この作品は、グラントヴァンダービルトマウスの代謝表現型のセンターに5 - U24 - DK059637 - 10によってサポートされていました。

<ol>
	<li>Ayala, J. E., Bracy, D. P., McGuinness, O. P., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Considerations+in+the+design+of+hyperinsulinemic-euglycemic+clamps+in+the+conscious+mouse.">Considerations in the design of hyperinsulinemic-euglycemic clamps in the conscious mouse.</a> Diabetes. 55, 390-397 (2006).</li><li>Berglund, E. D., Li, C. Y., Poffenberger, G., Ayala, J. E., Fueger, P. T., Willis, S. E., Jewell, M. M., Powers, A. C., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose+metabolism+in+vivo+in+four+commonly+used+inbred+mouse+strains.">Glucose metabolism in vivo in four commonly used inbred mouse strains.</a> Diabetes. 57, 1790-1799 (2008).</li><li>Niswender, K. D., Shiota, M., Postic, C., Cherrington, A. D., Magnuson, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+increased+glucokinase+gene+copy+number+on+glucose+homeostasis+and+hepatic+glucose+metabolism.">Effects of increased glucokinase gene copy number on glucose homeostasis and hepatic glucose metabolism.</a> J. Biol. Chem. 272, 22570-22575 (1997).</li><li>Kim, H. J., Higashimori, T., Park, S. Y., Choi, H., Dong, J., Kim, Y. J., Noh, H. L., Cho, Y. R., Cline, G., Kim, Y. B., Kim, J. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+effects+of+interleukin-6+and+-10+on+skeletal+muscle+and+liver+insulin+action+in+vivo.">Differential effects of interleukin-6 and -10 on skeletal muscle and liver insulin action in vivo.</a> Diabetes. 53, 1060-1067 (2004).</li><li>Kim, J. K., Fillmore, J. J., Chen, Y., Yu, C., Moore, I. K., Pypaert, M., Lutz, E. P., Kako, Y., Velez-Carrasco, W., Goldberg, I. J., Breslow, J. L., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue-specific+overexpression+of+lipoprotein+lipase+causes+tissue-specific+insulin+resistance.">Tissue-specific overexpression of lipoprotein lipase causes tissue-specific insulin resistance.</a> Proc. Natl. Acad. Sci. U. S. A. 98, 7522-7527 (2001).</li><li>Kim, J. K., Fillmore, J. J., Gavrilova, O., Chao, L., Higashimori, T., Choi, H., Kim, H. J., Yu, C., Chen, Y., Qu, X., Haluzik, M., Reitman, M. L., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+effects+of+rosiglitazone+on+skeletal+muscle+and+liver+insulin+resistance+in+A-ZIP/F-1+fatless+mice.">Differential effects of rosiglitazone on skeletal muscle and liver insulin resistance in A-ZIP/F-1 fatless mice.</a> Diabetes. 52, 1311-1318 (2003).</li><li>Kim, J. K., Fillmore, J. J., Sunshine, M. J., Albrecht, B., Higashimori, T., Kim, D. W., Liu, Z. X., Soos, T. J., Cline, G. W., O'Brien, W. R., Littman, D. R., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PKC-theta+knockout+mice+are+protected+from+fat-induced+insulin+resistance.">PKC-theta knockout mice are protected from fat-induced insulin resistance.</a> J. Clin. Invest. 114, 823-827 (2004).</li><li>Kim, J. K., Gavrilova, O., Chen, Y., Reitman, M. L., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanism+of+insulin+resistance+in+A-ZIP/F-1+fatless+mice.">Mechanism of insulin resistance in A-ZIP/F-1 fatless mice.</a> J. Biol. Chem. 275, 8456-8460 (2000).</li><li>Kim, J. K., Gimeno, R. E., Higashimori, T., Kim, H. J., Choi, H., Punreddy, S., Mozell, R. L., Tan, G., Stricker-Krongrad, A., Hirsch, . <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inactivation+of+fatty+acid+transport+protein+1+prevents+fat-induced+insulin+resistance+in+skeletal+muscle.">Inactivation of fatty acid transport protein 1 prevents fat-induced insulin resistance in skeletal muscle.</a> J. Clin. Invest. 113, 756-763 (2004).</li><li>Kim, J. K., Kim, H. J., Park, S. Y., Cederberg, A., Westergren, R., Nilsson, D., Higashimori, T., Cho, Y. R., Liu, Z. X., Dong, J., Cline, G. W., Enerback, S., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adipocyte-specific+overexpression+of+FOXC2+prevents+diet-induced+increases+in+intramuscular+fatty+acyl+CoA+and+insulin+resistance.">Adipocyte-specific overexpression of FOXC2 prevents diet-induced increases in intramuscular fatty acyl CoA and insulin resistance.</a> Diabetes. 54, 1657-1663 (2005).</li><li>Kim, J. K., Kim, Y. J., Fillmore, J. J., Chen, Y., Moore, I., Lee, J., Yuan, M., Li, Z. W., Karin, M., Perret, P., Shoelson, S. E., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+of+fat-induced+insulin+resistance+by+salicylate.">Prevention of fat-induced insulin resistance by salicylate.</a> J. Clin. Invest. 