USDA-NIFA/AFRI 2010-65105-20582、NSF 0918177、およびBedoukian研究の定款によってサポートされています。

<html> 1。昆虫<ol><li>へそorangeworm、Amyelois transitella（ウォーカー）（鱗翅目：メイガ科）フレズノ、カリフォルニア州のアーモンドの木から採取した成虫から発信された私たちの研究室でのコロニー。写真政権：私達の昆虫のコロニーは、± 2℃、75 ± 10％相対湿度、および午前16時08分H（暗い光）下で28℃（パーシバル社、IA）環境槽内に保持されます。 </li><li>新興蛾は、毎日収集性欲を刺激する、そしてプラスチックの箱（669 mLの弁当箱、13 × 13センチメートル、高さ、4.5センチ、ラバーメイド）に転送され、水に浸したペーパータオル（27.9、喉が渇いた超吸収X 27.9センチメートルの10層までで提供;セイフウェイ）。ボックスカバーは、空気循環を可能にするために穴あきです。この配置は、〜100％RHを提供します。 </li><li>電気生理学的測定のために我々は、高湿度下に保持されますが出現、後2〜4日の古い蛾を使用してください。 </li></ol> 2。電気生理学的準備<ol><li>蛾（男性または女性）が静かに大規模な（約2mm）を先端径を持つように上部から切断されたピペットチップ（200μL、米国サイエンティフィック株式会社）にプッシュされます。アンテナと頭の部分が先端から突出するまで蛾を穏やかに加湿ティッシュペーパーのスタブを持つもう一方の端を通してプッシュされます。 </li><li>露出頭は片目と公開される2つのアンテナのごく一部を残して不乾性粘土（Claytoon、バンアーケンインターナショナル、カリフォルニア州）で頭を覆うことによって固定化されている。この準備は、2つの電極ホルダーの間にEAGマニピュレーターMP - 12（Syntech、ドイツ）のプラットフォームに配置されます。 </li><li>私たちは、フィラメントと1.0ミリメートルホウケイ酸キャピラリーチューブから作られたガラス電極を使用して（WPI社、FL）とマイクロチップ未満の微小電極を得るためにモデルP - 97マイクロピペットプラー（サターインスツルメンツ、CA）でプルアップ。電極はバック感覚子リンパリンガー（Kaisslingとソーソン1980）で埋められます。一方の電極は、顕微鏡（ライカMZ75）の下で目の露出面積をimpalesと記録電極は、二つのアンテナを収納しながら、参照として機能します。接触を容易にするために、遠位1〜2触角のセグメントは、記録電極に挿入される前にカットされています。個々の感覚器からの​​記録信号の場合、我々はわずかに記録電極を顕微鏡（; 800 ×倍率オリンパスBX51WI）の下impaledされることを特徴この設定を変更します。活動電位は、感覚子の基部にガラス電極を挿入して記録されます。シグナルは、AUTOSPIKEソフトウェア（Syntech、ドイツ）と分析されるUB - IDACオフラインにハイインピーダンスプリアンプ（Syntech、ドイツ）と供給によって増幅されています。バンドは、100 -10000 Hzと活動電位との間でフィルタリング通過しているAC信号がノイズから彼らのピークツー谷の振幅を区別AUTOSPIKEアルゴリズムを使用してコンピュータによって抽出されます。個々のニューロンの応答は、自発的な周波数に対する活動電位の頻度の増加（または減少）（毎秒スパイク）として計算されます。 </li></ol> 3。化学的刺激と刺激のプロトコル<ol><li>最高純度の化学薬品が使用されます。へそのorangewormから女性フェロモンブレンドの主要成分、（Z、Z）-11,13 - hexadecadienalはBedoukianリサーチ社、CTから得られた。化学物質が希釈され、10μg/μLのと進法希釈のストック溶液を作るためにガラス蒸留ヘキサンとのW / Vは、作られています。ヘキサンに溶解した刺激の化学物質のアリコート（10μl）を溶媒としては、静かに換気フードの下に10秒間振とうすることにより蒸発させ、ストリップを5 mLのポリプロピレンに配置され、（8x40 mm）のワットマンろ紙片上にロードさ様々なボリュームが取り出されているからシリンジ（BDシリンジ、NJ）。ヘキサン単独で、空の注射器は、コントロールとして使用。 </li><li>準備がSyntech刺激コントローラ（CS - 55モデル、Syntech、ドイツ）が提供する加湿空気​​の流れで開催される20 ml /秒でそのための4mL /秒の刺激パルスは500ミリ秒のために追加されます。信号は、刺激パルスの開始前に2秒を開始、10秒間記録されます。触角の準備はどのCAの間に500 msのパルスで刺激されています。刺激を含む5mLのポリプロピレンシリンジから精製された空気の2mLをメイン空気の流れの上に追加されます。高応答、以下の少なくとも1分以上のギャップが、、刺激の間で許可される。 </li></ol> 4。代表的な結果控えめな蛾の触角（図1）女性のフェロモン成分で刺激、（Z、Z）-11,13 - hexadecadienal堅牢な用量依存性の応答（図2）を生成する。この主要なフェロモン成分はまた、単一ユニット記録（図3B）のトレースに表示される長いtrichodea感覚器（図3A）から用量依存性の興奮性応答を誘発する。 <img src="/files/ftp_upload/2489/2489fig1.jpg" alt="図1" /> 図1。ライヴへそorangeworm蛾抑制され、アンテナが露出しload/2489/2489fig2.jpg&quot;ALT =&quot;図2&quot;/&gt; 図2。用量依存性のEAG応答は（Z、Z）-11,13 - hexadecadienal、主要な女性のフェロモン成分に反応して誘発した。 <img src="/files/ftp_upload/2489/2489fig3.jpg" alt="図3" /> 男性のへそorangeworm蛾の図3。感覚器の毛髪状のは、用量依存的に（Z、Z）-11,13 - hexadecadienalに対応。 （A）オスへそorangeworm蛾のアンテナは、複数のセグメントであり、各セグメントは、構造体のような毛の数が多い、感覚器で飾られています。挿入図に詳細を示す走査型電子顕微鏡写真、（スケールバーはそれぞれ、200および50μMです）。毛状感覚子から（B）細胞外単一ユニット記録。

<ol>
	<li>Kaissling, K. -. E., Thorson, J., Sattelle, D. B., Hall, L. M., Hildebrand, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insect+olfactory+sensilla:+structural,+chemical+and+electrical+aspects+of+the+functional+organization.">Insect olfactory sensilla: structural, chemical and electrical aspects of the functional organization.</a> Receptors for Neurotransmitters. Hormones and Pheromones in Insects. , 261-282 (1980).</li></ol>

利害の衝突は宣言されません。

嗅覚信号を測定するために、ライブへそorangewormの蛾を抑制するために開発されたこの新しい方法は、堅牢で非常に信頼性が高いことが証明されている。我々は日常的にアーモンドやピスタチオなどの自然宿主の基質から小説誘引を分離し、識別するためにこの方法を採用しています。

<table border="1"><tbody><tr><td> 材料名 </td><td> カタログ＃/モデル </td><td> 会社 </td><td> コメント </td></tr><tr><td>顕微鏡</td><td> BX51WIモデル</td><td>オリンパス、米国</td><td></td></tr><tr><td>ステレオ顕微鏡</td><td> MZ75 </td><td>ライカマイクロシステムズ（株）米国</td><td></td></tr><tr><td>フィラメントと1.0ミリメートルホウケイ酸キャピラリーチューブ</td><td> 1B100F - 3 </td><td> WPI社、フロリダ州</td><td></td></tr><tr><td>マイクロピペットプラー</td><td> P - 97 </td><td>サターインスツルメンツ、CA </td><td></td></tr><tr><td>刺激コントローラ</td><td> CS - 55モデル</td><td> Syntech、ドイツ</td><td></td></tr><tr><td>ハイインピーダンスプリアンプ （ユニバーサルシングルエンドプローブ） </td><td></td><td> Syntech、ドイツ</td><td></td></tr><tr><td>アンプ/データ収集システム（USB - IDAC） </td><td></td><td> Syntech、ドイツ</td><td></td></tr><tr><td> EAGマニピュレーターMP - 12 </td><td></td><td> Syntech、ドイツ</td><td></td></tr><tr><td> （Z、Z）-11,13 - hexadecadienal </td><td></td><td> Bedoukianリサーチ社、コネチカット州。 </td><td></td></tr><tr><td>ワットマンろ紙</td><td> 1001070 </td><td>ワットマン米国</td><td></td></tr><tr><td> 5mLのポリプロピレンの注射器</td><td> 309633 </td><td> BDシリンジ、NJ </td><td></td></tr><tr><td>ピペットチップ（200μL） </td><td> 1111-0806 </td><td>米国サイエンティフィック株式会社</td><td></td></tr><tr><td> 669 mLの弁当箱、13 × 13センチメートル、高さ、4.5センチ、 </td><td></td><td>ラバーメイド</td><td></td></tr><tr><td>のどが渇いて超吸収、27.9 X 27.9センチメートル</td><td></td><td>セイフウェイ</td><td></td></tr><tr><td>非乾燥粘土</td><td> 18150 </td><td> Claytoon、バンアーケンインターナショナル、カリフォルニア州</td><td></td></tr><tr><td>環境室</td><td> I - 30BLLモデル</td><td></td><td></td></tr></tbody></table>

