Name: local application of drugs to study nicotinic acetylcholine receptor function in mouse brain slices
Uploaded: 2012-10-29T14:00:00
Description: Watch this Scientific Journal Video about Local Application of Drugs to Study Nicotinic Acetylcholine Receptor Function in Mouse Brain Slices at JoVE.com

यह काम राष्ट्रीय स्वास्थ्य संस्थान (एनआईएच) DA030396 अनुदान द्वारा समर्थित किया गया. Drenan प्रयोगशाला के सदस्यों के लिए उपयोगी चर्चा और पांडुलिपि की आलोचना के लिए धन्यवाद. एम आई के लिए विशेष धन्यवाद तकनीकी सहायता और जोनाथन थॉमस टिंग के लिए वयस्क माउस मस्तिष्क स्लाइस के संबंध में सलाह के लिए किम भाग गया.

1. मस्तिष्क टुकड़ा तैयारी और इलैक्ट्रोफिजियोलॉजी के लिए समाधान की तैयारी <ol><li> मस्तिष्क स्लाइस की तैयारी के लिए समाधान पहले 3, 4 में वर्णित किया गया है. 93 डी एन मिथाइल - glucamine, KCl 2.5, 1.2 नाह 2 4 पीओ, 30 3 NaHCO, 20 HE...

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M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subcellular+trafficking,+pentameric+assembly,+and+subunit+stoichiometry+of+neuronal+nicotinic+acetylcholine+receptors+containing+fluorescently+labeled+&#945;6+and+&#946;3+subunits.">Subcellular trafficking, pentameric assembly, and subunit stoichiometry of neuronal nicotinic acetylcholine receptors containing fluorescently labeled &#945;6 and &#946;3 subunits.</a> Mol. Pharmacol. 73, 27-41 (2008).</li><li>Keath, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+modulation+by+nicotine+of+substantia+nigra+versus+ventral+tegmental+area+dopamine+neurons.">Differential modulation by nicotine of substantia nigra versus ventral tegmental area dopamine neurons.</a> J. Neurophysiol. 98, 3388-3396 (2007).</li><li>Mansvelder, H. 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B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reliability+in+the+identification+of+midbrain+dopamine+neurons.">Reliability in the identification of midbrain dopamine neurons.</a> PLoS One. 5, e15222 (2010).</li><li>Margolis, E. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+ventral+tegmental+area+revisited:+is+there+an+electrophysiological+marker+for+dopaminergic+neurons.">The ventral tegmental area revisited: is there an electrophysiological marker for dopaminergic neurons.</a> J. Physiol. 577 (Pt 3), 907-924 (2006).</li><li>Couey, J. 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मोटे तौर पर इस अखबार में प्रस्तुत पद्धति मस्तिष्क टुकड़ा तैयारी में ligand-gated आयन चैनल समारोह के अध्ययन के लिए उपयोगी है. हालांकि, वहाँ कारकों की एक संख्या है कि काफी प्रयोगात्मक परिणाम है कि इस पद्धति का उ?...