108, 437-446 (2001).</li><li>Kim, J. K., Michael, P. r. e. v. i. s., Peroni, S. F., Mauvais-Jarvis, O. D., Neschen, F., Kahn, S., Kahn, B. B., R, C., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Redistribution+of+substrates+to+adipose+tissue+promotes+obesity+in+mice+with+selective+insulin+resistance+in+muscle.">Redistribution of substrates to adipose tissue promotes obesity in mice with selective insulin resistance in muscle.</a> J. Clin. Invest. 105, 1791-1797 (2000).</li><li>Kim, J. K., Zisman, A., Fillmore, J. J., Peroni, O. D., Kotani, K., Perret, P., Zong, H., Dong, J., Kahn, C. R., Kahn, B. B., Shulman, G. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose+toxicity+and+the+development+of+diabetes+in+mice+with+muscle-specific+inactivation+of+GLUT4.">Glucose toxicity and the development of diabetes in mice with muscle-specific inactivation of GLUT4.</a> J. Clin. Invest. 108, 153-160 (2001).</li><li>Haluzik, M., Gavrilova, O., LeRoith, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peroxisome+proliferator-activated+receptor-alpha+deficiency+does+not+alter+insulin+sensitivity+in+mice+maintained+on+regular+or+high-fat+diet:+hyperinsulinemic-euglycemic+clamp+studies.">Peroxisome proliferator-activated receptor-alpha deficiency does not alter insulin sensitivity in mice maintained on regular or high-fat diet: hyperinsulinemic-euglycemic clamp studies.</a> Endocrinology. 145, 1662-1667 (2004).</li><li>Haluzik, M., Yakar, S., Gavrilova, O., Setser, J., Boisclair, Y., LeRoith, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insulin+resistance+in+the+liver-specific+IGF-1+gene-deleted+mouse+is+abrogated+by+deletion+of+the+acid-labile+subunit+of+the+IGF-binding+protein-3+complex:+relative+roles+of+growth+hormone+and+IGF-1+in+insulin+resistance.">Insulin resistance in the liver-specific IGF-1 gene-deleted mouse is abrogated by deletion of the acid-labile subunit of the IGF-binding protein-3 complex: relative roles of growth hormone and IGF-1 in insulin resistance.</a> Diabetes. 52, 2483-2489 (2003).</li><li>Haluzik, M. M., Lacinova, Z., Dolinkova, M., Haluzikova, D., Housa, D., Horinek, A., Vernerova, Z., Kumstyrova, T., Haluzik, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improvement+of+insulin+sensitivity+after+peroxisome+proliferator-activated+receptor-alpha+agonist+treatment+is+accompanied+by+paradoxical+increase+of+circulating+resistin+levels.">Improvement of insulin sensitivity after peroxisome proliferator-activated receptor-alpha agonist treatment is accompanied by paradoxical increase of circulating resistin levels.</a> Endocrinology. 147, 4517-4524 (2006).</li><li>Kim, H., Haluzik, M., Asghar, Z., Yau, D., Joseph, J. W., Fernandez, A. M., Reitman, M. L., Yakar, S., Stannard, B., Heron-Milhavet, L., Wheeler, M. B., LeRoith, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Peroxisome+proliferator-activated+receptor-alpha+agonist+treatment+in+a+transgenic+model+of+type+2+diabetes+reverses+the+lipotoxic+state+and+improves+glucose+homeostasis.">Peroxisome proliferator-activated receptor-alpha agonist treatment in a transgenic model of type 2 diabetes reverses the lipotoxic state and improves glucose homeostasis.</a> Diabetes. 52, 1770-1778 (2003).</li><li>Deibert, D. C., DeFronzo, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epinephrine-induced+insulin+resistance+in+man.">Epinephrine-induced insulin resistance in man.</a> J. Clin. Invest. 65, 717-721 (1980).</li><li>Rizza, R. A., Cryer, P. E., Haymond, M. W., Gerich, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adrenergic+mechanisms+for+the+effects+of+epinephrine+on+glucose+production+and+clearance+in+man.">Adrenergic mechanisms for the effects of epinephrine on glucose production and clearance in man.</a> J. Clin. Invest. 65, 682-689 (1980).</li><li>Nunemaker, C. S., Wasserman, D. H., McGuinness, O. P., Sweet, I. R., Teague, J. C., Satin, L. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insulin+secretion+in+the+conscious+mouse+is+biphasic+and+pulsatile.">Insulin secretion in the conscious mouse is biphasic and pulsatile.</a> Am. J. Physiol. Endocrinol. Metab. 290, 523-529 (2006).</li><li>Nunemaker, C. S., Zhang, M., Wasserman, D. H., McGuinness, O. P., Powers, A. C., Bertram, R., Sherman, A., Satin, L. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Individual+mice+can+be+distinguished+by+the+period+of+their+islet+calcium+oscillations:+is+there+an+intrinsic+islet+period+that+is+imprinted+in+vivo.">Individual mice can be distinguished by the period of their islet calcium oscillations: is there an intrinsic islet period that is imprinted in vivo.</a> Diabetes. 54, 3517-3522 (2005).</li><li>Halter, J. B., Beard, J. C., Jr, P. o. r. t. e., D, . <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Islet+function+and+stress+hyperglycemia:+plasma+glucose+and+epinephrine+interaction.">Islet function and stress hyperglycemia: plasma glucose and epinephrine interaction.</a> Am. J. Physiol. 247, 47-52 (1984).</li><li>Jacobson, L., Ansari, T., McGuinness, O. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Counterregulatory+deficits+occur+within+24+h+of+a+single+hypoglycemic+episode+in+conscious,+unrestrained,+chronically+cannulated+mice.">Counterregulatory deficits occur within 24 h of a single hypoglycemic episode in conscious, unrestrained, chronically cannulated mice.</a> Am. J. Physiol. Endocrinol. Metab. 290, 678-684 (2006).</li><li>Jacobson, L., Ansari, T., Potts, J., McGuinness, O. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucocorticoid-deficient+corticotropin-releasing+hormone+knockout+mice+maintain+glucose+requirements+but+not+autonomic+responses+during+repeated+hypoglycemia.">Glucocorticoid-deficient corticotropin-releasing hormone knockout mice maintain glucose requirements but not autonomic responses during repeated hypoglycemia.</a> Am. J. Physiol. Endocrinol. Metab. 291, 15-22 (2006).</li><li>Ayala, J. E., Bracy, D. P., James, F. D., Julien, B. M., Wasserman, D. H., Drucker, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+glucagon-like+peptide-1+receptor+regulates+endogenous+glucose+production+and+muscle+glucose+uptake+independent+of+its+incretin+action.">The glucagon-like peptide-1 receptor regulates endogenous glucose production and muscle glucose uptake independent of its incretin action.</a> Endocrinology. 150, 1155-1164 (2009).</li><li>Fueger, P. T., Bracy, D. P., Malabanan, C. M., Pencek, R. R., Granner, D. K., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hexokinase+II+overexpression+improves+exercise-stimulated+but+not+insulin-stimulated+muscle+glucose+uptake+in+high-fat-fed+C57BL/6J+mice.">Hexokinase II overexpression improves exercise-stimulated but not insulin-stimulated muscle glucose uptake in high-fat-fed C57BL/6J mice.</a> Diabetes. 53, 306-314 (2004).</li><li>Fueger, P. T., Bracy, D. P., Malabanan, C. M., Pencek, R. R., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distributed+control+of+glucose+uptake+by+working+muscles+of+conscious+mice:+roles+of+transport+and+phosphorylation.">Distributed control of glucose uptake by working muscles of conscious mice: roles of transport and phosphorylation.</a> Am. J. Physiol. Endocrinol. Metab. 286, 77-84 (2004).</li><li>Fueger, P. T., Heikkinen, S., Bracy, D. P., Malabanan, C. M., Pencek, R. R., Laakso, M., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hexokinase+II+partial+knockout+impairs+exercise-stimulated+glucose+uptake+in+oxidative+muscles+of+mice.">Hexokinase II partial knockout impairs exercise-stimulated glucose uptake in oxidative muscles of mice.</a> Am. J. Physiol. Endocrinol. Metab. 285, 958-963 (2003).</li><li>Fueger, P. T., Hess, H. S., Posey, K. A., Bracy, D. P., Pencek, R. R., Charron, M. J., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Control+of+exercise-stimulated+muscle+glucose+uptake+by+GLUT4+is+dependent+on+glucose+phosphorylation+capacity+in+the+conscious+mouse.">Control of exercise-stimulated muscle glucose uptake by GLUT4 is dependent on glucose phosphorylation capacity in the conscious mouse.</a> J. Biol. Chem. , (2004).</li><li>Fueger, P. T., Li, C. Y., Ayala, J. E., Shearer, J., Bracy, D. P., Charron, M. J., Rottman, J. N., Wasserman, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose+kinetics+and+exercise+tolerance+in+mice+lacking+the+GLUT4+glucose+transporter.">Glucose kinetics and exercise tolerance in mice lacking the GLUT4 glucose transporter.</a> J. Physiol. 582, 801-812 (2007).</li><li>Shearer, J., Coenen, K. R., Pencek, R. R., Swift, L. L., Wasserman, D. H., Rottman, J. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long+chain+fatty+acid+uptake+in+vivo:+comparison+of+[125I]-BMIPP+and+[3H]-bromopalmitate.">Long chain fatty acid uptake in vivo: comparison of [125I]-BMIPP and [3H]-bromopalmitate.</a> Lipids. 43, 703-711 (2008).</li></ol>

利害の衝突は宣言されません。

高インスリン血症-正常血糖クランプ、またはインスリンクランプは、広く生体内でインスリンの作用を評価するための&quot;ゴールドスタンダード&quot;の方法と考えられている。この手法は、ヒト、イヌ、ラットやマウスを含むいくつかの種に適用されています。代謝性疾患のためのトランスジェニックマウスモデルの増加を考えると、マウスで使用するための技術の微細化は代謝の研究に大きな進歩を提供しています。 インスリンクランプの背後にある概念は単純ですが、実際にインスリンクランプ実験を行うためのさまざまなアプローチがあります。実験が行われる方法は、1を得られた結果に影響しますので、これは、些細な点ではありません。ここでは、ヴァンダービルトMMPCで使用されるプロトコルを提示する。我々のプロトコルや他の人のそれとの間の重要な違いは、我々は血液サンプルを得るために動脈カテーテルを使用することです。これはOBTのより広く使用されているアプローチとは対照的であるテール4、7、11、12、14から17の先端を切断することで血液サンプルをaining。動脈カテーテルからのサンプリングの利点は、実験が意識奔放マウスで実施されていることです。尾からのサンプリングはしばしば拘束を必要とし、大規模な血液サンプルが1を取得しているときにストレスの指標を向上させます。ストレスホルモンは、内因性グルコース産生を刺激し、グルコース処理18、19を損なう、潜在的にインスリン抵抗性の表現型の外観を与える。切断尾からサンプリングするため、そのストレスの多い性質の特殊な動物実験および使用委員会の承認を必要とする場合があります。動脈カテーテル法の手順は、尾を切断のマウスにストレスを回避するために開発されました。 インスリンクランプを行うことの重要な側面は、正常血糖を維持する能力です。正しくGIRは、血糖測定値に基づいて調整されるべきかを予測できるものアルゴリズムがありません。手術のような、インスリンクランプ実験を行う要員は、唯一の経験を通じて合理的な血糖値を維持するのに熟達するでしょう。ために彼らのより高い代謝率のマウスの研究から得られたデータは本質的に雑音になることに注意することは重要です。これは時間のグルコースとGIRのコースと血漿インスリン、endoRa、Rdと結果を解釈するためにあらゆるリーダーの能力のための重要なRgの絶対値を含むデータの完全なプレゼンテーションを、です。マウス（ヒトでの速度よりも約5倍）の高グルコースの流束率はグルコースのサンプリングの高周波数を保証。マウスの血液量が限られている間、一度、10分ごとの最小サンプリング周波数は、適切なクランプが確立されていることを確信する必要があります。 図4に示すように、クランプのインスリンレベルは、グループ間で異なる可能性があります。このような食事の介入、トランスジェニック操作やバックグラウンドの系統の違いなどの要因は、FASに影響を与える可能性がその後、クランプのインスリンレベルに影響を与える可能ティンのインスリンレベル、。クランプのインスリン濃度が異なる場合の結果を解釈するのが問題になる可能性があります。これは、実験群の間に相当するクランプのインスリンレベルを達成するインスリン注入速度を選択するためのパイロット実験を行うことにより対処することができます。また、ソマトスタチンは、膵臓のホルモンの分泌を抑制するために使用することができます、そしてインシュリンとグルカゴンは、実験的に制御するレートで交換することができます。この後者のアプローチは、より一般的にはマウスよりもラットにインスリンクランプで行われます。これらの実験的アプローチが取られていない場合、定常状態のGIRは、クランプインスリンレベルに正規化することができます、またはインスリン感受性指数（S I）は、S I = GIR /（G•ΔI）、などのクランプのデータから導出することができますGは、定常状態のグルコース濃度であり、ΔIは、空腹時やクランプのインスリン濃度との差です。どちらのアプローチで一つの仮定は達成クランプのインスリンレベルがあることです。インスリン感受性が直線的に研究されているグループに応じてインスリンのレベルに関連している範囲内で。インスリン抵抗性とインスリン感受性のグループを比較するときは、この後者の仮定は適用されない場合があります。理想的には、インスリンの用量反応曲線は、適切なインスリン注入を選択するために生成する必要があります。しかし、追加実験の必要性から、これはめったに行われません。 動脈カテーテルが提供する汎用性は、正常血糖クランプを超えた実験的なアプローチにも及ぶ。例えば、高血糖クランプは、グルコースが空腹時血糖値からの相対高血糖を維持するために可変速度で注入されている、 生体 2、20、21 の内因性膵臓機能を評価するために使用することができます。このテスト中に第一段階のインスリン分泌の測定は、血液サンプルの頻繁な買収（毎2〜5分間すなわち）、尾の先端からサンプルを取得するときには不可能ですが必要です。また、尾サンプリングに起因する上昇カテコールアミンがインスリン分泌を障害するとグルカゴンの分泌22を高めることができる。インスリンクランプのプロトコルはまた、反規制への反応2、23、24を評価するために相対的な低血糖に低下する血糖値を可能にするために変更することができます。動脈カテーテルはまた、運動25から30の間にグルコース代謝の動態を評価するために使用することができます。これは、単一の時点の前後の運動時または分離された筋肉を ex vivoで実施された従来の手法よりも著しく有利で ​​ある。ここで紹介するテクニックは、だけでなくグルコースでなく、脂肪酸の代謝31を評価するために使用することができます。

