1. Instrumentation 
<ol>
<li>To measure the response of olfactory sensory neurons to photolysis of caged compounds we use a flash lamp in combination with a typical patch-clamp recording system including: a patch-clamp amplifier, a recording electrode and a reference electrode connected to the head-stage of a patch-clamp amplifier, a digitizer, a computer, software for data acquisition, micro-manipulators, an epifluorescence microscope, a perfusion system, an anti-vibration table and a Faraday cage (Figure 2).</li>
<li...

<ol>
	<li>Ellis-Davies, G. C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Caged+compounds:+photorelease+technology+for+control+of+cellular+chemistry+and+physiology.">Caged compounds: photorelease technology for control of cellular chemistry and physiology.</a> Nat. Methods. 4, 619-628 (2007).</li><li>Pifferi, S., Boccaccio, A., Menini, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cyclic+nucleotide-gated+ion+channels+in+sensory+transduction.">Cyclic nucleotide-gated ion channels in sensory transduction.</a> FEBS Lett. 580, 2853-2859 (2006).</li><li>Bozza, T. C., Kauer, J. 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L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flash+photolysis+of+caged+compounds.">Flash photolysis of caged compounds.</a> Microelectrodes: Theory and Applications. , (1991).</li><li>Lagostena, L., Menini, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole-cell+recordings+and+photolysis+of+caged+compounds+in+olfactory+sensory+neurons+isolated+from+the+mouse.">Whole-cell recordings and photolysis of caged compounds in olfactory sensory neurons isolated from the mouse.</a> Chem. Senses. 28, 705-716 (2003).</li><li>Boccaccio, A., Lagostena, L., Hagen, V., Menini, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fast+adaptation+in+mouse+olfactory+sensory+neurons+does+not+require+the+activity+of+phosphodiesterase.">Fast adaptation in mouse olfactory sensory neurons does not require the activity of phosphodiesterase.</a> J. Gen. Physiol. 128, 171-184 (2006).</li><li>Boccaccio, A., Menini, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+development+of+cyclic+nucleotide-gated+and+Ca2++-activated+Cl-+currents+in+isolated+mouse+olfactory+sensory+neurons.">Temporal development of cyclic nucleotide-gated and Ca2+ -activated Cl- currents in isolated mouse olfactory sensory neurons.</a> J. 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Flash-Photolyse von caged Verbindungen mit Patch-Clamp-Aufnahmen kombiniert ist eine nützliche Technik, um schnelle und lokale Sprünge in der Konzentration der physiologisch aktive Moleküle sowohl innerhalb als auch außerhalb der Zellen zu erhalten. Mehrere Arten von Käfigen Verbindungen1 wurden synthetisiert, und diese Technik kann auf verschiedene Arten von Zellen, einschließlich kultivierten Zellen, die Ionenkanäle, die aktiviert oder moduliert werden durch Photolyse von einigen der verfügbaren caged Verbindu...