electrophysiological measurements from a moth olfactory system

昆虫の嗅覚システムは、アンテナ上にあるすべての嗅覚受容体の神経細胞（ORNs）から、個々に収容されたものからの応答を加算しているelectroantennograms（EAG）と、単一感覚子記録（SSR）の形で記録嗅覚誘発応答のためのユニークな機会を提供するそれぞれ感覚器、。これらのアプローチは、昆虫の化学通信のより良い理解を得るために悪用されている。特定された刺激は、病害虫の管理戦略の誘引や忌避剤のどちらかとして使用することができます。

Watch this Scientific Journal Video about Electrophysiological Measurements from a Moth Olfactory System at JoVE.com

Electrophysiological Measurements from a Moth Olfactory System

昆虫の嗅覚システムは、アンテナ上にあるすべての嗅覚受容体の神経細胞（ORNs）から、個々に収容されたものからの応答を加算しているelectroantennograms（EAG）と、単一感覚子記録（SSR）の形で記録嗅覚誘発応答のためのユニークな機会を提供するそれぞれ感覚器、。

蛾嗅覚系の電気生理学的測定

Insect olfactory systems provide unique opportunities for recording odorant-induced responses in the forms of electroantennograms (EAG) and single ...

electrophysiological-measurements-from-a-moth-olfactory-system

Research

JoVE Journal

Neuroscience

13.7K ビュー.  University of California, Davis. 昆虫の嗅覚システムは、アンテナ上にあるすべての嗅覚受容体の神経細胞（ORNs）から、個々に収容されたものからの応答を加算しているelectroantennograms（EAG）と、単一感覚子記録（SSR）の形で記録嗅覚誘発応答のためのユニークな機会を提供するそれぞれ感覚器、。