hyperinsulinemic-euglycemic clamps in conscious, unrestrained mice

2型糖尿病はインスリン作用の欠損によって特徴付けられる。高インスリン血症-正常血糖クランプ、またはインスリンクランプは、広く生体内でインスリンの作用を評価するための&quot;ゴールドスタンダード&quot;の方法と考えられている。インスリンクランプ時には、高インスリン血症は、一定のインスリン注入することによって達成される。正常血糖値は、可変レートで併用グルコース注入を介して維持されます。この変数はグルコース注入率（GIR）は、実験を通して短い間隔で血糖値を測定し、それに応じてGIRを調整することによって決定されます。強化インスリン作用を有するマウスは、より大きなGIRを必要とGIRは、全身インスリン作用の指標である。インスリンクランプは同位体2の管理を組み込むことができます[14 C]デオキシグルコースは、組織特異的グルコース取り込みを評価すると[3 - 3 H]グルコースは、内因性のグルコースの外観（endoRa）の速度を抑制するインスリンの能力、のマーカーを評価する肝臓でのグルコース産生、および刺激する全身グルコースの消失（路）の割合。 代謝性疾患の遺伝的マウスモデルで使用するためにインスリンクランプの小型化は、糖尿病の研究に大きな進歩をもたらした。インスリンクランプを実行するためのメソッドは、研究室によって異なります。インスリンクランプが行われる方法が大幅に得られた結果に影響を与えることができることに注意することは重要です。私たちは、意識的なマウス1と同様に様々なクランプ技法2を使用して、4つの一般的に使用される近交系マウスの代謝反応の評価にインスリンクランプを行うためにさまざまなアプローチの包括的な評価を発表している。ここでは、ヴァンダービルトマウス代謝表現型センター（;：www.mc.vanderbilt.edu / MMPC URL MMPC）により開発された意識、無拘束マウスにインスリンクランプを行うためのプロトコルを示す。これは、インスリンクランプ時に使用されるカテーテルを注入するための方法の説明が含まれています。で採用されているプロトコルバンダービルトMMPCは、ユニークな2カテーテルシステム3を利用しています。 Oneカテーテルは、注入のための頸静脈に挿入されます。第二カテーテルは、マウスを抑制または処理することなく、血液サンプリングを可能に頸動脈に挿入されます。このテクニックは、尾の切断端からのサンプリングです。インスリンクランプ時の血液サンプルを得るために最も一般的な方法に重要な利点を提供します。この後者のメソッドとは異なり、動脈カテーテルからサ ​​ンプリングすると、マウス1〜ストレスではありません。我々はまた、組織特異的なインスリン作用を評価するために同位体トレーサーの注入を使用するための方法を説明します。我々はまた、インスリンの留め金から得られた結果の適切なプレゼンテーションのためのガイドラインを提供しています。

Watch this Scientific Journal Video about Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice at JoVE.com

Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice

高インスリン血症 - 正常血糖クランプ、またはインスリンクランプは、インスリンの作用を評価するためのゴールドスタンダードです。<em&gt; in vivoで</em&gt;。マウスではインスリンクランプを行う方法について説明する。これは最小限のストレスで意識、無拘束マウスで実行する実験を許可する動脈カテーテル法のためのメソッドが含まれています。

意識、無拘束マウスにおける高インスリン血症、正常血糖クランプ

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold ...

hyperinsulinemic-euglycemic-clamps-in-conscious-unrestrained-mice

Research

JoVE Journal

Medicine

93.9K ビュー.  Sanford-Burnham Medical Research Institute at Lake Nona. 高インスリン血症 - 正常血糖クランプ、またはインスリンクランプは、インスリンの作用を評価するためのゴールドスタンダードです。 in vivoで。マウスではインスリンクランプを行う方法について説明する。これは最小限のストレスで意識、無拘束マウスで実行する実験を許可する動脈カテーテル法のためのメソッドが含まれています。

ビデオ: Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice

Transthoracic Echocardiography in Mice

In recent years, murine models have become the primary avenue for studying the molecular mechanisms of cardiac dysfunction resulting from changes in gene expression. Transgenic and gene targeting methods can be used to generate mice with altered cardiac size and function,1-3 and as a result, in vivo techniques are needed to evaluate their cardiac phenotype. Transthoracic echocardiography, pulse wave Doppler (PWD), and tissue Doppler imaging (TDI) can be used to provide dimensional measurements of the mouse heart and to quantify the degree of cardiac systolic and diastolic performance. Two-dimensional imaging is used to detect abnormal anatomy or movements of the left ventricle, whereas M-mode echo is used for quantification of cardiac dimensions and contractility.4,5 In addition, PWD is used to quantify localized velocity of turbulent flow,6 whereas TDI is used to measure the velocity of myocardial motion.7 Thus, transthoracic echocardiography offers a comprehensive method for the noninvasive evaluation of cardiac function in mice.

Transthoracic echocardiography offers a noninvasive method for the evaluation of cardiac function in mice. A combination of ultrasound and Doppler imaging modalities can be used to obtain dimensional measurements of the heart and intracardiac blood flow, which together provide an assessment of cardiac systolic and diastolic performance.

マウスにおける経胸壁心エコー

In recent years, murine models have become the primary avenue for studying the molecular mechanisms of cardiac dysfunction resulting from changes in gene ...

医学

Hyperinsulinemic-Euglycemic Clamp in the Conscious Rat

Type 2 diabetes (T2D) is rapidly rising in prevalence. Characterized by either inadequate insulin production or the inability to utilize insulin produced, T2D results in elevated blood glucose levels. The "gold-standard" in assessing insulin sensitivity is a hyperinsulinemic-euglycemic clamp or insulin clamp. In this procedure, insulin is infused at a constant rate resulting in a drop in blood glucose. To maintain blood glucose at a constant level, exogenous glucose (D50) is infused into the venous circulation. The amount of glucose infused to maintain homeostasis is indicative of insulin sensitivity. Here, we show the basic clamp procedure in the chronically catheterized, unrestrained, conscious rat. This model allows blood to be collected with minimal stress to the animal. Following the induction of anesthesia, a midline incision is made and the left common carotid artery and right jugular vein are catheterized. Inserted catheters are flushed with heparinized saline, then exteriorized and secured. Animals are allowed to recover for 4-5 days prior to experiments, with weight gain monitored daily. Only those animals who regain weight to pre-surgery levels are used for experiments. On the day of the experiment, rats are fasted and connected to pumps containing insulin and D50. Baseline glucose is assessed from the arterial line and used a benchmark throughout the experiment (euglycemia). Following this, insulin is infused at a constant rate into the venous circulation. To match the drop in blood glucose, D50 is infused. If the rate of D50 infusion is greater than the rate of uptake, a rise in glucose will occur. Similarly, if the rate is insufficient to match whole body glucose uptake, a drop will occur. Titration of glucose continues until stable glucose readings are achieved. Glucose levels and glucose infusion rates during this stable period are recorded and reported. Results provide an index of whole body insulin sensitivity. The technique can be refined to meet specific experimental requirements. It is further enhanced by the use of radioactive tracers that can determine tissue specific insulin-stimulated glucose uptake as well as whole body glucose turnover.

The hyperinsulinemic-euglycemic clamp is the "gold standard" for the assessment of insulin action. Insulin is infused at a constant rate stimulating glucose uptake. The amount of exogenous glucose infused to counter this drop is indicative of insulin sensitivity. Here the procedure is performed on a conscious, unrestrained rat.

覚醒ラットにおける高インスリン血症、正常血糖クランプ

Type 2 diabetes (T2D) is rapidly rising in prevalence. Characterized by either inadequate insulin production or the inability to utilize insulin produced, ...

Catheterization of the Carotid Artery and Jugular Vein to Perform Hemodynamic Measures, Infusions and Blood Sampling in a Conscious Rat Model

The success of a small animal model to study critical illness is, in part, dependent on the ability of the model to simulate the human condition. Intra-tracheal inoculation of a known amount of bacteria has been successfully used to reproduce the pathogenesis of pneumonia which then develops into sepsis. Monitoring hemodynamic parameters and providing standard clinical treatment including infusion of antibiotics, fluids and drugs to maintain blood pressure is critical to simulate routine supportive care in this model but to do so requires both arterial and venous vascular access. The video details the surgical technique for implanting carotid artery and common jugular vein catheters in an anesthetized rat. Following a 72 hr recovery period, the animals will be re-anesthetized and connected to a tether and swivel setup attached to the rodent housing which connects the implanted catheters to the hemodynamic monitoring system. This setup allows free movement of the rat during the study while continuously monitoring pressures, infusing fluids and drugs (antibiotics, vasopressors) and performing blood sampling.

Vascular accesses to measure hemodynamics, provide fluids and perform blood sampling are important to any small animal model study. We present a technique for implanting catheters into the carotid artery and the common jugular vein in an anesthetized rat for connecting to a system to perform monitoring, infusions and sampling.

配慮ラットモデルにおける血行動態の措置、煎茶や血液サンプリングを実行するために頸動脈と頸静脈のカテーテル

The success of a small animal model to study critical illness is, in part, dependent on the ability of the model to simulate the human condition. ...

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study model. As the use of mouse models are often a vital step in preclinical drug discovery and drug development1-8, it is necessary to design a system to introduce drugs into mice in a uniform, reproducible manner. Ideally, the system should permit the collection of blood samples at regular intervals over a set time course. The ability to measure drug concentrations by mass-spectrometry, has allowed investigators to follow the changes in plasma drug levels over time in individual mice1, 9, 10. In this study, paclitaxel was introduced into transgenic mice as a continuous arterial infusion over three hours, while blood samples were simultaneously taken by retro-orbital bleeds at set time points. Carotid artery infusions are a potential alternative to jugular vein infusions, when factors such as mammary tumors or other obstructions make jugular infusions impractical. Using this technique, paclitaxel concentrations in plasma and tissue achieved similar levels as compared to jugular infusion. In this tutorial, we will demonstrate how to successfully catheterize the carotid artery by preparing an optimized catheter for the individual mouse model, then show how to insert and secure the catheter into the mouse carotid artery, thread the end of the catheter out through the back of the mouse&#8217;s neck, and hook the mouse to a pump to deliver a controlled rate of drug influx. Multiple low volume retro-orbital bleeds allow for analysis of plasma drug concentrations over time.

