共查询到20条相似文献,搜索用时 15 毫秒
1.
Mutually Independent Cyclic AMP and Sodium Responses to Nerve Growth Factor in Embryonic Chick Dorsal Root Ganglia 总被引:1,自引:3,他引:1
Abstract: Chick embryo dorsal root ganglia display a rapid and transient rise in their cyclic AMP content when presented with nerve growth factor. These ganglia also depend on nerve growth factor for control of their intracellular Na+ and K+ levels. A sequential relationship between the cyclic AMP and Na+ responses is not readily apparent. Incubation of chick sensory ganglia in a sodium-free medium does not prevent the cyclic AMP response to nerve growth factor from occurring. When ganglia are first incubated with ouabain for 6 h, presentation of nerve growth factor elicits a cyclic AMP response, but no Na+ response. The cyclic AMP response therefore does not depend on the Na+ environment. An initial presentation of nerve growth factor to the ganglia for 30 min, followed by its withdrawal and subsequent re-administration at different intervals over several hours failed to result in a second cyclic AMP response. Nevertheless, the expected Na+ behaviors were still observed. Dibutyryl cyclic AMP is capable of eliciting a cyclic AMP response in chick sensory ganglia after 6 h of nerve growth factor deprivation. When both agents were presented simultaneously to the ganglia, only a single cyclic AMP response was obtained, corresponding in time to the response elicited by dibutyryl cyclic AMP alone-indicating that this drug acts on the NGF-sensitive cells. At the same time dibutyryl cyclic AMP alone failed to result in a Na+ response, leading one to conclude that the cyclic AMP response to nerve growth factor is truly not mediating the Na+ response. Additional support for the mutual independence of these two short-latency responses is provided by the apparent inability of nerve growth factor to cause a cyclic AMP response in chick embryo sympathetic ganglia, another traditional target for the factor, which is capable of displaying a Na+ response. 相似文献
2.
Robert W. Stach C. Richard Lyons J. Regino Perez-Polo 《Journal of neurochemistry》1987,49(4):1280-1285
Receptors for the nerve growth factor protein (NGF) have been isolated from three cell types [embryonic chicken sensory neurons (dorsal root sensory ganglia; DRG), rat pheochromocytoma (PC12) and human neuroblastoma (LAN-1) cells] and have been shown to be similar with respect to equilibrium dissociation constants. The present results demonstrate that there are multiple molecular weight species for NGF receptors from DRG neurons and PC12 cells. NGF receptors can be isolated from DRG as four different molecular species of 228, 187, 125, and 112 kilodaltons, and PC12 cells as three molecular species of 203, 118, and 107 kilodaltons. The NGF receptors isolated from DRG show different pH-binding profiles for high- and low-affinity binding. High-affinity binding displays a bell-shaped pH profile with maximum binding between pH 7.0 and 7.9, whereas low-affinity binding is constant between pH 5.0 and 9.1, with a twofold greater binding at pH 3.6. At 22 degrees C, the association rate constant was found to be 9.5 +/- 1.0 X 10(6) M-1 s-1. Two dissociation rate constants were observed. The fast dissociating receptor has a dissociation rate constant of 3.0 +/- 1.5 X 10(-2) s-1, whereas the slow dissociating receptor constant was 2.4 +/- 1.0 X 10(-4) s-1. The equilibrium dissociation constants calculated from the ratio of dissociation to association rate constants are 2.5 X 109-11) M for the high-affinity receptor (type I) and 3.2 X 10(-9) M for the low-affinity receptor (type II). These values are the same as those determined by equilibrium experiments on the isolated receptors.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
3.
