Name: analyzing responses of mouse olfactory sensory neurons using the air-phase electroolfactogram recording
Uploaded: 2010-03-02T17:57:00
Duration: 9 min 51 s
Description: Watch this Scientific Journal Video about Analyzing Responses of Mouse Olfactory Sensory Neurons Using the Air-phase Electroolfactogram Recording at JoVE.com

हम डॉ. Yijun गाने, और Hattar Kuruvilla झाओ त्रिकोणीय जीवविज्ञान विभाग, सलाह और मदद के लिए जॉन्स हॉपकिन्स विश्वविद्यालय के प्रयोगशाला के सदस्यों को धन्यवाद देता हूं. एनआईएच DC007395 और DC009946 अनुदान द्वारा समर्थित है.

<ol>
	<li>Ottoson, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+the+electrical+activity+of+the+olfactory+epithelium.">Analysis of the electrical activity of the olfactory epithelium.</a> Acta Physiologica Scandinavica. 35, 1-83 (1956).</li><li>Scott, J. W., Scott-Johnson, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+electroolfactogram:+a+review+of+its+history+and+uses.">The electroolfactogram: a review of its history and uses.</a> Microsc Res Tech. 58, 152-160 (2002).</li><li>Brunet, L. J., Gold, G. H., Ngai, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=General+anosmia+caused+by+a+targeted+disruption+of+the+mouse+olfactory+cyclic+nucleotide-gated+cation+channel.">General anosmia caused by a targeted disruption of the mouse olfactory cyclic nucleotide-gated cation channel.</a> Neuron. 17, 681-693 (1996).</li><li>Belluscio, L., Gold, G. H., Nemes, A., Axel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mice+deficient+in+G(olf)+are+anosmic.">Mice deficient in G(olf) are anosmic.</a> Neuron. 20, 69-81 (1998).</li><li>Zhao, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+expression+of+a+mammalian+odorant+receptor.">Functional expression of a mammalian odorant receptor.</a> Science. 279, 237-242 (1998).</li><li>Wong, S. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Disruption+of+the+type+III+adenylyl+cyclase+gene+leads+to+peripheral+and+behavioral+anosmia+in+transgenic+mice.">Disruption of the type III adenylyl cyclase gene leads to peripheral and behavioral anosmia in transgenic mice.</a> Neuron. 27, 487-497 (2000).</li><li>Munger, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+role+of+the+CNGA4+channel+subunit+in+Ca2+-calmodulin-dependent+odor+adaptation.">Central role of the CNGA4 channel subunit in Ca2+-calmodulin-dependent odor adaptation.</a> Science. 294, 2172-2175 (2001).</li><li>Michalakis, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Loss+of+CNGB1+protein+leads+to+olfactory+dysfunction+and+subciliary+cyclic+nucleotide-gated+channel+trapping.">Loss of CNGB1 protein leads to olfactory dysfunction and subciliary cyclic nucleotide-gated channel trapping.</a> J Biol Chem. 281, 35156-35166 (2006).</li><li>Buiakova, O. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+marker+protein+(OMP)+gene+deletion+causes+altered+physiological+activity+of+olfactory+sensory+neurons.">Olfactory marker protein (OMP) gene deletion causes altered physiological activity of olfactory sensory neurons.</a> Proc Natl Acad Sci U S A. 93, 9858-9863 (1996).</li><li>Nickell, W. T., Kleene, N. K., Gesteland, R. C., Kleene, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuronal+chloride+accumulation+in+olfactory+epithelium+of+mice+lacking+NKCC1.">Neuronal chloride accumulation in olfactory epithelium of mice lacking NKCC1.</a> J Neurophysiol. 95, 2003-2006 (2006).</li><li>Song, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+CNG+channel+desensitization+by+Ca2+/CaM+via+the+B1b+subunit+affects+response+termination+but+not+sensitivity+to+recurring+stimulation.">Olfactory CNG channel desensitization by Ca2+/CaM via the B1b subunit affects response termination but not sensitivity to recurring stimulation.</a> Neuron. 58, 374-386 (2008).</li><li>Cygnar, K. D., Zhao, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphodiesterase+1C+is+dispensable+for+rapid+response+termination+of+olfactory+sensory+neurons.">Phosphodiesterase 1C is dispensable for rapid response termination of olfactory sensory neurons.</a> Nat Neurosci. 12, 454-462 (2009).</li><li>Kleene, S. 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+electrochemical+basis+of+odor+transduction+in+vertebrate+olfactory+cilia.">The electrochemical basis of odor transduction in vertebrate olfactory cilia.</a> Chem Senses. 33, 839-859 (2008).</li><li>Chen, S., Lane, A. P., Bock, R., Leinders-Zufall, T., Zufall, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blocking+adenylyl+cyclase+inhibits+olfactory+generator+currents+induced+by+"IP(3)-odors".">Blocking adenylyl cyclase inhibits olfactory generator currents induced by "IP(3)-odors".</a> J Neurophysiol. 84, 575-5780 (2000).</li><li>Pinato, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electroolfactogram+responses+from+organotypic+cultures+of+the+olfactory+epithelium+from+postnatal+mice.">Electroolfactogram responses from organotypic cultures of the olfactory epithelium from postnatal mice.</a> Chem Senses. 33, 397-404 (2008).</li><li>Ezeh, P. I., Davis, L. M., Scott, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regional+distribution+of+rat+electroolfactogram.">Regional distribution of rat electroolfactogram.</a> J Neurophysiol. 73, 2207-2220 (1995).</li><li>Scott-Johnson, P. E., Blakley, D., Scott, J. W. <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+air+flow+on+rat+electroolfactogram.">Effects of air flow on rat electroolfactogram.</a> Chem Senses. 25, 761-768 (2000).</li></ol>

