the ADP/ATP carriers and Sal1p. Thus, during exponential growth on glucose, mitochondria are ATP consumers, as those from cells growing in anaerobic conditions or deprived of mitochondrial DNA which depend on cytosolic ATP and mitochondrial ATPase working in reverse to generate a mitochondrial membrane potential. In conclusion, the results show that growth on glucose requires ATP hydrolysis in mitochondria and recruits Sal1p as a Ca2+ -dependent mechanism to import ATP-Mg from the cytosol. Whether this mechanism is used under similar settings in higher eukaryotes is an open question. 相似文献
19.
Raquel Fernández-Durango José A. de Juan Horatio Zimman Francisco J. Moya Mario Garcia de la Coba Arturo Fernández-Cruz 《Journal of neurochemistry》1994,62(4):1482-1488
Abstract: Specific endothelin (ET) binding sites were characterized in membranes prepared from human cerebral cortices using binding assay and cross-linking analysis. The presence of immunoreactive (IR) ET-1 was studied by radioimmunoassay. Saturation binding experiments revealed that the K D and B max for 125 I-ET-1 and 125 l-ET-3 to membranes from gray matter were 25 ± 6 pM and 115 ± 15 fmol/mg of protein and 24 ± 5 p M and 108 ± 13 fmol/mg of protein, respectively. Similar results were obtained for white matter. In the presence of 10 n M sarafotoxin-6c, which is selective for ETB receptors, 125 I-ET-1 and 125 l-ET-3 binding was totally abolished. However, in the presence of 1 μ M BQ123, which is selective for ETA receptors, both bindings were not affected. These results suggest that the human cerebral cortex contains only ETB receptors. Cross-linking of 125 I-ET-1 and 125 l-ET-3 to membranes with disuccinimidyl suberate resulted in the labeling of two bands of 48 and 31 kDa. Concentrations of IR-ET-1 in the gray and white matter were 7.0 ± 3.2 and 2.5 ± 1.7 fmol/g wet weight, respectively. The demonstration of high-affinity ETB receptors and the presence of IRET-1 suggest that the peptide may act as a neurotransmitter or neuromodulator in the human cerebral cortex. 相似文献
20.
Shinichi Hatta Hiroki Ozawa Toshikazu Saito Hideyo Ohshika 《Journal of neurochemistry》1994,63(3):1104-1110
Abstract: The ability of the tubulin dimer to interact with and to modulate the Gi function inhibiting adenylyl cyclase was examined in cerebral cortex membranes from 2-month-old and 24-month-old rats. The hydrolysis-resistant GTP analogue 5'-guanylylimidodiphosphate (GppNHp)-dependent inhibition of adenylyl cyclase was significantly decreased in cerebral cortex membranes from 24-month-old rats. Tubulin, prepared from rat brains by polymerization with GppNHp, caused inhibition of adenylyl cyclase (∼28%) in 2-month-old rats. Tubulin-GppNHp-dependent inhibition of adenylyl cyclase in 24-month-old rats was significantly attenuated (∼15%). In 2-month-old rats, when tubulin, polymerized with the hydrolysis-resistant photoaffinity GTP analogue [32 P] P 3 (4-azidoanilido)- P 1 -5'-GTP ([32 P]AAGTP), was incubated with cerebral cortex membranes, AAGTP was transferred from tubulin to Giα . Transfer of AAGTP from tubulin to Giα was reduced in 24-month-old rats. Furthermore, photoaffinity labeling of [32 P]AAGTP to Giα in cortex membranes was significantly decreased in 24-month-old rats. No differences were observed in the amounts of Gsα , Giα , or Gβ subunits and tubulin, estimated by immunoblotting, in cortex membranes from 2-month-old and 24-month-old rats. These results suggest that the ability of tubulin to interact with Gi and thereby modulate the inhibitory regulation of adenylyl cyclase is reduced in the cerebral cortex of 24-month-old rats. 相似文献