ビデオ: Electrophysiological Measurements from a Moth Olfactory System

Electrophysiological Recordings from the Giant Fiber Pathway of D. melanogaster

When startled adult D. melanogaster react by jumping into the air and flying away. In many invertebrate species, including D. melanogaster, the "escape" (or "startle") response during the adult stage is mediated by the multi-component neuronal circuit called the Giant Fiber System (GFS). The comparative large size of the neurons, their distinctive morphology and simple connectivity make the GFS an attractive model system for studying neuronal circuitry. The GFS pathway is composed of two bilaterally symmetrical Giant Fiber (GF) interneurons whose axons descend from the brain along the midline into the thoracic ganglion via the cervical connective. In the mesothoracic neuromere (T2) of the ventral ganglia the GFs form electro-chemical synapses with 1) the large medial dendrite of the ipsilateral motorneuron (TTMn) which drives the tergotrochanteral muscle (TTM), the main extensor for the mesothoracic femur/leg, and 2) the contralateral peripherally synapsing interneuron (PSI) which in turn forms chemical (cholinergic) synapses with the motorneurons (DLMns) of the dorsal longitudinal muscles (DLMs), the wing depressors. The neuronal pathway(s) to the dorsovental muscles (DVMs), the wing elevators, has not yet been worked out (the DLMs and DVMs are known jointly as indirect flight muscles - they are not attached directly to the wings, but rather move the wings indirectly by distorting the nearby thoracic cuticle) (King and Wyman, 1980; Allen et al., 2006). The di-synaptic activation of the DLMs (via PSI) causes a small but important delay in the timing of the contraction of these muscles relative to the monosynaptic activation of TTM (~0.5 ms) allowing the TTMs to first extend the femur and propel the fly off the ground. The TTMs simultaneously stretch-activate the DLMs which in turn mutually stretch-activate the DVMs for the duration of the flight. The GF pathway can be activated either indirectly by applying a sensory (e.g."air-puff" or "lights-off") stimulus, or directly by a supra-threshold electrical stimulus to the brain (described here). In both cases, an action potential reaches the TTMs and DLMs solely via the GFs, PSIs, and TTM/DLM motoneurons, although the TTMns and DLMns do have other, as yet unidentified, sensory inputs. Measuring "latency response" (the time between the stimulation and muscle depolarization) and the "following to high frequency stimulation" (the number of successful responses to a certain number of high frequency stimuli) provides a way to reproducibly and quantitatively assess the functional status of the GFS components, including both central synapses (GF-TTMn, GF-PSI, PSI-DLMn) and the chemical (glutamatergic) neuromuscular junctions (TTMn-TTM and DLMn-DLM). It has been used to identify genes involved in central synapse formation and to assess CNS function.

Electrophysiological Recordings from the Giant Fiber Pathway of D. melanogaster

The Giant Fiber System is a simple neuronal circuit of adult Drosophila melanogaster containing the largest neurons in the fly. We describe the protocol for monitoring synaptic transmission through this pathway by recording post synaptic potentials in dorsal longitudinal (DLM) and tergotrochanteral (TTM) muscles following direct stimulation of the Giant Fiber interneurons.

の巨大線維経路からの電気生理学的記録 D.キイロ</em

When startled adult D. melanogaster react by jumping into the air and flying away. In many invertebrate species, including D. melanogaster, the "escape" ...

神経科学

Isolating Nasal Olfactory Stem Cells from Rodents or Humans

The olfactory mucosa, located in the nasal cavity, is in charge of detecting odours. It is also the only nervous tissue that is exposed to the external environment and easily accessible in every living individual. As a result, this tissue is unique for anyone aiming to identify molecular anomalies in the pathological brain or isolate adult stem cells for cell therapy. 
Molecular abnormalities in brain diseases are often studied using nervous tissue samples collected post-mortem. However, this material has numerous limitations. In contrast, the olfactory mucosa is readily accessible and can be biopsied safely without any loss of sense of smell1. Accordingly, the olfactory mucosa provides an "open window" in the adult human through which one can study developmental (e.g. autism, schizophrenia)2-4 or neurodegenerative (e.g. Parkinson, Alzheimer) diseases4,5. Olfactory mucosa can be used for either comparative molecular studies4,6 or in vitro experiments on neurogenesis3,7.
The olfactory epithelium is also a nervous tissue that produces new neurons every day to replace those that are damaged by pollution, bacterial of viral infections. This permanent neurogenesis is sustained by progenitors but also stem cells residing within both compartments of the mucosa, namely the neuroepithelium and the underlying lamina propria8-10. We recently developed a method to purify the adult stem cells located in the lamina propria and, after having demonstrated that they are closely related to bone marrow mesenchymal stem cells (BM-MSC), we named them olfactory ecto-mesenchymal stem cells (OE-MSC)11.
Interestingly, when compared to BM-MSCs, OE-MSCs display a high proliferation rate, an elevated clonogenicity and an inclination to differentiate into neural cells. We took advantage of these characteristics to perform studies dedicated to unveil new candidate genes in schizophrenia and Parkinson's disease4. We and others have also shown that OE-MSCs are promising candidates for cell therapy, after a spinal cord trauma12,13, a cochlear damage14 or in an animal models of Parkinson's disease15 or amnesia16.
In this study, we present methods to biopsy olfactory mucosa in rats and humans. After collection, the lamina propria is enzymatically separated from the epithelium and stem cells are purified using an enzymatic or a non-enzymatic method. Purified olfactory stem cells can then be either grown in large numbers and banked in liquid nitrogen or induced to form spheres or differentiated into neural cells. These stem cells can also be used for comparative omics (genomic, transcriptomic, epigenomic, proteomic) studies.

