Receptors & channels最新文献_第3页

Desensitization of muscarinic receptors. 毒蕈碱受体的脱敏。

Receptors & channels Pub Date : 2004-01-01 DOI: 10.3109/10606820490281175

N. Fraeyman, J. van Emmelo, R. Paulssen, K. Vermis

引用次数: 0

Production of the human D2S receptor in the methylotrophic yeast P. pastoris. 人D2S受体在甲基营养酵母帕斯德酵母中的产生。

Receptors & channels Pub Date : 2004-01-01 DOI: 10.3109/10606820490279466

Sylvia Grünewald, Winfried Haase, Eva Molsberger, Hartmut Michel, Helmut Reiländer

{"title":"Production of the human D2S receptor in the methylotrophic yeast P. pastoris.","authors":"Sylvia Grünewald, Winfried Haase, Eva Molsberger, Hartmut Michel, Helmut Reiländer","doi":"10.3109/10606820490279466","DOIUrl":"https://doi.org/10.3109/10606820490279466","url":null,"abstract":"<p><p>In order to evaluate the methylotrophic yeast Pichia pastoris as means for high-yield production of homogenous D(2S) receptor protein, we have expressed the unmodified D(2S) receptor and various D(2S) receptor fusion constructs under the transcriptional control of the highly inducible promotor of the P. pastoris alcoholoxidase 1 gene in strain SMD1163. Fusion of the D(2S) receptor gene to the alpha-factor preprosequence proved to be essential for receptor production. For the receptor fusion constructs a gene dosage of more than two copies per cell increased production levels three- to sixfold. Adding various dopaminergic ligands to the induction medium increased yields up to tenfold, reaching 51,500 +/- 5700 receptors/cell. Immunoblot analysis of the effect of tunicamycin on D(2S) receptor fusion proteins and immunoprecipitation of metabolically labeled wild-type and glycosylation-deficient D(2S) receptor fusion proteins revealed that the high-mannose-type glycosylation of the D(2S) receptor prevents cleavage of the alpha-factor prosequence by the Kex2 endopeptidase. Abolishing glycosylation restored correct processing. Immunogold electron microscopy showed that recombinant yeast cells overproducing the D(2S) receptor developed membrane stacks harboring the receptor protein. The pharmacological profile of the recombinant D(2S) receptor was similar to that reported for neuronal D(2) receptors independent of glycosylation and processing. In conclusion, the D(2S) receptor can readily be produced in P. pastoris with high yield suitable for receptor purification and future structural studies.</p>","PeriodicalId":20928,"journal":{"name":"Receptors & channels","volume":"10 1","pages":"37-50"},"PeriodicalIF":0.0,"publicationDate":"2004-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3109/10606820490279466","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"24201827","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 32

Comparison of the pharmacological properties of rat Na(V)1.8 with rat Na(V)1.2a and human Na(V)1.5 voltage-gated sodium channel subtypes using a membrane potential sensitive dye and FLIPR. 利用膜电位敏感染料和FLIPR比较大鼠Na(V)1.8与大鼠Na(V)1.2a和人Na(V)1.5电压门控钠通道亚型的药理学性质。

Receptors & channels Pub Date : 2004-01-01 DOI: 10.3109/10606820490270410

R. Vickery, S. Amagasu, R. Chang, N. Mai, E. Kaufman, J. Martin, J. Hembrador, M. O'Keefe, C. Gee, D. Marquess, Jacqueline A.M. Smith

{"title":"Comparison of the pharmacological properties of rat Na(V)1.8 with rat Na(V)1.2a and human Na(V)1.5 voltage-gated sodium channel subtypes using a membrane potential sensitive dye and FLIPR.","authors":"R. Vickery, S. Amagasu, R. Chang, N. Mai, E. Kaufman, J. Martin, J. Hembrador, M. O'Keefe, C. Gee, D. Marquess, Jacqueline A.M. Smith","doi":"10.3109/10606820490270410","DOIUrl":"https://doi.org/10.3109/10606820490270410","url":null,"abstract":"A novel, membrane potential sensitive dye and a fluorescence imaging plate reader (FLIPR) have been used to characterize the pharmacological properties of rat Na(v)1.8 voltage-gated sodium channels (VGSC) in parallel with rat Na(v)1.2a and human Na(v)1.5 VGSC subtypes, respectively. The sensitivity of recombinant Na(v)1.2a-CHO, Na(v)1.5-293-EBNA, and Na(v)1.8-F-11 cells to VGSC activators was subtype dependent. Veratridine evoked depolarization of Na(v)1.2a-CHO and Na(v)1.5-293-EBNA cells with pEC(50) values of 4.78 +/- 0.13 and 4.84 +/- 0.12, respectively (n = 3), but had negligible effect on Na(v)1.8-F-11 cells (pEC(50) < 4.5). Type I pyrethroids were without significant effect at all subtypes. In contrast, the type II pyrethroids deltamethrin and fenvalerate evoked direct depolarization of Na(v)1.8-F-11 and Na(v)1.5-293-EBNA cells. Deltamethrin potentiated the veratridine-evoked response in Na(v)1.8-F-11 cells by > or =20-fold, in contrast to a <or =3-fold potentiation of the response in Na(v)1.2a, and Na(v)1.5 cells. Tetrodotoxin (TTX) inhibited VGSC activator-evoked depolarization of Na(v)1.8-F-11 cells with a biphasic concentration-response curve. The calculated pIC(50) values were 8.05 +/- 0.25 (n = 4) and 4.32 +/- 0.21 (n = 4), corresponding to TTX inhibition of endogenous TTX-sensitive (TTX-S), and recombinant Na(v)1.8 TTX-resistant (TTX-R) VGSCs, respectively. With the exception of TTX, the potencies of a number of ion channel blockers for the Na(v)1.8, Na(v)1.2a, and Na(v)1.5 VGSC subtypes were similar. In summary, these high-throughput FLIPR assays represent a valuable tool for the determination of the relative potencies of compounds at different VGSC subtypes and may prove useful for the identification of novel subtype-selective inhibitors.","PeriodicalId":20928,"journal":{"name":"Receptors & channels","volume":"12 1","pages":"11-23"},"PeriodicalIF":0.0,"publicationDate":"2004-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84413805","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 30