This method was developed with the goal of delivering a steady drug solution via the carotid artery, to assess the pharmacokinetics of novel drugs in mouse models.

マウスにおけるタキサンの薬物動態学的および薬力学的分析のための頸動脈煎茶

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

Transcutaneous Assessment of Renal Function in Conscious Rodents

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and time-consuming. In addition, serial blood or urine samples are required, with the associated stress for the experimental animals. A recent technique significantly reduces the investment in time and resources, minimizing the invasiveness and the animal stress, but being equally valid as the traditional approaches. The method measures transcutaneously renal function. Using an optical device and the exogenous renal marker fluorescein isothiocyanate (FITC)-sinistrin, this technique is capable of measuring the elimination kinetics of the marker through the skin. With neither blood nor urine samples nor the associated laboratory assays needed, the results of the transcutaneous measurement are almost instantaneously available. The method has been already validated in different species and successfully applied in several models of renal pathology. Moreover, due to its minimally invasive characteristics, it is suitable for sequential measurements within the same animal. Here is provided a detailed protocol to carry out the transcutaneous assessment of renal function in rodents.

Determination of glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional procedures to measure this parameter are cumbersome and require a large investment of time. Here we describe a faster and minimally invasive method to determinate GFR transcutaneously.

コンシャスげっ歯類における腎機能の経皮評価

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and ...

Novel Approach for Simultaneous Recording of Renal Sympathetic Nerve Activity and Blood Pressure with Intravenous Infusion in Conscious, Unrestrained Mice.

Renal sympathetic nerves contribute significantly to both physiological and pathophysiological phenomena. Evaluating renal sympathetic nerve activity (RSNA) is of great interest in many areas of research such as chronic kidney disease, hypertension, heart failure, diabetes and obesity. Unequivocal assessment of the role of the sympathetic nervous system is thus imperative for proper interpretation of experimental results and understanding of disease processes. RSNA has been traditionally measured in anesthetized rodents, including mice. However, mice usually exhibit very low systemic blood pressure and hemodynamic instability for several hours during anesthesia and surgery. Meaningful interpretation of RSNA is confounded by this non-physiological state, given the intimate relationship between sympathetic nervous tone and cardiovascular status. To address this limitation of traditional approaches, we developed a new method for measuring RSNA in conscious, freely-moving mice. Mice were chronically instrumented with radio-telemeters for continuous monitoring of blood pressure as well as a jugular venous infusion catheter and custom-designed bipolar electrode for direct recording of RSNA. Following a 48-72 hour recovery period, survival rate was 100% and all mice behaved normally. At this time-point, RSNA was successfully recorded in 80% of mice, with viable signals acquired up to 4 and 5 days post-surgery in 70% and 50% of mice, respectively. Physiological blood pressures were recorded in all mice (116&plusmn;2 mmHg; n=10). Recorded RSNA increased with eating and grooming, as well-established in the literature. Furthermore, RSNA was validated by ganglionic blockade and modulation of blood pressure with pharmacological agents. Herein, an effective and manageable method for clear recording of RSNA in conscious, freely-moving mice is described.

Anesthetized mice exhibit non-physiological systemic blood pressure, which precludes meaningful assessment of autonomic tone given the intimate relationship between blood pressure and the autonomic nervous system. Thus, a novel method to simultaneously record renal sympathetic nerve activity and blood pressure with intravenous infusion in conscious mice is outlined.

腎交感神経活動と意識、持続静脈内投与による血圧の同時記録への新しいアプローチ気ままなマウス。

Renal sympathetic nerves contribute significantly to both physiological and pathophysiological phenomena. Evaluating renal sympathetic nerve activity ...

Assessment of Respiratory Function in Conscious Mice by Double-chamber Plethysmography

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to non-invasively assess some features of the respiratory function in humans as well as in laboratory animals. The present article focuses on the application of the double-chamber plethysmography (DCP) in small animals. It provides background information on the methodology as well as a detailed step-by-step procedure to successfully assess respiratory function in conscious, spontaneously breathing animals in a non-invasive manner. The DCP can be used to monitor the respiratory function of multiple animals in parallel, as well as to identify changes induced by aerosolized substances over a chosen time period and in a repeated manner. Experiments on control and allergic mice are used herein to demonstrate the utility of the technique, explain the associated outcome parameters, as well as to discuss the related advantages and shortcomings. Overall, the DCP provides valid and theoretically sound readouts that can be trusted to evaluate the respiratory function of conscious small animals both at baseline and after challenges with aerosolized substances.

The objective of the present article is to provide a detailed description of the recommended procedures to evaluate respiratory function in conscious mice by double-chamber plethysmography.