Nerve Growth Factor Stimulates Phospholipid Methylation in Target Ganglionic Neurons Independently of the Cyclic AMP and Sodium Pump Responses 总被引:2,自引:2,他引:0
Suspensions of neurons prepared from embryonic day 12 (E12) chick sympathetic ganglia were incubated with [methyl-3H]methionine in the absence of nerve growth factor (NGF). Presentation of the factor for different periods of time resulted in an approximate three-fold stimulation of radioactivity incorporated into total phospholipid, followed by a rapid decline thereafter. Both the magnitude and the time of the response were dependent on the NGF concentration used. Also examined were possible relationships of phospholipid methylation to two other short-latency responses to NGF, i.e., control of the Na+,K+-pump and elevation of cyclic AMP content. Incubation of E12 sympathetic neurons with known transmethylase inhibitors (shown to be active in the present system) failed to prevent reactivation of the Na+,K+-pump in response to NGF administration. E16 sympathetic neurons and E15 sensory neurons, which do not depend on exogenous NGF for control of their Na+,K+-pump, still show a stimulation of phospholipid methylation when challenged with the factor. Blockage of the pump with ouabain also fails to prevent a methylation response. Thus, the pump and methylation responses to NGF occur independently of each other. Intact E8 chick dorsal root ganglia, but not E12 sympathetic ganglia, display a rapid and transient rise in their cyclic AMP content when presented with NGF. At a concentration of 10 biological units/ml, NGF elicits a peak of phospholipid methylation at 4 min, and a peak of cyclic AMP at 10 min. Methylation inhibitors prevent the methylation response, but not that of cyclic AMP.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
4.
Suramin Induces Phosphorylation of the High-Affinity Nerve Growth Factor Receptor in PC12 Cells and Dorsal Root Ganglion Neurons 总被引:2,自引:0,他引:2
Jagjit S. Gill Denise C. Connolly †Michael J. McManus Nita J. Maihle ‡ Anthony J. Windebank 《Journal of neurochemistry》1996,66(3):963-972
Abstract: Suramin is a polysulfonated naphthylurea with demonstrated antineoplastic activity. Toxicity includes adrenal insufficiency and peripheral neuropathy. Although the mechanism of antitumor activity is unknown, inhibition of binding of growth factors to their receptors has been suggested. Growth factors inhibited by suramin include platelet-derived growth factor, fibroblast growth factor, transforming growth factor, epidermal growth factor, insulin-like growth factor, and nerve growth factor (NGF). In these studies, suramin was shown to be cytotoxic to PC12 cells in a dose-dependent manner. At lower doses and in surviving cells, we observed the induction of neurite outgrowth. To determine the mechanism of suramin-induced neurite outgrowth, PC12 cells were exposed to suramin and/or NGF for various time periods and treated cells were analyzed, by western blot analysis, for expression of tyrosine phosphoproteins. There was a similarity in the pattern of tyrosine-phosphorylated proteins in PC12 cells stimulated with suramin or NGF. Of particular interest was the rapid phosphorylation (by 1 min) of the high-affinity NGF (TrkA) receptor. Activation of other members of the signal-transduction cascade (Shc, p21 ras , Raf-1, ERK-1) revealed similar phosphorylation levels induced by suramin and NGF. Parallel studies were performed in rat dorsal root ganglion cultures; suramin potentiated neurite outgrowth and activated the NGF receptor on these cells. This finding of specific patterns of tyrosine phosphorylation of cellular proteins in response to suramin treatment demonstrated that suramin is a partial agonist for the NGF receptor in both PC12 cells and dorsal root ganglion neurons. 相似文献
5.
