摘要:

AbstractReceptor‐mediated cyclic GMP formation in N1E‐115 murine neuroblastoma cells appears to involve oxidative metabolism of arachidonic acid. Evidence in support of this includes the blockade of this response by lipoxygenase inhibitors, e.g., eicosatetraynoic acid (ETYA) or other metabolic perturbants, e.g., methylene blue. It was recently discovered that the lipoxygenase products 15‐hydroxyeicosatetraenoic (15‐HETE) acid and 12‐HETE, like ETYA, were inhibitors of M1muscarinic receptor‐mediated cyclic GMP formation. In the present report, the effects of monoHETEs are explored in more detail, particularly with regard to the function of the muscarinic receptor. Like 12‐HETE and 15‐HETE (IC50= 13 and 11 μM, respectively), 5‐HETE inhibited the cyclic GMP response to the muscarinic receptor (IC50= 10 μM). All three of these monoHETEs were shown also to be inhibitors of the cyclic GMP responses to receptors stimulated by carbachol, histamine, thrombin, neurotensin, and bradykinin. 15‐HETE was shown to inhibit the muscarinic receptor‐mediated response in a complex manner (apparent noncompetitive and uncompetitive components; IC50= 18 and 2 μM, respectively). 15‐HETE did not inhibit either the M1muscarinic receptor‐stimulated release of [3H]inositol phosphates from cellular phospholipids or the M2muscarinic receptor‐mediated inhibition of hormone (prostaglandin E1)‐induced AMP formation. It seemed possible that the monoHETEs could enter into biochemical pathways for arachidonate in N1E‐115 cells. [3H]Arachidonate and the three [3H]‐monoHETEs all rapidly labeled the membrane lipids of intact NIE‐115 cells, with each [3H]eicosanoid producing a unique labeling profile. [3H] 15‐HETE labeling was noteworthy in that 85% of the label found in the phospholipids was in phosphatidylinositol (PI; t1/2to steady state = 3 min). Exogenous 15‐HETE inhibited the labeling of PI by [3H]‐arachidonate (IC50= 28 μM) and elevated unesterified [3H]‐arachidonate levels. Thus, the mechanism of blockade of receptor‐mediated cyclic GMP responses by monoHETEs is likely to be more complex than the simple inhibition of cytosolic mechanisms, e.g., generation of a putative second messenger by lipoxygenase, and may involve also alterations of membrane function accompany

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02870.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractUsing an affinity‐purified monospecific polyclonal antibody against bovine brain synapsin I, the distribution of antigenically related proteins was investigated in the electric organs of the three strongly electric fishTorpedo marmorata, Electrophorus electricus, Malapterurus electricusand in the rat diaphragm. On application of indirect fluorescein isothiocyanate‐immunofluorescence and using α‐bungarotoxin for identification of synaptic sites, intense and very selective staining of nerve terminals was found in all of these tissues. Immunotransfer blots of tissue homogenates revealed specific bands whose molecular weights are similar to those of synapsin Ia and synapsin Ib. Moreover, synapsin I‐like proteins are still attached to the synaptic vesicles that were isolated in isotonic glycine solution fromTorpedoelectric organ by density gradient centrifugation and chromatography on Sephacryl‐1000. Our results suggest that synapsin I‐like proteins are also associated with cholinergic synaptic vesicles of electric organs and that the electric organ may be an ideal source for studying further the functional and molecular properties o

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02871.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractGlial fibrillary acidic protein (GFAP) was induced in rat C6 glioma cells grown in M199 and HAM F10 media by addition of 1 mMdibutyryl cyclic AMP. The amount of GFAP per cell increased 7‐and 33‐fold in M199 and HAM F10 media, respectively. GFAP could be induced in each phase of the cell culture except for the lag phase, where GFAP synthesis was delayed until the onset of the logarithmic growth. The induction took place under conditions where the total protein content of the cell decreased. Mear surement of the amount of vimentin indicated that GFAP was induced under conditions of low vimentin concentration. Our results do not support the hypothesis that GFAP induction depends on cell‐cell contact or cell proliferation. They indicate a shift from vimentin to GFAP synthesis by an as yet unknown mech

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02872.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractThe effects of some neurotransmitters, adenosine (Ad), and homocysteine (Hcys) on protein carboxyl methylation in synaptic plasma membranes from rat cerebral cortex were examined. Neither any of the neurotransmitters nor Ad had a detectable effect. Incubation of the membrane with DL‐Hcys alone (5 × 10‐5M), the combination of both Ad (5 × 10‐5) and DL‐Hcys (5 × 10‐5), orS‐adenosyl‐L‐homocysteine (SAH) (1 × 10‐6) strongly decreased the methyl ester formation. The inhibitory effect of the combination of both compounds may be interpreted in terms of the increased SAH concentration due to the presence of SAH hydrolase in the membrane. The inhibitory effect of Hcys alone was blocked by preincubation with Ad deaminase or Neplanocin A, a potent inhibitor of SAH hydrolase, suggesting the presence of Ad‐bound SAH hydrolase in the synaptic membrane. Ad‐bound SAH hydrolase activity estimated by the inhibition of methylation in the presence of Hcys was located in the membrane fractions including synaptosomes, myelin, and microsomes (about 70%), but the SAH hydrolase activity estimated on the basis of the inhibitory effect of the combination of both Ad and Hcys was localized exclusively in the soluble fraction (about 90%). The distribution of the latter activity is coincident with that of SAH hydrolase reported to date. Incubation of the synaptic membrane with Hcys alone and with the combination of both Ad and Hcys markedly increased the SAH concentration. The stimulatory effect of Hcys alone wa

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02873.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractThe proteins of lumbar CSF have been investigated by two‐dimensional gel electrophoresis, and their patterns have been compared with the corresponding serum protein patterns. Serum proteins in CSF have been identified by electroblotting and immunoreaction with antiserum against total human serum proteins. Proteins derived from brain have been identified with antiserum against human brain proteins. The most prominent CSF protein group has been identified as a multiple form of apolipoprotein E. The correct position of the glial fibrillary acidic protein has also been determined. The prefractionation of CSF proteins by size exclusion chromatography or by affinity chromatography followed by two‐dimensional electrophoresis has facilitated the detection of trace components in CSF and the corresponding se

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02874.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractThe synthesis of a tritiated derivative of the 5‐HT1Aphotoaffinity probe 8‐methoxy‐2‐[N‐n‐propyl,N‐3‐(2‐nitro‐4‐azidophenyl)aminopropyl]aminotetralin ([3H]8‐methoxy‐3′‐NAP‐amino‐PAT) allowed the use of this probe for attempting the irreversible labeling of specific binding sites in rat brain membranes. Sodium dodecyl‐sulfate‐polyacrylamide gel electrophoresis of proteins solubilized from hippocampal microsomal membranes that had been incubated with 20 nM[3H]8‐methoxy‐3′‐NAP‐amino‐PAT under UV light revealed a marked incorporation of3H label into a 63‐kilodalton protein termed PI. As expected of a possible correspondence between P1and 5‐HT1Areceptor binding sites,3H labeling by the photoaffinity probe could be prevented by selective 5‐HT1Aligands such as 8‐hydroxy‐2‐(di‐n‐propylamino)tetralin, ipsapirone, buspirone, and gepirone and by N‐ethylmaleimide, but not by the 5‐HT2antagonist ketanserin, noradrenaline‐and dopamine‐related drugs, monoamine oxidase inhibitors, and chlorimipramine. Furthermore, the regional and subcellular distributions of P1were identical to those of specific 5‐HT1Abinding sites. These results indicated that the binding subunit of the 5‐HT1Areceptor

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02875.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractSeveral ions commonly used as substitutes for Na+or Cl‐were found to inhibit directly the high‐affinity uptake of norepinephrine, dopamine, serotonin, and γ‐aminobutyric acid, but not glutamate or glutamine. When Na+was partially replaced by any of several different cations or sucrose the uptake of all neurotransmitters studied except that of serotonin was reduced more than could be accounted for by just the inhibitory effect of the cation substitute. In contrast, when Cl was partially replaced by any of several anions only the uptake of dopamine was reduced more than could be accounted for by the inhibitory effect of the anion substitute. These results suggest that for most neurotransmitters the electrochemical potential for Na+, but not for Cl, contributes to the uptake driving force. When either Na+or Cl wastotallyreplaced by an ion substitute or by sucrose the high‐affinity uptake was virtually abolished, an exception being that glutamate uptake was not affected when isethionate was substituted for Cl. The lack of uptake in the absence of either Na+or Cl may reflect a specific role for these ions in either increasing the affinity between the substrate and the carrier, or facilitating the translocation process. Alternatively, the transport carriers may undergo a nonspecific conformational change to an inactive form in the absence of Na+or Cl. A partial substitution of Na+with Li+or sucrose differentially affected the kinetics of uptake in that replacement with Li+, but not sucrose, usually resulted in a marked increase in theKmvalues. The results of this study emphasize the importance of taking into consideration the effects of ions used as substitutes in experiments undertaken to elucidate the roles of Na+and Cl in the high‐affinity uptake of monoamine and amino acid neurotr

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02876.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractCerebral cortical slices from rat brain were incubated at 37°C for 2 h in the presence of isoproterenol, noradrenaline, or adrenaline, and binding affinities and densities of adrenoceptor subtypes were subsequently examined in homogenized tissue. The density of α2‐and total β‐adre‐noceptors was estimated using the radioligands [3H]rauwolscine and [3H]dihydroalprenolol (DHA), respectively. The percentages of β1‐and β2‐adrenoceptors were denned by inhibiting the binding of [3H]DHA with the β1‐selective antagonist metoprolol. Exposure of slices to noradrenaline and adrenaline significantly decreased the maximal number of binding sites(Bmax) of α2‐adrenoceptors (48 and 37% respectively) without significantly affecting affinity; isoproterenol had no effect. Exposure to isoproterenol, noradrena line, and adrenaline significantly decreased the Bmaxof β‐adrenoceptors (by 60, 34, and 24%, respectively) but did not affect the affinity. Isoproterenol and adrenaline significantly decreased the density of β1‐adrenoceptors by 75 and 24% and β2‐radrenoceptors by 23 and 28%, respectively. Noradrenaline significantly decreased the density of β1‐adreno‐ceptors by 42% without affecting the number of β2‐adreno‐ceptors. These findings indicate that subtypes of adrenoceptors in rat cerebral cortex are differen

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02877.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractDorsal root ganglia, excised from the lumbar roots of the sciatic nerve of white Leghorn chicken embryos 6–13 days of age, were incubated usually for 5 h, at 36°C in 20 μI of a bicarbonate‐buffered physiological salt solution containing 5.5 mMglucose. [U‐14C]Glucose, [1‐14C]glucose, [6‐14C]glucose, or [5‐3H]uridine was also added. Lipid synthesis and lactate output were measured by incorporation of14C from [U‐14C]glucose and RNA synthesis by incorporation of3H from [5‐3H]uridine. Glucose uptake and labeled lactate output declined rapidly from 6 to 8–9 days of age, more slowly thereafter. Synthesis of lipids was relatively constant throughout the ages studied, without the increased rate at intermediate ages seen previously in sympathetic ganglia of the same species. RNA synthesis declined progressively throughout the ages studied. The output of C‐6 of glucose to CO2was about the same at all ages, whereas that of C‐1 declined rapidly from 6 to 7 days of age and then more slowly, but always remained higher than that of C‐6 and thus indicated that much glucose was metabolized via the

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02878.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY

摘要:

AbstractThe neural cell adhesion molecule (N‐CAM) is a complex of surface glycoproteins that are developmentally regulated and believed to be intimately involved in the orderly structuring of the CNS. Here the effect of chronic low‐level lead exposure on their expression in the postnatally developing cerebellum is described. Rat pups were chronically exposed to lead via their dam's drinking water which contained either 200 or 400 mg PbCl2/L from time of birth. Pup postnatal blood lead levels ranged between 10 and 20 μg/dl until day 16 after which they became elevated to 40 μg/dl on day 20. During this period the developmental sialylation state of N‐CAM, which is believed to regulate cell‐cell interaction, fibre outgrowth, and synapse formation, was monitored by rocket immunoelectrophoresis. In control animals the expected desialylation of N‐CAM was found to occur at times coincident with postnatal synapto‐genesis. In contrast, desialylation in animals chronically exposed to lead was found to be significantly impaired when blood lead levels exceeded 20 μg/dl. This could not be attributed to lead‐induced undernutrition or alterations in immu‐noprecipitate formation. These observations could account for the neurobehavioural deficits that are known to be induced at similar blood lead levels, and the potential contribution of impaired N‐CAM desialylation to synaptic elab

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1987.tb02879.x

出版商:Blackwell Publishing Ltd    

年代:1987

数据来源: WILEY