<table border="1"&gt;<tbody&gt;<tr&gt;<td<strong&gt; अभिकर्मक के नाम</strong</td&gt;<td<strong&gt; कंपनी</strong</td&gt;<td<strong&gt; सूचीपत्र संख्या</strong</td&gt;</tr&gt;<tr&gt;<td&gt; एन मिथाइल D-glucamine</td&gt;<td&gt; सिग्मा</td&gt;<tdM2004&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; KCl</td&gt;<td&gt; सिग्मा</td&gt;<tdP3911&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; नाह<sub&gt; 2</sub&gt; पीओ<sub&gt; 4</sub</td&gt;<td&gt; सिग्मा</td&gt;<tdS9638&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; NaHCO<sub&gt; 3</sub</td&gt;<td&gt; सिग्मा</td&gt;<tdS6014&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; HEPES</td&gt;<td&gt; सिग्मा</td&gt;<tdH3375&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; ग्लूकोज</td&gt;<td&gt; सिग्मा</td&gt;<tdG5767&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; ना<sup&gt; +</sup&gt; एस्कॉर्बेट</td&gt;<td&gt; सिग्मा</td&gt;<tdA4034&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; Thiourea</td&gt;<td&gt; सिग्मा</td&gt;<tdT8656&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; ना<sup&gt; +</sup&gt; पाइरूवेट</td&gt;<td&gt; सिग्मा</td&gt;<tdP2256&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; MgSO<sub&gt; 4</sub&gt; 7 •<sub&gt; 2</sub&gt; हे</td&gt;<td&gt; सिग्मा</td&gt;<td230391&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; CACL<sub&gt; 2</sub&gt; • 2H<sub&gt; 2</sub&gt; 0</td&gt;<td&gt; सिग्मा</td&gt;<td223506&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; NaCl</td&gt;<td&gt; सिग्मा</td&gt;<tdS9625&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; ना<sup&gt; +</supPentobarbital&gt;</td&gt;<td&gt; Vortech फार्मास्यूटिकल्स</td&gt;<td76351315&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; पोटेशियम gluconate</td&gt;<td&gt; सिग्मा</td&gt;<tdG4500&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; EGTA</td&gt;<td&gt; सिग्मा</td&gt;<tdE3889&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; मिलीग्राम एटीपी</td&gt;<td&gt; सिग्मा</td&gt;<tdA9187&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; GTP</td&gt;<td&gt; सिग्मा</td&gt;<tdG8877&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; DSK शून्य slicer 1 हिल</td&gt;<td&gt; टेड पेला इंक,</td&gt;<td</td&gt;</tr&gt;<tr&gt;<tdP&gt;-97 ज्वलंत / ब्राउन micropipette डांड़ी</td&gt;<td&gt; Sutter</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; आर सी-27 रिकॉर्डिंग कक्ष</td&gt;<td&gt; वार्नर</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; परफ्यूज़न TC-344B हीटर नियंत्रक</td&gt;<td&gt; वार्नर</td&gt;<td640101&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; एसएच 27B समाधान हीटर</td&gt;<td&gt; वार्नर</td&gt;<td640102&gt;</td&gt;</tr&gt;<tr&gt;<td&gt; Nikon FN-1</td&gt;<td&gt; Nikon</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; C-7500 सीसीडी वीडियो कैमरा</td&gt;<td&gt; हमामात्सू</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; Picospritzer III</td&gt;<td&gt; जनरल वाल्व कं</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; सांसद-285 micromanipulator</td&gt;<td&gt; Sutter</td&gt;<td</td&gt;</tr&gt;<tr&gt;<tdPA-100&gt; Piezoelectric अनुवादक</td&gt;<td&gt; Piezosystem जेना इंक,</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; 12V40 piezo प्रवर्धक</td&gt;<td&gt; Piezosystem जेना इंक,</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; Axopatch 200B</td&gt;<td&gt; आणविक उपकरणों कॉर्प</td&gt;<td</td&gt;</tr&gt;<tr&gt;<td&gt; Digidata 1440A</td&gt;<td&gt; आणविक उपकरणों कॉर्प</td&gt;<td</td&gt;</tr&gt;</tbody&gt;</table

local application of drugs to study nicotinic acetylcholine receptor function in mouse brain slices