Abstract: We have recently shown that the small GTP binding protein p21 ras is essential for nerve growth factor (NGF)-mediated survival of peripheral embryonic chick dorsal root ganglia (DRG) sensory but not sympathetic neurons. To investigate at which level of the signaling cascade the pathways diverge, we have studied the time-resolved pattern of NGF-stimulated tyrosine phosphorylation of proteins within 4 h after addition of the neurotrophin. In both chick sympathetic neurons [embryonic day (E) 12] and DRG sensory neurons (E9) NGF induces within 1 min the autophosphorylation of the receptor tyrosine kinase p140trk. However, the pattern of substrate protein tyrosine phosphorylation downstream of p140trk is distinctly different in both neuronal subtypes. In sympathetic neurons, we observe within 1 min the tyrosine phosphorylation of a new substrate protein, p105, reaching maximal levels at 3 min. Tyrosine phosphorylation of p105 remains elevated for up to 4 h. Subsequent to p105, NGF induces the tyrosine phosphorylation of p42, a protein belonging to the family of mitogen-activated protein (MAP) kinases. This stimulation is transient, reaching maximal levels at 10 min and returning to very low levels already after 2 h. In DRG sensory neurons, tyrosine phosphorylation of p105 is weak and very short lived, disappearing already after treatment with NGF for 10 min. In contrast, activation of MAP kinase p42 in DRG sensory neurons is more stable than in sympathetic neurons. All NGF-stimulated tyrosine phosphorylation events were inhibited by preincubation of neurons with the tropomyosin-related kinase (trk) inhibitor K252a. We suggest the working hypothesis that persistent tyrosine phosphorylation of p105 may play a role in the p21ras-independent NGF survival pathway of chick sympathetic neurons. 相似文献
6.
目的:检测脊神经切断大鼠背根节(DRG)神经元重复放电能力和钠电流的变化,并研究介导其电流变化的钠通道亚型的表达情况。方法:脊神经切断术后2~8d慢性痛大鼠模型背根节急性分离,对中等直径DRG神经元运用全细胞膜片钳技术记录神经元放电和钠电流的变化。对背根节神经元进行RT-PCR检测,分析其钠通道亚型的表达情况。结果:电流钳下,实验组DRG神经元在电流刺激下产生重复放电,而对照组神经元多诱发单个动作电位,电压钳记录发现实验组背根节神经元快钠电流和持续性钠电流幅值均明显大于对照组,PCR结果显示,Nav1.3、Nav1.7和Nav1.8通道亚型mRNA表达显著增高。结论:钠通道介导了脊神经受损模型的DRG神经元兴奋性增高,持续性钠电流可能通过调节阈下膜电位振荡的产生调节神经元兴奋性。 相似文献
7.
目的:检测脊神经切断大鼠背根节(DRG)神经元重复放电能力和钠电流的变化,并研究介导其电流变化的钠通道亚型的表达情况。方法:脊神经切断术后2~8d慢性痛大鼠模型背根节急性分离,对中等直径DRG神经元运用全细胞膜片钳技术记录神经元放电和钠电流的变化。对背根节神经元进行RT-PCR检测,分析其钠通道亚型的表达情况。结果:电流钳下,实验组DRG神经元在电流刺激下产生重复放电,而对照组神经元多诱发单个动作电位,电压钳记录发现实验组背根节神经元快钠电流和持续性钠电流幅值均明显大于对照组,PCR结果显示,Nav1.3、Nav1.7和Nav1.8通道亚型mRNA表达显著增高。结论:钠通道介导了脊神经受损模型的DRG神经元兴奋性增高,持续性钠电流可能通过调节阈下膜电位振荡的产生调节神经元兴奋性。 相似文献
8.
During development, neural crest-derived sensory neurons require nerve growth factor (NGF) for survival, but lose this dependency postnatally. Similarly, dissociated embryonic sensory neurons lose their NGF dependence during the first 3 weeks in cell culture. It has been hypothesized that, in sympathetic neurons, intracellular levels of calcium are related to trophic factor dependence. In vitro during the period in which embryonic-day-15 sensory neurons become independent of NGF, intracellular calcium concentrations progressively increased in parallel to the decline in NGF dependence. This elevation of intracellular calcium was directly related to the absolute age of the neurons, not to the length of time in culture. Without NGF, immature sensory, i.e., dependent, neurons survived in the presence of high extracellular potassium, a condition that produces elevated intracellular calcium. In another paradigm, measurements of intracellular calcium were determined in NGF-dependent neurons "committed to die" after NGF withdrawal. These measurements were determined prior to the time that extensive morphological changes, consistent with cell death, were noted by phase-contrast microscopy. No elevation in intracellular calcium was found in these dying neurons, but rather, a small decrease was observed prior to the disintegration of the neurons. These findings support the hypothesis that trophic factor dependence of neurons may be inversely related to levels of intracellular calcium. 相似文献
9.