Human gastric tissues simultaneously express the classical and alternative splicing cholecystokinin-B/gastrin receptors. 人胃组织同时表达经典剪接和选择性剪接胆囊收缩素- b /胃泌素受体。

Receptors & channels Pub Date : 2004-01-01 DOI: 10.3109/10606820490926179

Jianjiang Zhou, Manling Chen, Qunzhou Zhang, Jiankun Hu, Wenling Wang

引用次数: 5

Desensitization of Muscarinic Receptors 毒蕈碱受体的脱敏

Receptors & channels Pub Date : 2004-01-01 DOI: 10.1080/10606820490281175

N. Fraeyman, J. V. Emmelo, Rh Paulssen, K. Vermis

引用次数: 2

Structure/activity relationships of M2 muscarinic allosteric modulators. M2毒蕈碱变构调节剂的构效关系。

Receptors & channels Pub Date : 2003-01-01

K Mohr, C Tränkle, U Holzgrabe

引用次数: 0

Flip the tip: an automated, high quality, cost-effective patch clamp screen. 翻转提示:一个自动化，高品质，高性价比的膜片钳屏幕。

Receptors & channels Pub Date : 2003-01-01

Albrecht Lepple-Wienhues, Klaus Ferlinz, Achim Seeger, Arvid Schäfer

引用次数: 0

Microstructured apertures in planar glass substrates for ion channel research. 平面玻璃基板微结构孔径离子通道研究。

Receptors & channels Pub Date : 2003-01-01

Niels Fertig, Michael George, Michèle Klau, Christine Meyer, Armin Tilke, Constanze Sobotta, Robert H Blick, Jan C Behrends

引用次数: 0

Guest Editor's Introduction: An Evolution of Electrophysiology 客座编辑简介:电生理学的进化

Receptors & channels Pub Date : 2003-01-01 DOI: 10.3109/10606820308255

J. Worley

引用次数: 2

CYTOCENTERING: a novel technique enabling automated cell-by-cell patch clamping with the CYTOPATCH chip. 细胞定心:一种新颖的技术，可以使用CYTOPATCH芯片实现细胞对细胞的自动贴片夹紧。

Receptors & channels Pub Date : 2003-01-01 DOI: 10.1080/10606820308254

A. Stett, C. Burkhardt, U. Weber, Peter van Stiphout, T. Knott

{"title":"CYTOCENTERING: a novel technique enabling automated cell-by-cell patch clamping with the CYTOPATCH chip.","authors":"A. Stett, C. Burkhardt, U. Weber, Peter van Stiphout, T. Knott","doi":"10.1080/10606820308254","DOIUrl":"https://doi.org/10.1080/10606820308254","url":null,"abstract":"Automats for patch clamping suspended cells in whole-cell configuration must (1) bring isolated cells in contact with patch contacts, (2) form gigaseals, and (3) establish stable intracellular access that allows for high quality recording of ionic currents. Single openings in planar substrates seem to be intriguing simple solutions for these problems, but due to the low rate of formation of whole-cell configurations we discarded this approach. Single openings are not suited for both attracting cells to the opening by suction and forming gigaseals with subsequent membrane rupture. To settle the three tasks with a mechanical microstructure we developed the socalled CYTOCENTERING technique to apply to suspended cells the same operation sequence as in conventional patch clamping. With this method we immobilized selected cells from a flowing suspension on the tip of a patch pipette by suction with a success rate of 97% and formed gigaseals with a success rate of 68%. Subsequent whole-cell recordings and intracellular staining with Lucifer yellow proved the stable access to the cytoplasm. Currently, a chip with an embedded suction opening in glass surrounding the microstructured contact pipette is under development. The processing of this CYTOPATCH chip is compatible to large-volume production. The CYTOPATCH automat will allow for fully automated, parallel, and asynchronous whole-cell recordings.","PeriodicalId":20928,"journal":{"name":"Receptors & channels","volume":"81 1","pages":"59-66"},"PeriodicalIF":0.0,"publicationDate":"2003-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77564878","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 80