摘要:

AbstractCadmium (109Cd2+) uptake was studied in a preparation of isolated neurohypophysial nerve terminals. By use of a filter separation method, together with a permeabilizing agent (Triton X‐100), two cellular Cd2+pools have been distinguished. The uptake into the intraterminal pool was governed mainly by a process that displayed saturable kinetics, with aVmaxof 0.15 nmol of Cd2+/mg of protein /min and aKmof 0.18 mM, consistent with a transport system. The superficially bound Cd2+pool (Triton insensitive), which represented 30–50% of the total Cd2+bound to the cellular system, was very sensitive to the ionic composition of the incubation medium. Reducing the extracellular Ca2+or Na+concentration caused a significant increase in the size of the Triton‐insensitive Cd2+pool. Whereas Na+did not affect Cd2+uptake, Ca2+induced a small, but significant, increase of Cd2+uptake into the terminals. It is concluded that there is a significant intraterminal uptake of Cd2+, which could explain several physiological effects of thi

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06391.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractTreatment of bovine chromaffin cells with insulin‐like growth factor‐I (IGF‐I) caused the activation of a protein kinase that phosphorylates microtubule‐associated protein‐2 (MAP‐2) in vitro. Activation of MAP‐2 kinase by IGF‐I varied with the time of treatment (maximal at 10–15 min) and the concentration of IGF‐I (maximal at 10 nM). The IGF‐I‐activated MAP‐2 kinase was localized to the soluble fraction of chromaffin cell extracts and required Mg2+for activity. The IGF‐I‐activated kinase also phosphorylated myelin basic protein, but had little or no activity toward his‐tones or ribosomal S6 protein. To examine the role of protein tyrosine phosphoryiation in the activation of the MAP‐2 kinase, we isolated phosphotyrosine (PTyr)‐containing proteins from chromaffin cells by immunoaffinity adsorption on anti‐PTyr‐Sepharose beads. Anti‐PTyr‐Sepharose eluates from IGF‐I‐treated cells showed increased MAP‐2 kinase activity; thus, the MAP‐2 kinase (or a closely associated protein) appears to be a PTyr‐containing protein. Treatment of anti‐PTyr‐Sepharose eluates or crude chromaffin cell extracts with alkaline phosphatase significantly decreased kinase activity toward myelin basic protein, indicating that phosphoryiation o

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06392.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractAge‐dependent changes in the expression of neuropeptide Y (NPY) peptides and prepro‐NPY mRNA (NPY mRNA) were studied in rat adrenal gland and brain areas by means of radioimmunoassay, immunohistochemistry, and northern blot analysis. In the adrenal gland, NPY immunoreactívity (NPY‐I) increased by 80‐fold, mainly in the chromaffin cells, during aging (from 7 to 33 weeks old). The increase in NPY‐I was accompanied by a concomitant increase in the content of NPY mRNA (800 bases in size, by 16‐fold) and putative NPY pre‐mRNA, a result suggesting that this increase results from that in NPY gene expression, probably at the level of transcription. In contrast, in some brain areas, such as striatum and medulla oblongata plus pons, NPY‐I decreased in an age‐dependent manner, whereas NPY mRNA abundances in these areas increased by twofold with age (from 7 to 33 weeks old). The opposite changes between NPY and NPY mRNA content in specific brain areas suggested the accelerated turnover/degradation of NPY peptide in the brain areas. Furthermore, β‐actin mRNA abundance did not change in rat adrenal gland and brain areas during aging. Thus, the characteristic age‐related increase in NPY gene expression in rat adrenal gland and some brain areas seems to be important for physiological regulation of some neuronal functions, such as blood pr