激活素促进鸡胚神经节神经突起生长作用 总被引:4,自引:0,他引:4
为了探讨激活素(activin)促进鸡胚背根神经节(dorsal root ganglia,DRG)突起生长、维持神经节细胞生存作用及其与一氧化氮(NO)释放的关系,实验采用8 d的鸡胚分离背根神经节,原代培养法,观察鸡胚背根神经节的体外生长情况。研究结果表明,添加激活素A培养的背根神经节有明显的神经突起生长,形成密集的网络,背根神经节可存活8~10 d;而阴性对照组几乎无神经突起生长,背根神经节可存活3~4 d。添加激活素A的背根神经节单层培养神经节细胞也可长期存活;而阴性对照组在培养第5 d几乎无神经节细胞生存。NO检测结果显示,添加激活素A培养的背根神经节上清NO分泌水平明显降低,与阴性对照组比较差异显著(P<0.05);激活素A与神经生长因子(nerve growth factor,NGF)具有协同抑制背根神经节NO分泌作用。激活素结合蛋白(follistatin)明显抑制激活素A诱导的背根神经节神经突起生长。研究结果提示,激活素可维持鸡胚神经节细胞存活并刺激神经突起生长,其作用与抑制神经损伤因子NO的释放有关。 相似文献
10.
James R. Munis Joseph T. Steiner Martial Ruat Solomon H. Snyder 《Journal of neurochemistry》1998,70(3):1323-1326
Abstract: The enzyme diamine oxidase (DAO) catalyzes the oxidative deamination of histamine, diamines, and polyamines. DAO has been localized to several tissues, including thymus, kidney, intestine, seminal vesicles, placenta, and pregnancy plasma. DAO is not constitutively expressed in the mammalian brain, but it becomes detectable following focal injury. Although the physiologic role of DAO remains unknown, the observation that it is present at the interface between rapidly dividing and quiescent cells in several tissues suggests that it might be involved in regulating cell division or differentiation at tissue boundaries. In addition, the observation that DAO is expressed in the brain following injury suggests that the protein might play a role in the CNS response to focal neuronal damage. To test that hypothesis, we assessed the ability of purified DAO to alter the pattern of neuronal differentiation and nerve growth in vitro. In chick dorsal root ganglion explant cultures, purified porcine DAO induced neurite outgrowth in the low nanomolar range. Addition of aminoguanidine, which inhibits DAO enzyme activity, did not inhibit the protein's neurotrophic activity. These findings suggest that DAO can function as a neurotrophic ligand independent of its enzymatic activity. 相似文献
11.
Nerve Growth Factor Receptors in Human Neuroblastoma Cells 总被引:2,自引:2,他引:2
Receptors for the nerve growth factor protein (NGFR) present in the human neuroblastoma cell line LAN-1 were characterized. LAN-1 cells display high-affinity (type I, with KD value of 5.9 X 10(-11) M) and low-affinity (type II, with KD value of 9.2 X 10(-9) M) binding to NGF. NGFR were fractionated by preparative isoelectric focusing in a granulated gel (PEGG). High-affinity binding was found in the 5.9-6.2 pH region of the PEGG, and low-affinity binding in the 4.6-4.8 and 8.8-9.3 pH ranges. After further analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) we observed both 92.5- and 200-kDa molecular species associated with NGF binding activity. The 200-kDa protein was found in fractions displaying high-affinity NGF binding and the 92.5-kDa protein in fractions displaying low-affinity NGF binding. Equilibrium binding analysis of NGF in PEGG fractions confirmed the presence of two specific saturable binding sites with KD values similar to those observed for whole dissociated cells. When NGFR II activity from the acidic region of the PEGG chromatogram was incubated with NGFR II from the basic region of the PEGG chromatogram, there was no change in NGF binding or in the number of apparent NGF receptors. However, incubation of these same fractions with a fraction having only NGFR I showed an apparent increase in high-affinity NGF binding and a decrease in low-affinity NGF binding. Immunoprecipitation of this "mixed" fraction and analysis on SDS-PAGE under reduced and nonreduced conditions showed 200-kDa and 92.5-kDa proteins under nonreduced conditions and a 92.5-kDa protein under reduced conditions. Our findings are consistent with the hypothesis that there are two distinct NGF receptors in NGF-responsive cells. The interconvertibility of low- and high-affinity receptors and the possible existence of a modulator type protein or of "silent" type receptors are also in agreement with our findings. 相似文献
12.