Tobacco use leads to numerous health problems, including cancer, heart disease, emphysema, and stroke. Addiction to cigarette smoking is a prevalent neuropsychiatric disorder that stems from the biophysical and cellular actions of nicotine on nicotinic acetylcholine receptors (nAChRs) throughout the central nervous system. Understanding the various nAChR subtypes that exist in brain areas relevant to nicotine addiction is a major priority.
Experiments that employ electrophysiology techniques such as whole-cell patch clamp or two-electrode voltage clamp recordings are useful for pharmacological characterization of nAChRs of interest. Cells expressing nAChRs, such as mammalian tissue culture cells or Xenopus laevis oocytes, are physically isolated and are therefore easily studied using the tools of modern pharmacology. Much progress has been made using these techniques, particularly when the target receptor was already known and ectopic expression was easily achieved. Often, however, it is necessary to study nAChRs in their native environment: in neurons within brain slices acutely harvested from laboratory mice or rats. For example, mice expressing "hypersensitive" nAChR subunits such as &alpha;4 L9&prime;A mice 1 and &alpha;6 L9&prime;S mice 2, allow for unambiguous identification of neurons based on their functional expression of a specific nAChR subunit. Although whole-cell patch clamp recordings from neurons in brain slices is routinely done by the skilled electrophysiologist, it is challenging to locally apply drugs such as acetylcholine or nicotine to the recorded cell within a brain slice. Dilution of drugs into the superfusate (bath application) is not rapidly reversible, and U-tube systems are not easily adapted to work with brain slices.
In this paper, we describe a method for rapidly applying nAChR-activating drugs to neurons recorded in adult mouse brain slices. Standard whole-cell recordings are made from neurons in slices, and a second micropipette filled with a drug of interest is maneuvered into position near the recorded cell. An injection of pressurized air or inert nitrogen into the drug-filled pipette causes a small amount of drug solution to be ejected from the pipette onto the recorded cell. Using this method, nAChR-mediated currents are able to be resolved with millisecond accuracy. Drug application times can easily be varied, and the drug-filled pipette can be retracted and replaced with a new pipette, allowing for concentration-response curves to be created for a single neuron. Although described in the context of nAChR neurobiology, this technique should be useful for studying many types of ligand-gated ion channels or receptors in neurons from brain slices.

हमारे प्रयोगों में, हम नियमित रूप से dopamine (डीए) उत्पादन वेंट्रल टेगमेंटल (VTA) क्षेत्र और सब्सटेंशिया निग्रा पार्स कॉम्पेक्टा (एसएनसी) के न्यूरॉन्स से रिकॉर्ड. , वोल्टेज दबाना मोड में acetylcholine या इन कोशिकाओं को...

Watch this Scientific Journal Video about Local Application of Drugs to Study Nicotinic Acetylcholine Receptor Function in Mouse Brain Slices at JoVE.com

Local Application of Drugs to Study Nicotinic Acetylcholine Receptor Function in Mouse Brain Slices

इस पत्र में, हम ligand-gated आयन चैनल समारोह तीव्रता पृथक मस्तिष्क स्लाइस की न्यूरॉन्स में अध्ययन करने के लिए एक उपयोगी विधि का वर्णन है. इस विधि के मानक पैच दबाना तकनीक का उपयोग कर दर्ज न्यूरॉन्स के लिए दवाओं की स्थानीय आवेदन के लिए एक दवा से भरे micropipette का इस्तेमाल शामिल है.

ड्रग्स के स्थानीय आवेदन करने के लिए माउस मस्तिष्क स्लाइसें में निकोटिनिक acetylcholine रिसेप्टर समारोह अध्ययन

Tobacco use leads to numerous health problems, including cancer, heart disease, emphysema, and stroke. Addiction to cigarette smoking is a prevalent ...

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Drugs Acting on Autonomic Nervous System: Cholinergic Agonists and Antagonists Agents

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Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) ...

Probing Nicotinic Acetylcholine Receptor Function in Mouse Brain Slices via Laser Flash Photolysis of Photoactivatable Nicotine

Photoactivatable निकोटीन के लेजर फ्लैश Photolysis के माध्यम से माउस मस्तिष्क स्लाइस में कोर्टेक्स Acetylcholine रिसेप्टर समारोह की जांच

Acetylcholine (ACh) acts through receptors to modulate a variety of neuronal processes, but it has been challenging to link ACh receptor function with ...

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset

जीर्ण-cortical Electrocorticographic सरणी के आम Marmoset में पुरानी आरोपण

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent ...