Channelrhodopsin-2 ectopically expressed in the retina can recover the response to blue light in genetically blind mice and rats, but is unable to restore visual function due to optic nerve or optic tract lesions. Lon...Channelrhodopsin-2 ectopically expressed in the retina can recover the response to blue light in genetically blind mice and rats, but is unable to restore visual function due to optic nerve or optic tract lesions. Long Evans rats at postnatal day 1 were used for primary culture of visual cortical cells and 24 hours later, cells were transfected with recombinant adenovirus carrying channelrhodopsin-2 and green fluorescent protein genes. After 2 4 days of transfection, green fluorescence was visible in the cultured cells. Cells were stimulated with blue light （470 nm）, and light-induced action potentials were recorded in patch-clamp experiments. Our findings indicate that channelrhodopsin-2-recombinant adenovirus transfection of primary cultured visual cortical cells can control the production of action potentials via blue light stimulation.展开更多
We have delivered viral vectors containing either Chop2 fused with GFP, Channelrhodopsin-2 （ChR2）, or Halorhodopsin （HaloR） fused with mCherry （to form light gated cation channels or chloride pumps, respectively...We have delivered viral vectors containing either Chop2 fused with GFP, Channelrhodopsin-2 （ChR2）, or Halorhodopsin （HaloR） fused with mCherry （to form light gated cation channels or chloride pumps, respectively）, into the dorsal cochlear nucleus （DCN）. One to eighteen months later we examined the CN and inferior colliculus （IC） for evidence of virally transfected cells and processes. Production of ChR2 and HaloR was observed throughout the DCN. Rhodopsin localization within neurons was determined, with elongate, fusiform and giant cells identified based on morphology and location within the DCN. Production of ChR2 and HaloR was found at both the injection site as well as in regions projecting to and from the DCN. Light driven neuronal activity in the DCN was dependent upon the wavelength and intensity of the light, with only the appropriate wavelength resulting in activation and higher intensity light resulting in more neuronal activity. Transfecting cells via viral delivery of rhodopsins can be useful as a tract tracer and as a neuronal marker to delineate pathways. In the future rhodopsin delivery and activation may be developed as an alternative to electrical stimulation of neurons.展开更多
Optogenetics is a combination of optics and genetics technology that can be used to activate or inhibit specific cells in tissues. It has been used to treat Parkinson’s disease, epilepsy and neurological diseases, bu...Optogenetics is a combination of optics and genetics technology that can be used to activate or inhibit specific cells in tissues. It has been used to treat Parkinson’s disease, epilepsy and neurological diseases, but rarely Alzheimer’s disease. Adeno-associated virus carrying the CaMK promoter driving the optogenetic channelrhodopsin-2 (CHR2) gene (or without the CHR2 gene, as control) was injected into the bilateral dentate gyri, followed by repeated intrahippocampal injections of soluble low-molecular-weight amyloid-β1–42 peptide (Aβ1–42). Subsequently, the region was stimulated with a 473 nm laser (1–3 ms, 10 Hz, 5 minutes). The novel object recognition test was conducted to test memory function in mice. Immunohistochemical staining was performed to analyze the numbers of NeuN and synapsin Ia/b-positive cells in the hippocampus. Western blot assay was carried out to analyze the expression levels of glial fibrillary acidic protein, NeuN, synapsin Ia/b, metabotropic glutamate receptor-1a (mGluR-1a), mGluR-5, N-methyl-D-aspartate receptor subunit NR1, glutamate receptor 2, interleukin-1β, interleukin-6 and interleukin-10. Optogenetic stimulation improved working and short-term memory in mice with Alzheimer’s disease. This neuroprotective effect was associated with increased expression of NR1, glutamate receptor 2 and mGluR-5 in the hippocampus, and decreased expression of glial fibrillary acidic protein and interleukin-6. Our results show that optogenetics can be used to regulate the neuronal-glial network to ameliorate memory functions in mice with Alzheimer’s disease. The study was approved by the Animal Resources Committee of Jinan University, China (approval No. LL-KT-2011134) on February 28, 2011.展开更多
目的研究光感基因技术对脊髓栓系神经源性膀胱组织及其受体P2X1、P2X3的影响。方法将60只未成年SD雌性大鼠(尿流动力学检查无异常)随机分组,建立脊髓栓系神经源性膀胱动物模型,分为对照组(n=15)、脊髓栓系无光感基因组(n=21)、脊髓栓系...目的研究光感基因技术对脊髓栓系神经源性膀胱组织及其受体P2X1、P2X3的影响。方法将60只未成年SD雌性大鼠(尿流动力学检查无异常)随机分组,建立脊髓栓系神经源性膀胱动物模型,分为对照组(n=15)、脊髓栓系无光感基因组(n=21)、脊髓栓系有光感基因组(n=21)。HE染色观察膀胱组织的病理学改变,Real time PCR和Western blotting检测膀胱逼尿肌受体P2X1、P2X3在分子和蛋白水平的表达情况。结果在建模过程中,诱发电位监测发现术中有3只动物脊髓受损,其余动物建模成功。与脊髓栓系无光感基因组比较,在HE染色结果中脊髓栓系有光感基因组逼尿肌细胞形态大小相对均一,肌纤维排列较有序,趋近于对照组。Real time PCR和Western blotting检测结果显示,与对照组比较,脊髓栓系无光感基因组P2X1、P2X3 mRNA和相对蛋白表达量均升高,差异有统计学意义(P<0.05);而经蓝光照射一定时间后,脊髓栓系有光感基因组中两种受体mRNA和相对蛋白表达量均降低,与脊髓栓系无光感基因组比较,差异有统计学意义(P<0.05);与对照组比较,差异无统计学意义(P>0.05)。结论光感基因技术对脊髓栓系神经源性膀胱组织及其受体P2X1、P2X3有一定的调控作用。展开更多
基金sponsored by the National Natural Science Foundation of China (General Program),No.81070749,31070882,30970758Key Project of Chongqing Science & Technology Commission,No. 2010AB5118
文摘Channelrhodopsin-2 ectopically expressed in the retina can recover the response to blue light in genetically blind mice and rats, but is unable to restore visual function due to optic nerve or optic tract lesions. Long Evans rats at postnatal day 1 were used for primary culture of visual cortical cells and 24 hours later, cells were transfected with recombinant adenovirus carrying channelrhodopsin-2 and green fluorescent protein genes. After 2 4 days of transfection, green fluorescence was visible in the cultured cells. Cells were stimulated with blue light （470 nm）, and light-induced action potentials were recorded in patch-clamp experiments. Our findings indicate that channelrhodopsin-2-recombinant adenovirus transfection of primary cultured visual cortical cells can control the production of action potentials via blue light stimulation.