Acute Regulation of the Epidermal Growth Factor Receptor in Response to Nerve Growth Factor 总被引:2,自引:0,他引:2
PC12 cells possess specific receptors for both nerve growth factor and epidermal growth factor, and by an unknown mechanism, nerve growth factor is able to attenuate the propagation of a mitogenic response to epidermal growth factor. The differentiation response of PC12 cells to nerve growth factor, therefore, predominates over the proliferative response to epidermal growth factor. We have observed that the addition of nerve growth factor to PC12 cells rapidly produces a decrease in surface 125I-epidermal growth factor binding capacity. Unlike previously described nerve growth factor effects on 125I-epidermal growth factor binding capacity, which required several days of nerve growth factor exposure, the decreases we report occur within minutes of nerve growth factor addition: A 50% decrease in 125I-epidermal growth factor binding capacity is evident at 10 min. This rapid nerve growth factor response is concentration dependent; inhibition of 125I-epidermal growth factor binding is detectable at nerve growth factor levels as low as 0.2 ng/ml and is maximal at approximately 50 ng/ml, consistent with known ranges of biological activity. No demonstrable differences in the rate of epidermal growth factor receptor synthesis or degradation were observed in cells acutely exposed to nerve growth factor. Scatchard analysis revealed that acute nerve growth factor treatment decreased the number of both high- and low-affinity 125I-epidermal growth factor binding sites, while the receptor affinity remained unchanged. We have also investigated the involvement of various potential intracellular mediators of nerve growth factor action and of known intracellular modulatory systems of the epidermal growth factor receptor for their capacity to participate in this nerve growth factor activity. 相似文献
13.
Eugenia Yakovchenko Michael Whalin Vilen Movsesyan Gordon Guroff 《Journal of neurochemistry》1996,67(2):540-548
Abstract: Receptors for insulin-like growth factor I (IGF-I) were studied on PC12EY cells, a subclone of PC12. Differentiation of PC12EY cells with nerve growth factor (NGF) did not alter either the number of IGF-I receptors nor their affinity for IGF-I. IGF-I receptors remained fully functional during differentiation, promoting increases in thymidine incorporation, glucose uptake, amino acid uptake, and the phosphorylation of the S6 protein of the ribosomes. IGF-I also increased the proportion of differentiated cells found in S-phase. But although the addition of IGF-I to naive cells caused an increase in cell number, there was no comparable increase when IGF-I was added to differentiated cells. Thus, although the receptor for IGF-I continues to be present and functional, IGF-I fails to induce cell proliferation in differentiated PC12 cells. 相似文献
14.
Regulation by Interleukin-1 of Nerve Growth Factor Secretion and Nerve Growth Factor mRNA Expression in Rat Primary Astroglial Cultures 总被引:10,自引:4,他引:6
Primary cultures of neonatal rat cortical astrocytes contain low cellular levels (about 2 pg/mg of protein) of nerve growth factor (NGF), but secrete significant amounts of NGF into the culture medium (about 540 pg of NGF/mg of cell protein/38-h incubation). Incubation of astrocytes with interleukin-1 (IL-1) increased the cellular content of NGF and the amount secreted by about threefold. In comparison, cerebellar astrocytes secreted significant amounts of NGF, and the secretion was also stimulated by IL-1. The stimulatory action of IL-1 on astrocytes prepared from cortex was dose- and time-dependent. Concentrations of IL-1 causing half-maximal and maximal stimulation of NGF secretion were 1 and 10 U/ml, respectively). Maximal NGF secretion induced by IL-1 (10 U/ml) was seen following 38 h of incubation. The basal secretion of NGF was reduced by about 50% under Ca2(+)-free conditions; however, the percent stimulation of NGF secretion by IL-1 was the same in the absence or presence of Ca2+. The stimulatory action of IL-1 was specific, because other glial growth factors and cytokines were almost ineffective in stimulating NGF secretion from cortical astroglial cells. IL-1 treatment also increased cellular NGF mRNA content twofold. The results indicate that IL-1 specifically triggers a cascade of events, independent of cell growth, which regulate NGF mRNA content and NGF secretion by astrocytes. 相似文献
15.