चूहे को संभाला और पशु और जॉन्स हॉपकिन्स विश्वविद्यालय की देखभाल का प्रयोग करें समितियों द्वारा अनुमोदित विधियों के साथ euthanized थे.

इस प्रोटोकॉल में वर्णित सेटअप के साथ, घ्राण उपकला की सतह पर odorant उत्तेजनाओं ऊतक की तैयारी के बीच लगातार हो सकता है, जंगली और प्रकार उत्परिवर्ती चूहों के बीच तुलना के लिए अनुमति देता है, भले ही सटीक odorant एकाग्रता और गतिशीलता से अनजान हैं. कई कारकों, विशेष रूप से रिकॉर्डिंग स्थान और humidified हवा के प्रवाह की दर, सभी छवियाँ में बदलाव के कारण. केयर वही turbinate पर इसी तरह की स्थिति से रिकॉर्ड करने के लिए भिन्नता को कम से कम करने के लिए लिया जाना चाहिए. यह आसानी से लगातार सिर के एक ही पक्ष से रिकॉर्डिंग और माइक्रोस्कोप, गंध वितरण ट्यूब, और ऊतकों के नमूनों के बीच हवा अपरिवर्तित तालिका पर micromanipulators के पदचिह्न रखने के द्वारा प्राप्त किया जा सकता है. इसके अलावा, ऊतकों के नमूनों तुरंत humidified हवा की धारा में रखा जाना चाहिए विच्छेदन के बाद ऊतक के अत्यधिक सुखाने को रोकने के लिए. चूहों पर सभी छवियाँ रिकॉर्डिंग भी तैयार माउस पर एक तरल छिड़काव तंत्र के साथ बाहर कर सकते हैं किया जा 7, 14, 15, या सिर बरकरार छोड़ने और turbinates 16, 17 के ऊपर एक छोटे से छेद drilled में इलेक्ट्रोड डालने से turbinates. सभी छवियाँ रिकॉर्डिंग के प्रत्येक बदलाव की अपनी ताकत है: ऊतक के रूप में इस प्रोटोकॉल में वर्णित तैयारी पर हवा चरण रिकॉर्डिंग सेटअप की एक न्यूनतम राशि की आवश्यकता होती है और आसान आचरण रहे हैं, एक तरल छिड़काव तंत्र का उपयोग करते हुए रिकॉर्डिंग औषधीय अभिकर्मकों के उपयोग की सुविधा है, हालांकि कई odorants के हाइड्रोफोबिक प्रकृति गंध डिलीवरी पेचीदा हो, अंत में, रिकॉर्डिंग में जो सिर बरकरार रह गया है, &#39;कृत्रिम सूंघ&#39; प्रयोगों में इस्तेमाल किया जा सकता है, हालांकि इलेक्ट्रोड प्लेसमेंट जब turbinates पूरी तरह से उजागर कर रहे हैं की तुलना में अधिक मुश्किल है.

