摘要:

Abstract:S‐Antigen is a soluble cell protein unique to the retina and pineal gland. In the former, it is a well‐characterized molecule that participates in light‐induced signal transduction in photoreceptor cells. In the latter, the functional role is presently not known. The expression of S‐antigen and its mRNA was examined in the rat retina and pineal gland throughout the diurnal cycle and with light interruption of the dark cycle. A cDNA for rat S‐antigen was isolated from a pineal gland library to examine the mRNAs. A 1.7‐kb mRNA for S‐antigen was observed in both the pineal gland and the retina. Retinal S‐antigen mRNA was expressed throughout the diurnal cycle and increased with light interruption of the dark cycle. In contrast, pineal gland S‐antigen mRNA levels were detectable only during the dark and were absent preceding and during light. The phenotypic expression of immunoreactive S‐antigen, identified with two S‐antigen monoclonal antibodies (MAbs), MAb A9C6 and MAb CIOC10, was analyzed by sodium dodecyl sulfate (SDS)‐polyacrylamide gel (PAGE) and isoelectric focusing (IEF) electrophoresis. Immunoblot analysis of gels after SDS‐PAGE revealed a single 46‐kDa protein in retina. In contrast, two bands of ∼43 and 46 kDa were identified in the pineal gland. Immunoblots of the retinal extracts separated by IEF electrophoresis revealed five S‐antigen isomers, which vary quantitatively throughout the diurnal cycle and when light interrupted the dark cycle. Immunoblots of the pineal gland samples separated by IEF electrophoresis indicated that the pineal gland possesses four pineal gland‐specific forms of S‐antigen in addition to the five forms present in the retina. The differences observed in the mRNA and protein analyses suggest tissue‐specific structural components for S‐antigen in the retina and pineal gland th

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04927.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:3‐Bromopyruvate inhibits pyruvate decarboxylase in brain homogenates and causes a 90% drop in acetylcholine tissue content at a concentration of 2 mM. Stereotaxic injection of 3‐bromopyruvate into the basal forebrain causes after 7 days a 40% drop of acetylcholine concentration and pyruvate decarboxylase activity in the cortex and hippocampus, and greater decreases at the site of injection. However, values return to normal 18 days after injection. Choline ace‐tyltransferase is partially inhibited only at the site of injection after 7 days. Choline transport and choline concentration are not affected at either 7 or 18 days after injection. Impairments in spontaneous alternation and in retention of passive avoidance were seen only 7 days after the injection. The results suggest that stereotaxic injection of bromopyruvate can induce discrete reversible cholinergic lesions on a time scale useful for behavioural experiments and for comparison with neurodegener

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04928.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:We have studied the effect of a dietary deprivation of n‐3 fatty acids on the activity of the dopamine (DA)‐de‐pendent adenylate cyclase in the rat retina. Experiments were conducted in 6‐month‐old rats raised on semipurified diets containing either safflower oil (n‐3 deficient diet) or soybean oil (control diet). The levels of docosahexaenoic acid [22:6 (n‐3)] in retinal phospholipids were significantly decreased in n‐3 deficient rats (35–42% of control levels). This was compensated by a rise in 22:5 (n‐6), the total content of poly‐unsaturated fatty acids (PUFA) remaining approximately constant. Adenylate cyclase activity was measured in retinal membrane preparations from dark‐adapted or light‐exposed rats. The enzyme activity was stimulated by DA and SKF 38393 in a light‐dependent fashion. The activation was lower in rats exposed to light than in dark‐adapted animals, suggesting a down‐regulation of the DI DA receptors by light. The activation by guanine nucleotides and forskolin was also decreased in light‐exposed rats. There was no significant effect of the dietary regimen on the various adenylate cyclase activities and their response to light. Furthermore, the guanine nucleotide‐ and DA‐dependent adenylate cyclase activities of retinal membranes were found to be relatively resistant to changes in membrane fluidity induced in vitro by benzyl alcohol. The results indicate that in the absence of changes in total PUFA content, a decreased ratio of n‐3 to n‐6 fatty acids in membrane phospholipids does not significantly affect the properti

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04929.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:Ganglioside analysis of human motor and sensory nerves revealed that ceramide compositions of sensory nerve GD1a, GD1b, and GMl differed apparently from those in the motor nerve. These gangliosides from sensory nerve contained a large amount of long‐chain fatty acids and d18:1 as a major long chain base. On the contrary, the motor nerve gangliosides contained C16–18fatty acids and a large amount of d20:1 besides d 18:1. Furthermore, these gangliosides were enriched more in the axon fraction than in the myelin fraction. LM1, which was a major ganglioside in myelin from human peripheral nerve, was composed of similar ceramide compositions in the two nerves. The present findings suggest that the characteristic ceramide species of nerve gangliosides may reflect in part properties of their own neur