<table border="1"><tbody><tr><th> Ausrüstung </th><th> Firma </th><th> Katalog-Nummer </th><th> Kommentare </th></tr><tr><td> Adapter-Modul Flash-Lampe Mikroskop </td><td> Rapp Optoelectronic </td><td> Blitzwürfel 70 </td><td></td></tr><tr><td> Air Tisch </td><td> TMC </td><td> MICRO-g 63-534 </td><td></td></tr><tr><td> Digitizer </td><td> Axon Instruments </td><td> Digidata 1322a </td><td></td></tr><tr><td> Data Acquisition Software </td><td> Axon Instruments </td><td> pClamp 8 </td><td></td></tr><tr><td> Data Analysis Software </td><td> WaveMetrics </td><td> Igor </td><td></td></tr><tr><td> Spiegel für Adapter-Modul </td><td> Rapp Optoelectronic </td><td> M70/100 </td><td></td></tr><tr><td> Elektrodenhalter </td><td> Axon Instruments </td><td> 1-HL-U </td><td></td></tr><tr><td> Faraday-Käfig </td><td> Custom made </td><td></td><td></td></tr><tr><td> Filter-Würfel </td><td> Olymp </td><td> U-MWU </td><td> Excitation Filter entfernt </td></tr><tr><td> Flash-Lampe </td><td> Rapp Optoelectronic </td><td> JML-C2 </td><td></td></tr><tr><td> Pinzetten Dumont Nr. 55 </td><td> World Precision Instruments </td><td> 14099 </td><td></td></tr><tr><td> Glaskapillaren </td><td> World Precision Instruments </td><td> PG10165-4 </td><td></td></tr><tr><td> Glasboden Gericht </td><td> World Precision Instruments </td><td> FD35-100 </td><td></td></tr><tr><td> Illuminator </td><td> Olymp </td><td> Highlight 3100 </td><td></td></tr><tr><td> Inverse Mikroskop </td><td> Olymp </td><td> IX70 </td><td></td></tr><tr><td> Mikromanipulatoren </td><td> Luigs &amp; Neumann </td><td> SM I </td><td></td></tr><tr><td> Micropipette Puller </td><td> Narishige </td><td> PP-830 </td><td></td></tr><tr><td> Monitor </td><td> HesaVision </td><td> MTB-01 </td><td></td></tr><tr><td> Neutralfilter </td><td> Omega Optical </td><td> variiert </td><td></td></tr><tr><td> Ziel 100X </td><td> Zeiss </td><td> Fluar 440285 </td><td> Entweder Zeiss oder Olympus </td></tr><tr><td> Ziel 100X </td><td> Olymp </td><td> UPLFLN 100XOI2 </td><td> Entweder Zeiss oder Olympus </td></tr><tr><td> Optische UV Kurzpassfilter </td><td> Rapp Optoelectronic </td><td> SP400 </td><td></td></tr><tr><td> Patch-Clamp-Verstärkers </td><td> Axon Instruments </td><td> Axopatch 200B </td><td></td></tr><tr><td> Photo Diode Assembly </td><td> Rapp Optoelectronic </td><td> PDA </td><td></td></tr><tr><td> Quartz-Lichtleiter </td><td> Rapp Optoelectronic </td><td> variiert </td><td> Wir verwenden 600 pm Durchmesser </td></tr><tr><td> Silberdraht </td><td> World Precision Instruments </td><td> AGT1025 </td><td></td></tr><tr><td> Silber Boden Pellet </td><td> Warner Instrumente </td><td> 64-1309 </td><td></td></tr><tr><td> Xenon-Bogenlampe </td><td> Rapp Optoelectronic </td><td> XBL-JML </td><td></td></tr></tbody></table> <table border="1"><tbody><tr><th> Reagens </th><th> Firma </th><th> Katalog-Nummer </th></tr><tr><td> BCMCM-caged cAMP </td><td> BioLog </td><td> B016 </td></tr><tr><td> Rinderserumalbumin (BSA) </td><td> Sigma </td><td> A8806 </td></tr><tr><td> CaCl2-Standard-Lösung 0,1 M </td><td> Fluka </td><td> 21059 </td></tr><tr><td> Caged Ca: DMNP-EDTA </td><td> Invitrogen </td><td> D6814 </td></tr><tr><td> Cystein </td><td> Sigma </td><td> C9768 </td></tr><tr><td> Concanavalin A-Typ V (ConA) </td><td> Sigma </td><td> C7275 </td></tr><tr><td> CsCl </td><td> Sigma </td><td> C4036 </td></tr><tr><td> DMSO </td><td> Sigma </td><td> D8418 </td></tr><tr><td> DNAse I </td><td> Sigma </td><td> D4527 </td></tr><tr><td> EDTA </td><td> Sigma </td><td> E9884 </td></tr><tr><td> EGTA </td><td> Sigma </td><td> E4378 </td></tr><tr><td> Glucose </td><td> Sigma </td><td> G5767 </td></tr><tr><td> HEPES </td><td> Sigma </td><td> H3375 </td></tr><tr><td> KCl </td><td> Sigma </td><td> P3911 </td></tr><tr><td> KOH </td><td> Sigma </td><td> P1767 </td></tr><tr><td> Leupeptin </td><td> Sigma </td><td> L0649 </td></tr><tr><td> MgCl2 </td><td> Fluka </td><td> 63020 </td></tr><tr><td> Papain </td><td> Sigma </td><td> P3125 </td></tr><tr><td> Poly-L-Lysin </td><td> Sigma </td><td> P1274 </td></tr><tr><td> NaCl </td><td> Sigma </td><td> S9888 </td></tr><tr><td> NaOH </td><td> Sigma </td><td> S5881 </td></tr><tr><td> NaPyruvate </td><td> Sigma </td><td> P2256 </td></tr></tbody></table>

flash photolysis of caged compounds in the cilia of olfactory sensory  neurons

Photolyse von caged-Verbindungen ermöglicht die Produktion von schnellen und lokalisierten Anstieg der Konzentration von verschiedenen physiologisch aktive Verbindungen 1. Caged-Verbindungen sind Moleküle aus physiologisch inaktiv durch eine chemische Käfig, der durch einen Blitz von UV-Licht gebrochen werden kann. Hier zeigen wir, wie Patch-Clamp-Aufnahmen mit der Photolyse von caged-Verbindungen für das Studium der olfaktorischen Wahrnehmung in dissoziierten Maus Riechzellen kombiniert zu erhalten. Der Prozess der olfaktorischen Wahrnehmung (Abbildung 1) erfolgt in den Cilien von Riechzellen, wo Geruchsstoff Bindung an Rezeptoren führt zu einer Erhöhung der cAMP, dass zyklische Nukleotid-gesteuerte (CNG)-Kanäle 2 öffnet. Ca Eintrag durch CNG-Kanäle aktiviert Ca-aktivierte Cl-Kanäle. Wir zeigen, wie die Neuronen aus dem olfaktorischen Epithel der Maus 3 und wie CNG-Kanäle oder Ca-aktivierte Cl-Kanäle durch Photolyse von caged cAMP 4 oder caged Ca 5 aktivieren distanzieren &lt;/ Sup&gt;. Wir verwenden eine Blitzlampe 6,7 gegenüber UV-Blitze, die Ciliargegend gelten Uncage cAMP oder Ca während Patch-Clamp-Aufnahmen getroffen werden, um den Strom in der whole-cell Voltage-Clamp-Konfiguration 8-11 messen.

Watch this Scientific Journal Video about Flash Photolysis of Caged Compounds in the Cilia of Olfactory Sensory  Neurons at JoVE.com

Flash Photolysis of Caged Compounds in the Cilia of Olfactory Sensory  Neurons

Photolyse von caged-Verbindungen ermöglicht die Produktion von schnellen und lokalisierten Anstieg der Konzentration von verschiedenen physiologisch aktiven Verbindungen. Hier zeigen wir, wie Patch-Clamp-Aufnahmen mit der Photolyse von caged cAMP kombiniert oder caged Ca für das Studium der olfaktorischen Wahrnehmung in dissoziierten Maus Riechzellen erhalten.

Flash-Photolyse von Caged Compounds in den Zilien der Riechzellen

Photolysis of caged compounds allows the production of rapid and localized increases in the concentration of various physiologically active compounds1. ...

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