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Air delivery tube</td>
<td>equipment</td>
<td>Custom fabricated</td>
<td>&nbsp;</td>
<td>The barrel of a 1-mL syringe with a T-fitting can be used as a substitute</td>
</tr>
<tr>
<td>Air table</td>
<td>equipment</td>
<td>Newport</td>
<td>LW3030B-OPT</td>
<td>&nbsp;</td>
<tr>
<td>Amplifier</td>
<td>equipment</td>
<td>Warner</td>
<td>DP-301</td>
<td>&nbsp;</td>
<tr>
<td>Computer and Data Acquisition Software</td>
<td>equipment</td>
<td>Axograph 4.9.2 on Apple Macintosh</td>
<td>&nbsp;</td>
<td>Updated versions of Axograph for Mac OS X and Windows are available from http://axographx.com/.</td>
</tr>
<tr>
<td>Butane torch</td>
<td>equipment</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>A cr&egrave;me br&ucirc;l&egrave;e torch works well</td>
</tr>
<tr>
<td>Digitizer</td>
<td>equipment</td>
<td>Axon Instruments</td>
<td>Digidata 1322A</td>
<td>&nbsp;</td>
<tr>
<td>Dissecting Scope</td>
<td>equipment</td>
<td>Scienscope</td>
<td>SSZ</td>
<td>&nbsp;</td>
<tr>
<td>Electrode holder</td>
<td>equipment</td>
<td>Harvard Apparatus</td>
<td>64-1021</td>
<td>&nbsp;</td>
<tr>
<td>Magnetic Holding Devices (12 mm)</td>
<td>equipment</td>
<td>World Precision Instruments</td>
<td>M10</td>
<td>&nbsp;</td>
<tr>
<td>Micromanipulators</td>
<td>equipment</td>
<td>World Precision Instruments</td>
<td>M3301R M3301L</td>
<td>&nbsp;</td>
<tr>
<td>Micropipette Puller</td>
<td>equipment</td>
<td>Sutter Instrument Co.</td>
<td>P2000</td>
<td>&nbsp;</td>
<tr>
<td>Oscilloscope</td>
<td>equipment</td>
<td>Tektronix </td>
<td>5110</td>
<td>&nbsp;</td>
<tr>
<td>Picospritzer III</td>
<td>equipment</td>
<td>Parker Instrumentation</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Silicone tubing</td>
<td>equipment</td>
<td>Nalge Nunc</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Specimen stage</td>
<td>equipment</td>
<td>Custom fabricated</td>
<td>&nbsp;</td>
<td>Any small solid object can be used to elevate the mounting dish. Immobilize the dish with modeling clay.</td>
</tr>
<tr>
<td>18 gage needles</td>
<td>material</td>
<td>Becton Dickinson</td>
<td>305195</td>
<td>&nbsp;</td>
<tr>
<td>2 oz. glass bottles</td>
<td>material</td>
<td>VWR International</td>
<td>16152-201</td>
<td>&nbsp;</td>
<tr>
<td>Glass capillaries</td>
<td>material</td>
<td>World Precision Instruments</td>
<td>TW150F-6</td>
<td>&nbsp;</td>
<tr>
<td>Silicone stoppers size 16D</td>
<td>material</td>
<td>Chemware</td>
<td>D1069809</td>
<td>&nbsp;</td>
<tr>
<td>Silver wire</td>
<td>material</td>
<td>World Precision Instruments</td>
<td>AGW1010</td>
<td>&nbsp;</td>
<tr>
<td>SylGuard 184</td>
<td>material</td>
<td>Dow Corning </td>
<td>SYLG184</td>
<td>From World Precision Instruments</td>
</tr>
<tr>
<td>Agarose</td>
<td>reagent</td>
<td>Invitrogen</td>
<td>15510-027</td>
<td>&nbsp;</td>
<tr>
<td>Amyl acetate</td>
<td>reagent</td>
<td>Aldrich</td>
<td>W504009</td>
<td>&nbsp;</td>
<tr>
<td>Calcium chloride (CaCl2)</td>
<td>reagent</td>
<td>Sigma</td>
<td>C-1016</td>
<td>&nbsp;</td>
<tr>
<td>Dimethyl sulfoxide (DMSO)</td>
<td>reagent</td>
<td>Sigma</td>
<td>D5879</td>
<td>&nbsp;</td>
<tr>
<td>HEPES</td>
<td>reagent</td>
<td>Fisher</td>
<td>BP310</td>
<td>&nbsp;</td>
<tr>
<td>Heptaldehyde</td>
<td>reagent</td>
<td>Aldrich</td>
<td>H2120</td>
<td>&nbsp;</td>
<tr>
<td>Magnesium chloride hexahydrate (MgCl2+6H2O)</td>
<td>reagent</td>
<td>Sigma</td>
<td>M9272</td>
<td>&nbsp;</td>
<tr>
<td>Sodium chloride (NaCl)</td>
<td>reagent</td>
<td>JT Baker</td>
<td>3624-05</td>
<td>&nbsp;</td>
<tr>
<td>flowmeter</td>
<td>equipment</td>
<td>Gilmont</td>
<td>GF-2260</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