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06393.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractChanges in transglutaminase (TG) activity in superior cervical ganglia (SCG) and nodose ganglia (NG) excised from adult rats were examined following application of selected membrane transport‐altering agents, including GM1‐ganglioside (GM1) and α‐sialylcholesterol (α‐SC). Although TG activity of freshly dissected SCG and NG was relatively low, it increased gradually during 30 min of incubation, and it stayed at this elevated level for 2 h. Addition of α‐SC at its maximal effective concentration, 20 μM, stimulated TG activity more than eightfold in SCG and more than twofold in NG by 30 min. Addition of GM1 at its most effective concentration, 5 nM, had similar effects, but of lesser magnitude. Cydoheximide, a potent inhibitor of protein biosynthesis, did not affect the GM1‐or α‐SC‐evoked increases in ganglionic TG activity, suggesting that enzyme activation rather than synthesis of new enzyme was occurring. The stimulation of TG activity in both ganglia caused by either GM1 or α‐SC was associated with a decrease inKmand an increase inVmaxvalues. Addition of cholera toxin B. which specifically masks the oligosaccharide chain of GM1. reduced the GM1‐induced increase in TG activity by approximately 60% in SCG and 88% in NG. The α‐SC‐induced increase in TG activity was only partially mimicked by free cholesterol. Although application of either dibutyryl cyclic AMP or dibutyryl cyclic GMP produced little change in TG activity of either ganglion, phorbol ester clearly inhibited the enzymic activity. Because TG is a calcium‐dependent enzyme, we measured45Ca2+influx into either ganglion, and found that it was reduced by GM1 and a‐SC in SCG and by α‐SC in NG. The GM1‐induced decrease in45Ca2+influx was prevented by cholera toxin B in SCG, whereas the α‐SC‐induced decrease was not mimicked by cholesterol in either ganglion. These results suggest that the sialic acid moiety of GM1 and α‐SC is responsible for the activation of ganglionic TG and that activation may occur via a protein kinase C‐r

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06394.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractThis study demonstrates the presence of both atrial natriuretic peptide (ANP) precursor and ANP transcripts in the rat olfactory bulb (OB), a key brain structure involved in the generation of olfaction‐dependent behavior. In addition to synthesizing ANP, the OB contains ANP‐transducing receptors coupled to the guanylate cyclase system but it is devoid of ANP “clearance receptors.” The characterization of biologically active ANP receptors and the evidence for in situ ANP synthesis in this region of the CNS adds credence to the hypothesis that the peptide plays a putative role in ol

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06395.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

Abstract:Neuronal stimulation can rapidly activate several immediate early genes that code for transcription factors. We have used primary cortical cultures to study the regulation of four of these genes, c‐fos, c‐jun, jun‐B, andzif268.Im‐munocytochemical studies with antibodies to Jun‐B, c‐Jun, and c‐Fos demonstrate intense staining in the nuclei of a subset of cortical neurons in mature cultures (21–25 days in vitro) but not young cultures (3–7 days in vitro). To assess whether this immunoreactivity may be induced by spontaneous synaptic activity that develops with a similar profile, we examined the effects of agents that reduce this synaptic activity. Tetrodotoxin orN‐methyl‐d‐aspartate receptor antagonists suppress basal immunoreactivity to Jun‐B and c‐Fos, but not c‐Jun, indicating that the basal level of c‐Jun expression is not dependent on electrical activity. Pierotoxin, an agent that increases synaptic excitation indirectly by blocking inhibitory synaptic currents mediated by γ‐aminobutyric acidAreceptors, markedly increases the percentage of neurons displaying immunoreactivity to c‐Fos, c‐Jun, Jun‐B, and Zif268. Northern analysis suggests that the increases in immunostaining induced by picrotoxin are secondary to a rapid increase in mRNA for these proteins. These findings provide evidence for rapid transcriptional regulation of immediate early genes

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06396.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractMuscarinic acetylcholine receptors (mAChRs) share with many other receptors of the guanine nucleotide‐binding protein‐coupled receptor family a highly conserved cysteine residue in the putative cytoplasmic carboxyl‐terminal region of the protein. Because elimination of this cysteine in the β2‐adrenergic receptor has been reported to decrease functional responsiveness, we determined if this cysteine residue is essential for mAChR‐effector coupling by replacing Cys457of the m2mAChR with glycine and expressing wild‐type and mutant receptor in Chinese hamster ovary (CHO) cells. The mutant and wild‐type receptors exhibited similar affinities for binding of muscarinic ligands. In addition, the mutation did not affect cell surface localization or receptor‐mediated inhibition of adenylate cyclase. These results indicate that the cysteine residue in the carboxyl‐terminal domain of the m2mAChR is not required for ligand binding or mAChR‐mediated inhibition of adenylate

