OBJECTIVE Individuals vary in sensitivity to the behavioral effects of nicotine,resulting in differences in their vulnerability to addiction.The role of rearing environment in determining individual sensitivity to nic...OBJECTIVE Individuals vary in sensitivity to the behavioral effects of nicotine,resulting in differences in their vulnerability to addiction.The role of rearing environment in determining individual sensitivity to nicotine is unclear.The neuropharmacological mechanisms mediating the effect of rearing environment on the actions of nicotine are also understood.Thus,the contribution of rearing environment in determining the sensitivity to the locomotor effects of nicotine and regulating α4β2*-and α7-nicotinic acetylcholine(n ACh) receptor expressionwas determined in rats reared in isolated(IC) or enriched(EC) conditions.METHODS To measure locomotor activity,adolescent rats(postnatal day 21-51)were injected with saline(1 mL·kg^(-1)) or nicotine(0.3 mg·kg^(-1)) subcutaneously,then placed in chamberswhere ambulatory activity was monitored for 30-min by computer for 14 daily sessions.α4β2*-andα7-n ACh receptor expression in the mesolimbic dopamine pathway was determined by quantitative autoradiography of [125 I]-epibatidine and [125 I]-bungarotoxinbinding,respectively,in 16 μmol·L^(-1) coronal sections.Values for receptor expression in fmol are ±s of 8 brains and compared by two-tailed,unpaired t-test with P<0.05 considered significant.RESULTS EC-rats are similarly sensitive as IC-rats to the locomotor effects of nicotine.[125 I]-epibatidine binding in the ventral tegmental area of EC-rats was reduced(2.8±0.3 fmo L) compared to IC-rats(4.0±0.4 fmo L);there was no difference in the nucleus accumbens.There was no difference between EC-and IC-rats in α7-n ACh receptor expression in the mesolimbic dopamine pathway.CONCLUSION Rearing environment differentially regulates n ACh receptor subtypes in EC and IC rats.These data suggest regulation of n ACh receptors by environmental factors may be a mechanism for the protective effect of enrichment against altered sensitivity to nicotine in genetically vulnerable individuals.The characterization of these mechanisms will aid in development of novel pharmacological tools mimicking the protection afforded by environmental enrichment in nicotine-sensitive individuals.展开更多
Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. Th...Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. The present study was to determine the effects of activation of α7nAChR on oxidant stress-induced injury in endo- thelial cells. Methods Cultured human umbilical vein endothelial cells were treated with H202 （400 μmol · L^-1） or H202plus PNU-282987 （ 10 μmol · L^-1 ）. Cell viability and membrane integrity were measured. AnnexinV ＋ PI assay, immunoblotting of bcl-2, bax and cleaved caspase-3, and immunofluorescence of apoptosis inducing factor （AIF） were performed to evaluate apoptosis. Protein expression of vascular peroxidase-1 （ VPO-1 ） and phosphor- JNK were measured by immunoblotting. Results Activation of α7nAChR by a selective agonist PNU-282987 pre-vented H202-indced decrease of cell viability and increase of lactate dehydrogenase release. Activation of α7nAChR markedly reduced cell apoptosis and intracellular oxidative stress level. Moreover, activation of α7nAChR reduced H2 02 -induced VPO-1 protein upregulation and JNK1/2 phosphorylation. The inhibitory effect of α7nAChR activa- tion on VPO-1 was blocked by JNK inhibitor SP600125. In addition, pretreatment of α7nAChR antagonist methyl- lycaconitine blocked the cytoprotective effect of PNU-282987. Conclusion These results provide the first evidence that activation of α7nAChR protects against oxidant stress-induced damage by suppressing VPO-1 in a JNK signa- ling pathway-dependent manner in endothelial cells.展开更多
目的探讨β2-烟碱型乙酰胆碱受体(β2-n ACh R)在海马CA1和CA3锥体神经元的A型γ-氨基丁酸受体(GABA_A-R)发育中的作用。方法应用β2-n ACh R基因敲除小鼠(β2-KO组)制备急性分离的海马CA1和CA3锥体神经元,应用穿孔膜片钳记录技术记录GA...目的探讨β2-烟碱型乙酰胆碱受体(β2-n ACh R)在海马CA1和CA3锥体神经元的A型γ-氨基丁酸受体(GABA_A-R)发育中的作用。方法应用β2-n ACh R基因敲除小鼠(β2-KO组)制备急性分离的海马CA1和CA3锥体神经元,应用穿孔膜片钳记录技术记录GABA_A-R选择性激动剂蝇蕈醇在CA1和CA3锥体神经元诱导的GABA电流,测试其平衡电位(E_(Mus))和动力学指标,并与野生型小鼠(WT组)进行比较。结果β2-KO组小鼠(n=4)CA1锥体神经元(n=7)的E_(Mus)为-31.7±3.5 m V,与WT组相比向去极化偏移(P<0.05);CA3锥体神经元(n=4)的E_(Mus)为-16.1±4.6 m V,同样较WT组偏向去极化方向(P<0.01);与WT组小鼠不同,β2-KO组小鼠CA3和CA1神经元的E_(Mus)差异有统计学意义(P<0.05)。β2-KO组小鼠CA1和CA3神经元上都显示GABA_A-R的失敏显著减慢,衰减时间分别为2.2±0.2 s、3.2±0.1 s(WT组为1.6±0.1 s、2.3±0.1 s,P<0.05或P<0.01)。结论含β2的n ACh R可能参与促进小鼠海马CA1和CA3锥体细胞中GABA_A-R的功能成熟。展开更多
【目的】探讨α7烟碱乙酰胆碱受体（α7 n Ach R）激动剂对血管紧张素Ⅱ（AngⅡ）诱导的心肌成纤维细胞增殖和胶原合成的影响及其分子机制。【方法】培养出生2~3d大鼠心肌成纤维细胞（CF）,取传代2~4代细胞,分为1空白对照组（control）;...【目的】探讨α7烟碱乙酰胆碱受体（α7 n Ach R）激动剂对血管紧张素Ⅱ（AngⅡ）诱导的心肌成纤维细胞增殖和胶原合成的影响及其分子机制。【方法】培养出生2~3d大鼠心肌成纤维细胞（CF）,取传代2~4代细胞,分为1空白对照组（control）;2模型组（AngⅡ10-7 mol/L）;3ACh预处理组：ACh＋AngⅡ组（ACh 10-4 mol/L＋AngⅡ10-7 mol/L）;4α7n ACh R激动剂组：α7n ACh R激动剂＋AngⅡ组（PNU-282987 5×10-6 mol/L＋AngⅡ10-7 mol/L）;5α7n ACh R拮抗剂组：ACh＋甲基牛扁碱柠檬酸盐（MLA）＋AngⅡ组（ACh 10-4 mol/L＋AngⅡ10-7 mol/L＋MLA 1×10-6 mol/L）。干预24 h后,应用CCK-8试剂盒检测CF的增殖能力,western blot检测胶原蛋白Ι、α-SMA、α7烟碱乙酰胆碱受体（α7n ACh R）、核中p65蛋白表达;Real-time PCR检测α7n ACh R的m RNA表达。【结果】AngⅡ干预24h,CF增殖显著提高,胶原蛋白Ι、α-SMA、核中p65蛋白的表达明显升高,α7n ACh R的m RNA和蛋白水平均表达降低;Ach、α7n ACh R激动剂预处理后抑制CF增殖,胶原蛋白Ι、α-SMA和核中p65蛋白的表达降低,α7n ACh R的m RNA和蛋白水平均表达升高;Ach＋MLA预处理,CF增殖再次提高,胶原蛋白Ι、纤维化因子α-SMA、核中p65蛋白的表达再次明显升高,α7n ACh R的m RNA和蛋白水平均表达降低。【结论】α7n ACh R激动剂抑制AngⅡ诱导的心肌成纤维细胞的增殖和胶原合成,其机制可能与NF-κB信号通路有关。展开更多
基金supported by Nebraska Cancer and Smoking Disease Research Programs LB506and LB595 to CS BOCKMAN and DJ STAIRS
文摘OBJECTIVE Individuals vary in sensitivity to the behavioral effects of nicotine,resulting in differences in their vulnerability to addiction.The role of rearing environment in determining individual sensitivity to nicotine is unclear.The neuropharmacological mechanisms mediating the effect of rearing environment on the actions of nicotine are also understood.Thus,the contribution of rearing environment in determining the sensitivity to the locomotor effects of nicotine and regulating α4β2*-and α7-nicotinic acetylcholine(n ACh) receptor expressionwas determined in rats reared in isolated(IC) or enriched(EC) conditions.METHODS To measure locomotor activity,adolescent rats(postnatal day 21-51)were injected with saline(1 mL·kg^(-1)) or nicotine(0.3 mg·kg^(-1)) subcutaneously,then placed in chamberswhere ambulatory activity was monitored for 30-min by computer for 14 daily sessions.α4β2*-andα7-n ACh receptor expression in the mesolimbic dopamine pathway was determined by quantitative autoradiography of [125 I]-epibatidine and [125 I]-bungarotoxinbinding,respectively,in 16 μmol·L^(-1) coronal sections.Values for receptor expression in fmol are ±s of 8 brains and compared by two-tailed,unpaired t-test with P<0.05 considered significant.RESULTS EC-rats are similarly sensitive as IC-rats to the locomotor effects of nicotine.[125 I]-epibatidine binding in the ventral tegmental area of EC-rats was reduced(2.8±0.3 fmo L) compared to IC-rats(4.0±0.4 fmo L);there was no difference in the nucleus accumbens.There was no difference between EC-and IC-rats in α7-n ACh receptor expression in the mesolimbic dopamine pathway.CONCLUSION Rearing environment differentially regulates n ACh receptor subtypes in EC and IC rats.These data suggest regulation of n ACh receptors by environmental factors may be a mechanism for the protective effect of enrichment against altered sensitivity to nicotine in genetically vulnerable individuals.The characterization of these mechanisms will aid in development of novel pharmacological tools mimicking the protection afforded by environmental enrichment in nicotine-sensitive individuals.
文摘Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. The present study was to determine the effects of activation of α7nAChR on oxidant stress-induced injury in endo- thelial cells. Methods Cultured human umbilical vein endothelial cells were treated with H202 （400 μmol · L^-1） or H202plus PNU-282987 （ 10 μmol · L^-1 ）. Cell viability and membrane integrity were measured. AnnexinV ＋ PI assay, immunoblotting of bcl-2, bax and cleaved caspase-3, and immunofluorescence of apoptosis inducing factor （AIF） were performed to evaluate apoptosis. Protein expression of vascular peroxidase-1 （ VPO-1 ） and phosphor- JNK were measured by immunoblotting. Results Activation of α7nAChR by a selective agonist PNU-282987 pre-vented H202-indced decrease of cell viability and increase of lactate dehydrogenase release. Activation of α7nAChR markedly reduced cell apoptosis and intracellular oxidative stress level. Moreover, activation of α7nAChR reduced H2 02 -induced VPO-1 protein upregulation and JNK1/2 phosphorylation. The inhibitory effect of α7nAChR activa- tion on VPO-1 was blocked by JNK inhibitor SP600125. In addition, pretreatment of α7nAChR antagonist methyl- lycaconitine blocked the cytoprotective effect of PNU-282987. Conclusion These results provide the first evidence that activation of α7nAChR protects against oxidant stress-induced damage by suppressing VPO-1 in a JNK signa- ling pathway-dependent manner in endothelial cells.
文摘目的探讨β2-烟碱型乙酰胆碱受体(β2-n ACh R)在海马CA1和CA3锥体神经元的A型γ-氨基丁酸受体(GABA_A-R)发育中的作用。方法应用β2-n ACh R基因敲除小鼠(β2-KO组)制备急性分离的海马CA1和CA3锥体神经元,应用穿孔膜片钳记录技术记录GABA_A-R选择性激动剂蝇蕈醇在CA1和CA3锥体神经元诱导的GABA电流,测试其平衡电位(E_(Mus))和动力学指标,并与野生型小鼠(WT组)进行比较。结果β2-KO组小鼠(n=4)CA1锥体神经元(n=7)的E_(Mus)为-31.7±3.5 m V,与WT组相比向去极化偏移(P<0.05);CA3锥体神经元(n=4)的E_(Mus)为-16.1±4.6 m V,同样较WT组偏向去极化方向(P<0.01);与WT组小鼠不同,β2-KO组小鼠CA3和CA1神经元的E_(Mus)差异有统计学意义(P<0.05)。β2-KO组小鼠CA1和CA3神经元上都显示GABA_A-R的失敏显著减慢,衰减时间分别为2.2±0.2 s、3.2±0.1 s(WT组为1.6±0.1 s、2.3±0.1 s,P<0.05或P<0.01)。结论含β2的n ACh R可能参与促进小鼠海马CA1和CA3锥体细胞中GABA_A-R的功能成熟。
文摘【目的】探讨α7烟碱乙酰胆碱受体（α7 n Ach R）激动剂对血管紧张素Ⅱ（AngⅡ）诱导的心肌成纤维细胞增殖和胶原合成的影响及其分子机制。【方法】培养出生2~3d大鼠心肌成纤维细胞（CF）,取传代2~4代细胞,分为1空白对照组（control）;2模型组（AngⅡ10-7 mol/L）;3ACh预处理组：ACh＋AngⅡ组（ACh 10-4 mol/L＋AngⅡ10-7 mol/L）;4α7n ACh R激动剂组：α7n ACh R激动剂＋AngⅡ组（PNU-282987 5×10-6 mol/L＋AngⅡ10-7 mol/L）;5α7n ACh R拮抗剂组：ACh＋甲基牛扁碱柠檬酸盐（MLA）＋AngⅡ组（ACh 10-4 mol/L＋AngⅡ10-7 mol/L＋MLA 1×10-6 mol/L）。干预24 h后,应用CCK-8试剂盒检测CF的增殖能力,western blot检测胶原蛋白Ι、α-SMA、α7烟碱乙酰胆碱受体（α7n ACh R）、核中p65蛋白表达;Real-time PCR检测α7n ACh R的m RNA表达。【结果】AngⅡ干预24h,CF增殖显著提高,胶原蛋白Ι、α-SMA、核中p65蛋白的表达明显升高,α7n ACh R的m RNA和蛋白水平均表达降低;Ach、α7n ACh R激动剂预处理后抑制CF增殖,胶原蛋白Ι、α-SMA和核中p65蛋白的表达降低,α7n ACh R的m RNA和蛋白水平均表达升高;Ach＋MLA预处理,CF增殖再次提高,胶原蛋白Ι、纤维化因子α-SMA、核中p65蛋白的表达再次明显升高,α7n ACh R的m RNA和蛋白水平均表达降低。【结论】α7n ACh R激动剂抑制AngⅡ诱导的心肌成纤维细胞的增殖和胶原合成,其机制可能与NF-κB信号通路有关。