analyzing responses of mouse olfactory sensory neurons using the air-phase electroolfactogram recording

Animals depend on olfaction for many critical behaviors, such as finding food sources, avoiding predators, and identifying conspecifics for mating and other social interactions. The electroolfactogram (EOG) recording is an informative, easy to conduct, and reliable method to assay olfactory function at the level of the olfactory epithelium. Since the 1956 description of the EOG by Ottoson in frogs1, the EOG recording has been applied in many vertebrates including salamanders, rabbits, rats, mice, and humans (reviewed by Scott and Scott-Johnson, 2002, ref. 2). The recent advances in genetic modification in mice have rekindled interest in recording the EOG for physiological characterization of olfactory function in knock-out and knock-in mice. EOG recordings have been successfully applied to demonstrate the central role of olfactory signal transduction components3-8, and more recently to characterize the contribution of certain regulatory mechanisms to OSN responses9-12.
Odorant detection occurs at the surface of the olfactory epithelium on the cilia of OSNs, where a signal transduction cascade leads to opening of ion channels, generating a current that flows into the cilia and depolarizes the membrane13. The EOG is the negative potential recorded extracellularly at the surface of the olfactory epithelium upon odorant stimulation, resulting from a summation of the potential changes caused by individual responsive OSNs in the recording field2. Comparison of the amplitude and kinetics of the EOG thus provide valuable information about how genetic modification and other experimental manipulations influence the molecular signaling underlying the OSN response to odor. 
Here we describe an air-phase EOG recording on a preparation of mouse olfactory turbinates. Briefly, after sacrificing the mouse, the olfactory turbinates are exposed by bisecting the head along the midline and removing the septum. The turbinate preparation is then placed in the recording setup, and a recording electrode is placed at the surface of the olfactory epithelium on one of the medial turbinates. A reference electrode is electrically connected to the tissue through a buffer solution. A continuous stream of humidified air is blown over the surface of the epithelium to keep it moist. The vapor of odorant solutions is puffed into the stream of humidified air to stimulate the epithelium. Responses are recorded and digitized for further analysis.

Watch this Scientific Journal Video about Analyzing Responses of Mouse Olfactory Sensory Neurons Using the Air-phase Electroolfactogram Recording at JoVE.com

Analyzing Responses of Mouse Olfactory Sensory Neurons Using the Air-phase Electroolfactogram Recording

electroolfactogram रिकॉर्डिंग (सभी छवियाँ) एक जानकारीपूर्ण, घ्राण उपकला के स्तर पर घ्राण समारोह का आकलन करने के आचरण करने के लिए आसान और विश्वसनीय तरीका है. यह एक रिकॉर्डिंग सेटअप, माउस ऊतक तैयारी, डेटा संग्रह, और बुनियादी डेटा विश्लेषण प्रोटोकॉल का वर्णन करता है.