Magdalena Torres M. France Bader Dominique Aunis M. Teresa Miras-Portugal 《Journal of neurochemistry》1987,48(1):233-235
Chromaffin cells both recently isolated or in culture present a high-affinity adenosine transporter with a Km value of 1 microM. When cells were exposed to nerve growth factor (NGF; 10 ng/ml), the adenosine transporter affinity decreased to 3 microM. This value was maintained from 3 days after plating to the end of the culture period. A change in the transport capacity was observed, with a significant increase (approximately 200-260%) in NGF-cultured cells throughout the period studied. 相似文献
16.
Production of 1,2-Diacylglycerol in PC12 Cells by Nerve Growth Factor and Basic Fibroblast Growth Factor 总被引:10,自引:5,他引:5
The addition of nerve growth factor (NGF) or basic fibroblast growth factor (bFGF) to PC12 cells prelabeled with [3H]inositol and preincubated for 15 min in the presence of 10 mM LiCl stimulated the production of inositol phosphates with maximal increases of 120-180% in inositol monophosphate (IP), 130-200% in inositol bisphosphate (IP2), and 45-50% in inositol trisphosphate (IP3) within 30 min. The majority of the overall increase (approximately 85%) was in IP; the remainder was recovered as IP2 and IP3 (approximately 10% as IP2 and 5% as IP3). Under similar conditions, carbachol (0.5 mM) stimulated about a 10-fold increase in IP, a sixfold increase in IP2, and a fourfold increase in IP3. The mass level of 1,2-diacylglycerol (DG) in PC12 cells was found to be dependent on the incubation conditions; in growth medium [Dulbecco's modified Eagle's medium (DME) plus serum], it was around 6.2 mol %, in DME without serum, 2.5 mol %, and after a 15-min incubation in Dulbecco's phosphate-buffered saline, 0.62 mol %. The addition of NGF and bFGF induced an increase in the mass level of DG of about twofold within 1-2 min, often rising to two- to threefold by 15 min, and then decreasing slightly by 30 min. This increase was dependent on the presence of extracellular Ca2+, and was inhibited by both phenylarsine oxide (25 microM) and 5'-deoxy-5'-methylthioadenosine (3 mM). Under similar conditions, 0.5 mM carbachol stimulated the production of DG to the same extent as 200 ng/ml NGF and 50 ng/ml bFGF. Because carbachol is much more effective in stimulating the production of inositol phosphates, the results suggest that both NGF and bFGF stimulate the production of DG primarily from phospholipids other than the phosphoinositides. 相似文献
17.
Mari Oshima Linnea Weiss William C. Dougall Mark I. Greene Gordon Guroff 《Journal of neurochemistry》1995,65(1):427-433
Abstract: A small number of p185c- neu receptors have been found on PC12 cells. These receptors show some basal phosphorylation in quiescent cells. When the cells are treated with nerve growth factor (NGF) for a short time, some increase in phosphorylation is seen, mainly on serine and threonine residues, and this is accompanied by a slight shift in the apparent molecular weight. Epidermal growth factor (EGF) also increases the phosphorylation of p185c- neu , in this case on tyrosine residues. Neither heregulin-β1 nor gp30 stimulates the tyrosine phosphorylation of p185c- neu , and neither has a proliferative effect on the cells. Treatment of the cells with NGF for 5 days produces a 70–80% reduction in the number of p185c- neu receptors. This down-regulation does not occur when PC12nnr5 cells, which lack the high-affinity NGF receptor, p140 trk , are treated with NGF.The level of p185c- neu mRNA is not altered by NGF treatment, suggesting that the down-regulation is due to either a translational or a posttranslational alteration. 相似文献
18.