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04930.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:The monoclonal L5 antibody reacts with anN‐glycosidically linked carbohydrate structure which is present on the neural cell adhesion molecule L1, neural chondroitin sulfate proteoglycans, and other not yet identified glycosylated proteins. Using this antibody, we isolated and characterized proteoglycans from adult mouse brain and cultured astrocytes biosynthetically labeled with Na235SO4and a3H‐amino acid mixture. Our data suggest that the L5 proteoglycans of both sources are identical in their biochemical properties. The apparent molecular mass of the L5 proteoglycan is approximately 500 kDa. Digestion of the iodinated L5 proteoglycan from mouse brain and of the [35S]methionine‐labeled L5 proteoglycan from cultured astrocytes with proteinase‐free chondroitinases ABC and AC revealed three major core proteins with apparent molecular masses of approximately 380, 360, and 260 kDa. These represent molecularly distinct protei

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04931.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:Brain concentrations of salsolinol (SAL), a simple tetrahydroisoquinoline (sTIQ) condensation product of do‐pamine (DA) and acetaldehyde, are reported to increase in chow‐fed rats drinking ethanol/H2O ad libitum. However, our analyses showed that rat chow contains traces of SAL and, as previously reported, appreciable 3,4‐dihydroxyphe‐nylalanine (DOPA), a sTIQ precursor. To examine the effect of consumption of ethanol in a DOPA‐ and SAL‐free diet on endogenous sTIQs, we analyzed two brain regions and blood plasma of rats undergoing prolonged intake (3 weeks and 23 weeks) of liquid diet containing 6.6% ethanol or iso‐caloric carbohydrate. SAL and three other DA‐related sTIQs were quantitated using capillary gas chromatography/mass spectrometry in the selected ion mode with deuterated standards. In accord with studies on ethanol/chow‐fed rats. sTIQ concentrations in hypothalamus were elevated after 3 weeks of ethanol, although after 23 weeks, hypothalamic sTIQs were either unchanged or reduced (O‐methylated SAL). Furthermore, sTIQ concentrations in corpus triatum and, with one exception, plasma were not altered by ethanol ingestion for either duration. (However, 23 weeks of ethanol intake significantly reduced the striatal concentrations of DA and its acid metabolite, presumably reflecting neurotoxicity.) Reasoning that DOPA in diet might underlie the reported ethanol‐dependent increases in striatal sTIQs, we found that l‐DOPA supplementation (500 μg/rat/day) of EtOH/Iiquid diet‐fed rats for 13 weeks tended to increase striatal SAL. Overall, the data indicate that elevations in endogenous sTIQ concentrations due to prolonged ethanol intake depend on the brain region, duration of intake, and even associated dietary constituents. In that regard, the higher striatal SAL concentrations in rats drinking ethanol ad libitum could have been facilitated by DOPA and perhap

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04932.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:The receptor‐ionophore complex of theN‐methyl‐d‐aspartate (NMDA)‐sensitive receptor was solubilized by deoxycholic acid from rat brain using (+)‐[3H]5‐methyl‐10,11‐dihydro‐5H‐dibenzo[a,b]cyclohepten‐5,10‐imine ([3H]MK‐801) binding as a marker for the receptor. Gel filtration of the solubilized preparations on a Sephadex G‐25 column revealed significant [3H]MK‐801 binding sensitive to potentiation by glutamate and glutamate/glycine, which was prevented by competitive antagonists for the NMDA and strychnine‐insensitive glycine (GlyB) sites. In contrast to NMDA and glycine, spermidine markedly potentiated the amount of [3H]MK‐801 binding in solubilized preparations by increasing the apparent affinity of the ligand. In the presence of all three stimulants, the solubilized preparations exhibited pharmacological profiles similar to those in the membrane preparations. These results clearly indicate that the whole macromolecular NMDA receptor‐ionophore complex is solubilized u

