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>
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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 fotolyse van gekooide verbindingen in combinatie met patch-clamp opnames is een handige techniek om een ​​snelle en lokale sprongen te krijgen in de concentratie van fysiologisch actieve moleculen binnen en buiten cellen. Verschillende soorten gekooide compounds1 zijn gesynthetiseerd, en deze techniek kan worden toegepast op verschillende soorten cellen, met inbegrip van gekweekte cellen die ion kanalen die kunnen worden geactiveerd of gemoduleerd door fotolyse van enkele van de beschikbare gekooide verbinding...

<table border="1"><tbody><tr><th> Uitrusting </th><th> Vennootschap </th><th> Catalogus nummer </th><th> Reacties </th></tr><tr><td> Adapter module flitslamp aan microscoop </td><td> Rapp Opto-elektronische </td><td> FlashCube 70 </td><td></td></tr><tr><td> Lucht tafel </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 voor adapter module </td><td> Rapp Opto-elektronische </td><td> M70/100 </td><td></td></tr><tr><td> Elektrode houder </td><td> Axon Instruments </td><td> 1-HL-U </td><td></td></tr><tr><td> Faraday-kooi </td><td> Op maat gemaakt </td><td></td><td></td></tr><tr><td> Filter kubus </td><td> Olympus </td><td> U-MWU </td><td> Excitatiefilter verwijderd </td></tr><tr><td> Flitslamp </td><td> Rapp Opto-elektronische </td><td> JML-C2 </td><td></td></tr><tr><td> Pincet Dumont # 55 </td><td> Wereld precisie-instrumenten </td><td> 14099 </td><td></td></tr><tr><td> Glazen capillairen </td><td> Wereld precisie-instrumenten </td><td> PG10165-4 </td><td></td></tr><tr><td> Glazen bodem schaal </td><td> Wereld precisie-instrumenten </td><td> FD35-100 </td><td></td></tr><tr><td> Illuminator </td><td> Olympus </td><td> Highlight 3100 </td><td></td></tr><tr><td> Omkeermicroscoop </td><td> Olympus </td><td> IX70 </td><td></td></tr><tr><td> Micromanipulators </td><td> Luigs &amp; Neumann </td><td> SM I </td><td></td></tr><tr><td> Micropipet 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> Grijsfilters </td><td> Omega Optical </td><td> varieert </td><td></td></tr><tr><td> Doelstelling 100X </td><td> Zeiss </td><td> Fluar 440285 </td><td> Ofwel Zeiss of Olympus </td></tr><tr><td> Doelstelling 100X </td><td> Olympus </td><td> UPLFLN 100XOI2 </td><td> Ofwel Zeiss of Olympus </td></tr><tr><td> Optisch UV-filter shortpass </td><td> Rapp Opto-elektronische </td><td> SP400 </td><td></td></tr><tr><td> Patch-clamp versterker </td><td> Axon Instruments </td><td> Axopatch 200B </td><td></td></tr><tr><td> Foto Diode Assembly </td><td> Rapp Opto-elektronische </td><td> PDA </td><td></td></tr><tr><td> Kwarts lichtgeleider </td><td> Rapp Opto-elektronische </td><td> varieert </td><td> We maken gebruik van 600 micrometer diameter </td></tr><tr><td> Zilverdraad </td><td> Wereld precisie-instrumenten </td><td> AGT1025 </td><td></td></tr><tr><td> Zilver grond pellet </td><td> Warner instrumenten </td><td> 64-1309 </td><td></td></tr><tr><td> Xenon booglamp </td><td> Rapp Opto-elektronische </td><td> XBL-JML </td><td></td></tr></tbody></table> <table border="1"><tbody><tr><th> Reagens </th><th> Vennootschap </th><th> Catalogus nummer </th></tr><tr><td> BCMCM-kooi cAMP </td><td> Biolog </td><td> B016 </td></tr><tr><td> Bovine serum albumine (BSA) </td><td> Sigma </td><td> A8806 </td></tr><tr><td> CaCl2 standaard oplossing 0,1 M </td><td> Fluka </td><td> 21059 </td></tr><tr><td> Gekooide Ca: DMNP-EDTA </td><td> Invitrogen </td><td> D6814 </td></tr><tr><td> Cysteine </td><td> Sigma </td><td> C9768 </td></tr><tr><td> Concanavaline Een type 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> Papaïne </td><td> Sigma </td><td> P3125 </td></tr><tr><td> Poly-L-lysine </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

Fotolyse van gekooide verbindingen maakt de productie van snelle en gelokaliseerde toename van de concentratie van diverse fysiologisch actieve verbindingen 1. Gekooide verbindingen zijn moleculen gemaakt fysiologisch inactief door een chemische kooi die kan worden doorbroken door een flits van ultraviolet licht. Hier laten we zien hoe de patch-clamp opnames gecombineerd met fotolyse van gekooide verbindingen voor de studie van olfactorische transductie in gedissocieerde muis olfactorische sensorische neuronen te verkrijgen. Het proces van olfactorische transductie (figuur 1) vindt plaats in de cilia van olfactorische sensorische neuronen, waar odorant binding aan receptoren leidt tot de toename van cAMP dat cyclische nucleotide-gated (CNG) kanaal 2 wordt geopend. Ca ingang door CNG kanalen activeert Ca-geactiveerde Cl kanalen. We laten zien hoe de neuronen distantiëren van de muis olfactorische epitheel 3 en hoe CNG kanalen of Ca-geactiveerde Cl-kanalen activeren door fotolyse van gekooide cAMP 4 of gekooid Ca 5 &lt;/ Sup&gt;. We maken gebruik van een flitslamp 6,7 aan ultraviolette flitsen van toepassing op de ciliaire regio vrij maken cAMP of Ca, terwijl patch-clamp opnames worden genomen om de stroom te meten in de whole-cell voltage-clamp configuratie 8-11.

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

Fotolyse van gekooide verbindingen maakt de productie van snelle en gelokaliseerde toename van de concentratie van diverse fysiologisch actieve verbindingen. Hier laten we zien hoe de patch-clamp opnames gecombineerd met fotolyse van gekooide cAMP of gekooid Ca voor de studie van olfactorische transductie in gedissocieerde muis olfactorische sensorische neuronen te verkrijgen.

Flash fotolyse van Gekooide verbindingen in de Cilia van olfactorische sensorische neuronen

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

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