ダブル室容積脈波による意識下マウスにおける呼吸機能の評価

Air volume changes created by a conscious subject breathing spontaneously within a body box are at the basis of plethysmography, a technique used to ...

Repeated Orotracheal Intubation in Mice

The literature describes several methods for mouse intubation that either require visualization of the glottis through the oral cavity or incision in the ventral neck for direct confirmation of cannula placement in the trachea. The relative difficulty or the tissue trauma induced to the subject by such procedures can be an impediment to an investigator&#8217;s ability to perform longitudinal studies. This article illustrates a technique in which physical manipulation of the mouse following the use of a depilatory to remove hair from the ventral neck permits transcutaneous visualization of the trachea for orotracheal intubation regardless of degree of skin pigmentation. This method is innocuous to the subject and easily achieved with a limited understanding of murine anatomy. This refined approach facilitates repeated intubation, which may be necessary for monitoring progression of disease or instillation of treatments. Using this method may result in a reduction of the number of animals and technical skill required to measure lung function in mouse models of respiratory disease.

The goal of this article is to describe a refined method of intubation of the laboratory mouse. The method is noninvasive and, therefore, ideal for studies that require serial monitoring of respiratory function and/or instillation of treatments into the lung.

マウスにおける反復性の耳管挿管

The literature describes several methods for mouse intubation that either require visualization of the glottis through the oral cavity or incision in the ...

Implantation of Combined Telemetric ECG and Blood Pressure Transmitters to Determine Spontaneous Baroreflex Sensitivity in Conscious Mice

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. The baroreflex is a key homeostatic mechanism to counteract acute, short-term changes in arterial BP and to maintain BP in a relatively narrow physiological range. BP is sensed by baroreceptors located in the aortic arch and carotid sinus. When BP changes, signals are transmitted to the central nervous system and are then communicated to the parasympathetic and sympathetic branches of the autonomic nervous system to adjust HR. A rise in BP causes a reflex decrease in HR, a drop in BP causes a reflex increase in HR.
Baroreflex sensitivity (BRS) is the quantitative relationship between changes in arterial BP and corresponding changes in HR. Cardiovascular diseases are often associated with impaired baroreflex function. In various studies reduced BRS has been reported in e.g., heart failure, myocardial infarction, or coronary artery disease. 
Determination of BRS requires information from both BP and HR, which can be recorded simultaneously using telemetric devices. The surgical procedure is described beginning with the insertion of the pressure sensor into the left carotid artery and positioning of its tip in the aortic arch to monitor arterial pressure followed by the subcutaneous placement of the transmitter and ECG electrodes. We also describe postoperative intensive care and analgesic management. After a two-week period of post-surgery recovery long-term ECG and BP recordings are performed in conscious and unrestrained mice. Finally, we include examples of high-quality recordings and the analysis of spontaneous baroreceptor sensitivity using the sequence method.

The baroreflex is a heart-rate regulation mechanism by the autonomic nervous system in response to blood-pressure changes. We describe a surgical technique to implant telemetry transmitters for continuous and simultaneous measurement of electrocardiogram and blood pressure in mice. This can determine spontaneous baroreflex sensitivity, an important prognostic marker for cardiovascular disease.

意識のあるマウスにおける自発的な圧反射感受性を決定するための遠隔ECGと血圧伝達器の組み合わせの埋め込み

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. ...

Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits

Patients with ion channelopathies are at a high risk of developing seizures and fatal cardiac arrhythmias. There is a higher prevalence of heart disease and arrhythmias in people with epilepsy (i.e., epileptic heart.) Additionally, cardiac and autonomic disturbances have been reported surrounding seizures. 1:1,000 epilepsy patients/year die of sudden unexpected death in epilepsy (SUDEP). The mechanisms for SUDEP remain incompletely understood. Electroencephalograms (EEG) and electrocardiograms (ECG) are two techniques routinely used in the clinical setting to detect and study the substrates/triggers for seizures and arrhythmias. While many studies and descriptions of this methodology are in rodents, their cardiac electrical activity differs significantly from humans. This article provides a description of a non-invasive method for recording simultaneous video-EEG-ECG-oximetry-capnography in conscious rabbits. As cardiac electrical function is similar in rabbits and humans, rabbits provide an excellent model of translational diagnostic and therapeutic studies. In addition to outlining the methodology for data acquisition, we discuss the analytical approaches for examining neuro-cardiac electrical function and pathology in rabbits. This includes arrhythmia detection, spectral analysis of EEG and a seizure scale developed for restrained rabbits.

Using simultaneous video-EEG-ECG-oximetry-capnography, we developed a methodology to evaluate the susceptibility of rabbit models to develop provoked arrhythmias and seizures. This novel recording system establishes a platform to test the efficacy and safety of therapeutics and can capture the complex cascade of multi-system events that culminate in sudden death.

意識的に拘束されたウサギにおける発作、不整脈および無呼吸の識別のためのマルチシステムモニタリング

Patients with ion channelopathies are at a high risk of developing seizures and fatal cardiac arrhythmias. There is a higher prevalence of heart disease ...