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04933.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:The effect of chronic versus acute administration of lithium on receptor‐linked phosphoinositide metabolism was assessed by comparing the change in the cerebral cortex levels of myo‐inositol 1‐phosphate in response to pilocarpine, physostigmine, or pargyline in rats. Rats were exposed to either 29 consecutive days of LiCl injections or 27 and 39 days of dietary Li2CO3, followed by injected LiCl at the end of the diet to insure a constant level of exposure to the drug. In each experiment, an acute group received a single injection of LiCl 20–24 h before they were killed. One hour before being killed, some of the animals acutely exposed to lithium and some of the animals chronically exposed to lithium each received pilocarpine, physostigmine, or pargyline. At the conclusion of the experiment, the rats were killed and brain levels of myo‐inositol 1‐phosphate and lithium were determined. A differential production of myo‐inositol 1‐phosphate in groups receiving acute versus chronic lithium would provide evidence of a change in receptor‐linked phosphoinositide metabolism due to the chronic administration of lithium. Brain levels of mvo‐inositol 1‐phosphate are dependent on tissue lithium concentrations; consequently, significant differences observed in brain lithium levels between the groups receiving acute versus chronic lithium prevented a meaningful assessment of the effect of the mode of lithium administration on the production of myo‐inositol 1‐phosphate in those groups. Stepwise multiple regression analysis and the measured brain lithium levels were used to assess the response of myo‐inositol 1‐phosphate levels to stimulation in animals receiving acute or chronic lithium treatment. The results of the analysis indicate that there would have been no differences in the response to drugs when comparing the two routes of lithium administration if the lithium levels had been identical in both the acute and chronic lithium groups. These results also suggest that multiple regression analysis can be used with an established data base to aid in the interpretation of the results of experiments in which changes in brain myo‐inositol 1‐phosphate levels are used as an index of phosphoinositide metabolism. The possibility is discussed that the effects of lithium on phosphoinositide signalling are already present in the group receiving a single acute dose of LiCl and that this effect persists over the

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04934.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:Specific binding sites with pharmacological properties typical of serotonin 5‐HT3receptors were identified in membranes of the murine hybridoma cell line NG 108‐15, using [3H]zacopride as a ligand. Optimal solubilization of these sites (yield. 50%) could be achieved using the detergent 3–[3–(cholamidopropyl)dimethylammonio]‐1‐propane sul‐fonate (CHAPS) at 24 mMplus0.5 MNaCl in 25 mMTris‐HCl, pH 7.4. Specific [3H]zacopride binding to soluble sites in the 100,000‐gCHAPS extract was saturable and showed characteristics (Bmax= 425 ± 81 fmol/mg of protein;KD= 0.19 ± 0.02 nM) closely related to those of membrane‐bound sites (Bmax= 932 ± 183 fmol/mg of protein;KD= 0.60 ± 0.03 nM). Determination of association (K+1=0.17 nMmin−1) and dissociation (k‐1=0.02 min−1) rate constants for the soluble sites gave aKDvalue of 0.12 nM, a result consistent with that calculated from saturation studies. As assessed from the displacement potencies (IC50) of 10 different drugs, the pharmacological profile of [3H]zacopride specific binding sites was essentially the same (r= 0.99) in the CHAPS‐soluble extract and in cell membranes, although some increase in the affinity for 5‐HT3antagonists (zacopride, ICS 205–930, and MDL 72222) and decrease in the affinity for 5‐HT3agonists (2‐methyl‐5‐hydroxytryptamine and phenylbiguanide) were noted for the soluble sites. Sucrose density gradient sedimentation of the CHAPS‐soluble extract gave a Svedberg coefficient of 12S for the material with [3H]zacopride specific binding capacity. Chromatographic analyses using Sephacryl S‐400 and wheat germ agglutinin‐agarose columns indicated marked enrichment (by 2.5– and 10‐fold, respectively) in [3H]zacopride specific binding activity in the corresponding eluates compared with the starting soluble extract, a finding suggesting that both steps are of potential interest for the partial purification of solubilized 5‐HT3receptors. Two soluble materials with apparent molecular masses of ∼600 and ∼36 kDa were found to bind [3H]zacopride specifically in the Sephacryl S‐400 eluate. Interestingly, molecular mass determination by radiation inactivation of [3H]zacopride bi

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04935.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY

摘要:

Abstract:Modifications of plasma membrane acyl‐linked phospholipid fatty acid composition were produced by supplementing the culture medium with essential fatty acids. The plasma membrane fraction was purified by Percoll gradient centrifugation from dissociated fetal rat brain cells grown in a serum‐free culture medium. Both the concentration dependence and the time course of the modifications were examined. Supplementation of the medium with essential polyunsaturated fatty acid, linolenic acid (18:3ω3) or lin‐oleic acid (18:2ω6), produced incorporation of the elongated and desaturated products of ω3 or ω6 class, respectively, i.e., the incorporation was class specific. Within each class, the most unsaturated and elongated members, i.e., terminal members, were preferentially incorporated until they reached a maximum concentration within 6–7 days. At higher concentrations of supplemented fatty acids, additional class specific incorporation in plasma membrane was produced by an increase in the concentration of intermediate members. At the same time, the concentration of monounsaturated fatty acids declined and that of saturated fatty acids remained unchanged. The modifications in fatty acid composition were reversible, with the time course similar to that of incorporation. The total plasma membrane phospholipid and sterol contents did not change with alterations of fatty acid composition, but did change with time in culture. This preparation should prove useful for investigating the role of polyunsaturated fatty acids in brain cell functions, including neuronal e

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1990.tb04936.x

出版商:Blackwell Publishing Ltd    

年代:1990

数据来源: WILEY