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06397.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractExperiments examined the effects of peripheral and central administration of the vesicular acetylcholine transport blocker vesamicol (AH5183) on the content, synthesis, and release of acetylcholine in the rat brain in vivo. In time course studies, a single intraperitoneal dose ofdl‐vesamicol (5 mg/kg) rapidly and reversibly (within 2 h) doubled the content of acetylcholine in the striatum and hippocampus, without affecting choline levels or the rate of transmitter synthesis. In microdialysis experiments, the same peripheral dose of drug produced a reversible 55% reduction in endogenous striatal acetylcholine release. A similar inhibitory effect was produced by direct intrastriatal perfusion with vesamicol. Moreover, this effect of vesamicol was (a) concentration‐dependent and saturable (EC50= 68 nM), (b) rapidly reversible, (c) stereospecific for thel‐isomer, and (d) poorly mimicked by a vesamicol analog with lower plasma membrane permeability. This profile of effects is consistent with an interaction with a specific vesamicol receptor as defined by previous in vitro binding studies. These results support a functional role for vesamicol receptors in modulating central cholinergic transmission in

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06398.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractThe effects of ryanodine, a selective inhibitor of the Ca2+‐induced Ca2+release mechanism, on caffeine‐evoked changes in cytosolic Ca2+concentration ([Ca2+]i) and cate‐cholamine secretion were investigated using cultured bovine adrenal chromaffin cells. Caffeine (5–40 mM) caused a concentration‐dependent transient rise in [Ca2+]iand catecholamine secretion in Ca2+/Mg2+‐free medium containing 0.2 mM EGTA. Ryanodine (5 × 10–5M) alone had no effect on either [Ca2+]ior catecholamine secretion. Although the application of ryanodine plus caffeine caused the same increase in both [Ca2+]iand catecholamine secretion as those induced by caffeine alone, ryanodine (4 × 10–7–5 × 10–5M) irreversibly prevented the increase in both [Ca2+]iand catecholamine secretion resulting from subsequent caffeine application over a range of concentrations. The secretory response to caffeine was markedly enhanced by replacement of Na+with sucrose in Ca2/Mg2+‐free medium, and this enhanced response was also blocked by ryanodine. Caffeine was found to decrease the susceptibility of the secretory apparatus to Ca2+in digitonin‐permeabilized cells. These results indicate that caffeine mobilizes Ca2+from intracellular stores, the function of which is irreversibly blocked by ryanodine, resulting in the increase in catecholamine secretion in the bovi

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06399.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY

摘要:

AbstractInclusion of 1.1% elemental tellurium in the diet of postweanling rats produces a peripheral neuropathy due to a highly synchronous primary demyelination of sciatic nerve; this demyelination is followed closely by remyelination. Sciatic nerves from animals fed tellurium for various times were removed and incubated ex vivo for 1 h with [14C]acetate, and radioactivity incorporated into individual lipid classes was determined. In nerves from rats exposed to tellurium, there was a profound and selective block in the conversion of radioactive acetate to cholesterol. Another radioactive precursor, [3H]water, gave similar results. We suggest that tellurium feeding inhibits squalene epoxidase activity and that the consequent lack of cholesterol destabilizes myelin, thereby causing destruction of the larger internodes. Ex vivo incubation experiments were also carried out with liver slices. As with nerve, tellurium feeding caused accumulation in squalene of label from radioactive acetate, whereas labeling of cholesterol was greatly inhibited. Unexpectedly, however, incorporation of label from [3H]water into both squalene and cholesterol was increased. Relevant is the demonstration that liver was the primary site of bulk accumulation of squalene, which accounted for 10% of liver dry weight at 5 days. Thus, accumulation of squalene (and other mechanisms, possibly including up‐regulation of cholesterol biosynthetic pathways) drives squalene epoxidase activity at normal levels in liver even in the presence of inhibitors of this enzyme. This is reflected by continuing incorporation of [3H]water into cholesterol; incorporation of this precursor takes place at many of the postsqualene biosynthetic steps for sterol formation. [14C]Acetate entering the sterol pathway before squalene in liver is greatly diluted in specific activity when it reaches the large squalene pool, and thus increased squalene epoxidase activity does not transfer significant14C label to sterols. In contrast to the situation with liver, synthesis of sterols is markedly depressed in sciatic nerve, and squalene does not accumulate to high level

ISSN:0022-3042    

DOI:10.1111/j.1471-4159.1991.tb06400.x

出版商:Blackwell Publishing Ltd    

年代:1991

数据来源: WILEY