माउस घ्राण संवेदी न्यूरॉन्स Electroolfactogram एयर चरण रिकॉर्डिंग का उपयोग विश्लेषण प्रतिक्रियाएँ

Animals depend on olfaction for many critical behaviors, such as finding food sources, avoiding predators, and identifying conspecifics for mating and ...

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Neuroscience

Lentivirus-mediated Genetic Manipulation and Visualization of Olfactory Sensory Neurons in vivo

Lentivirus-mediated Genetic Manipulation and Visualization of Olfactory Sensory Neurons in vivo

Lentivirus की मध्यस्थता जेनेटिक और घ्राण संवेदी न्यूरॉन्स के हेरफेर विज़ुअलाइज़ेशन Vivo में</em

Development of a precise olfactory circuit relies on accurate projection of olfactory sensory neuron (OSN) axons to their synaptic 
targets in the ...

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

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Preparation of an Awake Mouse for Recording Neural Responses and Injecting Tracers

एक जाग माउस के तंत्रिका प्रतिक्रियाएँ रिकॉर्डिंग और tracers इंजेक्शन के लिए तैयार

It is well known that anesthesia alters neural response properties in various regions of the brain.13.  In the auditory system, fundamental response ...

Odorant-induced Responses Recorded from Olfactory Receptor Neurons using the Suction Pipette Technique

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Animals sample the odorous environment around them through the chemosensory systems located in the nasal cavity. Chemosensory signals affect complex ...

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Intracellular रिकॉर्डिंग, संवेदी क्षेत्र मानचित्रण, और जोंक में संवर्धन के पहचाने गए न्यूरॉन्स, Hirudo medicinalis</i

The freshwater leech, Hirudo medicinalis, is a versatile model organism that has been used to address scientific questions in the fields of ...

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

में घ्राण रिसेप्टर न्यूरॉन्स के पूरा पर्वत immunolabeling<em&gt; ड्रोसोफिला</em&gt; एंटीना

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this ...

Recording Light-evoked Postsynaptic Responses in Neurons in Dark-adapted, Mouse Retinal Slice Preparations Using Patch Clamp Techniques

रिकॉर्डिंग पैच दबाना तकनीकों का प्रयोग काले अनुकूलित, माउस रेटिना टुकड़ा तैयारी में न्यूरॉन्स में में postsynaptic प्रतिक्रियाएँ प्रकाश पैदा

The retina is the gateway to the visual system. To understand visual signal processing mechanisms, we investigate retinal neural network functions. ...

Perforated Patch-clamp Recording of Mouse Olfactory Sensory Neurons in Intact Neuroepithelium: Functional Analysis of Neurons Expressing an Identified Odorant Receptor

बरकरार neuroepithelium में माउस घ्राण संवेदी न्यूरॉन्स के छिद्रित पैच दबाना रिकॉर्डिंग: एक की पहचान odorant रिसेप्टर व्यक्त न्यूरॉन्स की कार्यात्मक विश्लेषण

Analyzing the physiological responses of olfactory sensory neurons (OSN) when stimulated with specific ligands is critical to understand the basis of ...

Using Single Sensillum Recording to Detect Olfactory Neuron Responses of Bed Bugs to Semiochemicals

Semiochemicals करने के लिए बिस्तर कीड़े की घ्राण न्यूरॉन जवाब का पता लगाने के लिए एकल Sensillum रिकॉर्डिंग का उपयोग

The insect olfactory system plays an important role in detecting semiochemicals in the environment. In particular, the antennal sensilla which house ...

Olfactory Behaviors Assayed by Computer Tracking Of Drosophila in a Four-quadrant Olfactometer

Olfactory Behaviors Assayed by Computer Tracking Of Drosophila in a Four-quadrant Olfactometer

घ्राण व्यवहार कंप्यूटर पर नज़र रखने का यत्न से<em&gt; ड्रोसोफिला</em&gt; एक चार चतुर्थ भाग Olfactometer में

A key challenge in neurobiology is to understand how neural circuits function to guide appropriate animal behaviors. Drosophila melanogaster is an ...