基金supported by Ralph Wilson Foundation(to A.G.H)Capita Foundation(to A.G.H)
文摘We have delivered viral vectors containing either Chop2 fused with GFP, Channelrhodopsin-2 （ChR2）, or Halorhodopsin （HaloR） fused with mCherry （to form light gated cation channels or chloride pumps, respectively）, into the dorsal cochlear nucleus （DCN）. One to eighteen months later we examined the CN and inferior colliculus （IC） for evidence of virally transfected cells and processes. Production of ChR2 and HaloR was observed throughout the DCN. Rhodopsin localization within neurons was determined, with elongate, fusiform and giant cells identified based on morphology and location within the DCN. Production of ChR2 and HaloR was found at both the injection site as well as in regions projecting to and from the DCN. Light driven neuronal activity in the DCN was dependent upon the wavelength and intensity of the light, with only the appropriate wavelength resulting in activation and higher intensity light resulting in more neuronal activity. Transfecting cells via viral delivery of rhodopsins can be useful as a tract tracer and as a neuronal marker to delineate pathways. In the future rhodopsin delivery and activation may be developed as an alternative to electrical stimulation of neurons.
基金supported by the National Natural Science Foundation of China,No.81171191(to LYZ)the Shenzhen Special Fund Project on Strategic Emerging Industry Development of China,No.JCYJ20160422170522075(to LYZ)the Shenzhen Healthcare Research Project of China,No.201601015(to LYZ)
文摘Optogenetics is a combination of optics and genetics technology that can be used to activate or inhibit specific cells in tissues. It has been used to treat Parkinson’s disease, epilepsy and neurological diseases, but rarely Alzheimer’s disease. Adeno-associated virus carrying the CaMK promoter driving the optogenetic channelrhodopsin-2 (CHR2) gene (or without the CHR2 gene, as control) was injected into the bilateral dentate gyri, followed by repeated intrahippocampal injections of soluble low-molecular-weight amyloid-β1–42 peptide (Aβ1–42). Subsequently, the region was stimulated with a 473 nm laser (1–3 ms, 10 Hz, 5 minutes). The novel object recognition test was conducted to test memory function in mice. Immunohistochemical staining was performed to analyze the numbers of NeuN and synapsin Ia/b-positive cells in the hippocampus. Western blot assay was carried out to analyze the expression levels of glial fibrillary acidic protein, NeuN, synapsin Ia/b, metabotropic glutamate receptor-1a (mGluR-1a), mGluR-5, N-methyl-D-aspartate receptor subunit NR1, glutamate receptor 2, interleukin-1β, interleukin-6 and interleukin-10. Optogenetic stimulation improved working and short-term memory in mice with Alzheimer’s disease. This neuroprotective effect was associated with increased expression of NR1, glutamate receptor 2 and mGluR-5 in the hippocampus, and decreased expression of glial fibrillary acidic protein and interleukin-6. Our results show that optogenetics can be used to regulate the neuronal-glial network to ameliorate memory functions in mice with Alzheimer’s disease. The study was approved by the Animal Resources Committee of Jinan University, China (approval No. LL-KT-2011134) on February 28, 2011.
文摘目的研究光感基因技术对脊髓栓系神经源性膀胱组织及其受体P2X1、P2X3的影响。方法将60只未成年SD雌性大鼠(尿流动力学检查无异常)随机分组,建立脊髓栓系神经源性膀胱动物模型,分为对照组(n=15)、脊髓栓系无光感基因组(n=21)、脊髓栓系有光感基因组(n=21)。HE染色观察膀胱组织的病理学改变,Real time PCR和Western blotting检测膀胱逼尿肌受体P2X1、P2X3在分子和蛋白水平的表达情况。结果在建模过程中,诱发电位监测发现术中有3只动物脊髓受损,其余动物建模成功。与脊髓栓系无光感基因组比较,在HE染色结果中脊髓栓系有光感基因组逼尿肌细胞形态大小相对均一,肌纤维排列较有序,趋近于对照组。Real time PCR和Western blotting检测结果显示,与对照组比较,脊髓栓系无光感基因组P2X1、P2X3 mRNA和相对蛋白表达量均升高,差异有统计学意义(P<0.05);而经蓝光照射一定时间后,脊髓栓系有光感基因组中两种受体mRNA和相对蛋白表达量均降低,与脊髓栓系无光感基因组比较,差异有统计学意义(P<0.05);与对照组比较,差异无统计学意义(P>0.05)。结论光感基因技术对脊髓栓系神经源性膀胱组织及其受体P2X1、P2X3有一定的调控作用。