Expression of Nerve Growth Factor and Nerve Growth Factor Receptor Genes in Human Tissues and in Prostatic Adenocarcinoma Cell Lines 总被引:8,自引:0,他引:8
Nerve growth factor (NGF) mRNAs were detected and quantified in a variety of normal and neoplastic human tissues by northern blot hybridization. Human heart contained the highest NGF mRNA levels, whereas lower but comparable levels were found in the placenta, prostate, and kidney. All tissues examined coexpressed the low-affinity NGF receptor (LNGFR), whereas none of these tissues expressed the high-affinity NGF receptor encoded by the trk protooncogene. The widespread distribution of the LNGFR suggests that it plays a role in the regulation of normal cell growth. No overexpression of NGF or LNGFR mRNA was detected in neoplastic tissues, whereas LNGFR-like immunoreactivity was localized outside of tumor cells. Transforming growth factor-alpha and protooncogene c-fos expression in these tissues did not show a systematic correlation with NGF/LNGFR expression. Furthermore, regulation of the human NGF gene was studied in DU145 cells, a prostatic adenocarcinoma cell line that synthesizes significant NGF mRNA levels. Serum induced, whereas dexamethasone inhibited, NGF mRNA synthesis in these cells. Serum induction was preceded by a rapid and transient activation of the c-fos protooncogene. 相似文献
19.
Polyclonal antibodies to ganglioside GM1 have been prepared and characterised by direct and competitive enzyme-linked immunoassay. An immunoglobulin fraction was prepared from a rabbit antisera showing high specificity and antibody titre for GM1 relative to the other major brain gangliosides. The anti-GM1 immunoglobulin fraction and B-cholera toxin specifically labelled neurons in primary cultures of embryonic chick dorsal root ganglia and there was a good correlation between the relative increase in binding of anti-GM1 immunoglobulin and B-cholera toxin following neuraminidase treatment of a variety of cell types. At antibody concentrations that show saturable binding to endogenous ganglioside in the neuronal membrane, the anti-GM1 immunoglobulin fraction did not interfere with the nerve growth factor (NGF)-mediated fibre outgrowth and neuronal survival as indexed by measurement of neurofilament protein levels. Similarly, at levels in excess of those shown to stimulate thymocyte proliferation, B-cholera toxin was also without effect. These data are not consistent with GM1 in the neuronal membrane functioning as a receptor molecule for NGF and/or other differentiation factors present in the tissue culture media. 相似文献
20.
Previous studies from this laboratory have shown that the phosphorylation of the S6 protein of the ribosomes is catalyzed by at least two different and separable kinase activities in PC12 cells. One of these activities is increased by treatment of the cells with nerve growth factor, the other by treatment of the cells with epidermal growth factor. The present work shows that these two factors stimulate the phosphorylation of S6 with quite different kinetics, and that both the number of phosphates incorporated into S6 and the phosphopeptide pattern of S6 are different in cells treated with nerve growth factor than in cells treated with epidermal growth factor. The characteristics of the nerve growth factor-sensitive S6 kinase and of the epidermal growth factor-sensitive kinase were also clearly different. Substrate specificity and inhibitor studies indicated that neither was identical to cyclic AMP-dependent kinase, kinase C, or the calcium/calmodulin-dependent kinases. However, two major phosphopeptides produced by S6 phosphorylation in nerve growth factor-treated cells were also seen on phosphorylation of S6 by cyclic AMP-dependent kinase in vitro. In addition, when rat liver 40S ribosomal subunits were pretreated with cyclic AMP-dependent kinase in vitro, the action of the nerve growth factor-sensitive S6 kinase was increased about twofold. 相似文献