We describe here a method for biopsying olfactory mucosa from rat and human nasal cavities. These biopsies can be used for either identifying molecular anomalies in brain diseases or isolating multipotent adult stem cells that can be utilized for cell transplantation in animal models of brain trauma/disease.

げっ歯類やヒトから鼻嗅覚幹細胞を単離する

The olfactory mucosa, located in the nasal cavity, is in charge of detecting odours. It is also the only nervous tissue that is exposed to the external ...

Electrophysiological Measurements and Analysis of Nociception in Human Infants

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are based on behavioural and physiological body reactions, such as crying, body movements or changes in facial expression. While these measures demonstrate that infants mount a response following noxious stimulation, they are limited: they are based on activation of subcortical somatic and autonomic motor pathways that may not be reliably linked to central sensory processing in the brain. Knowledge of how the central nervous system responds to noxious events could provide an insight to how nociceptive information and pain is processed in newborns.
The heel lancing procedure used to extract blood from hospitalised infants offers a unique opportunity to study pain in infancy. In this video we describe how electroencephalography (EEG) and electromyography (EMG) time-locked to this procedure can be used to investigate nociceptive activity in the brain and spinal cord.
This integrative approach to the measurement of infant pain has the potential to pave the way for an effective and sensitive clinical measurement tool.

The assessment and treatment of pain in infants is difficult because infants cannot verbally report their experience. In this video we describe quantitative electrophysiological methods and analysis techniques that can be used to measure the response to noxious events from the infant nervous system.

電気生理学的測定とヒト幼児における痛覚の分析

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are ...

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

Animals sample the odorous environment around them through the chemosensory systems located in the nasal cavity. Chemosensory signals affect complex behaviors such as food choice, predator, conspecific and mate recognition and other socially relevant cues. Olfactory receptor neurons (ORNs) are located in the dorsal part of the nasal cavity embedded in the olfactory epithelium. These bipolar neurons send an axon to the olfactory bulb (see Fig. 1, Reisert &amp; Zhao1, originally published in the Journal of General Physiology) and extend a single dendrite to the epithelial border from where cilia radiate into the mucus that covers the olfactory epithelium. The cilia contain the signal transduction machinery that ultimately leads to excitatory current influx through the ciliary transduction channels, a cyclic nucleotide-gated (CNG) channel and a Ca2+-activated Cl- channel (Fig. 1). The ensuing depolarization triggers action potential generation at the cell body2-4.
In this video we describe the use of the "suction pipette technique" to record odorant-induced responses from ORNs. This method was originally developed to record from rod photoreceptors5 and a variant of this method can be found at jove.com modified to record from mouse cone photoreceptors6. The suction pipette technique was later adapted to also record from ORNs7,8. Briefly, following dissociation of the olfactory epithelium and cell isolation, the entire cell body of an ORN is sucked into the tip of a recording pipette. The dendrite and the cilia remain exposed to the bath solution and thus accessible to solution changes to enable e.g. odorant or pharmacological blocker application. In this configuration, no access to the intracellular environment is gained (no whole-cell voltage clamp) and the intracellular voltage remains free to vary. This allows the simultaneous recording of the slow receptor current that originates at the cilia and fast action potentials fired by the cell body9. The difference in kinetics between these two signals allows them to be separated using different filter settings. This technique can be used on any wild type or knockout mouse or to record selectively from ORNs that also express GFP to label specific subsets of ORNs, e.g. expressing a given odorant receptor or ion channel.

Olfactory receptor neurons (ORNs) convert odor signals first into a receptor current that in turn triggers action potentials that are conveyed to second order neurons in the olfactory bulb. Here we describe the suction pipette technique to record simultaneously the odorant-induced receptor current and action potentials from mouse ORNs.

吸引ピペット技術を用いた嗅覚受容ニューロンから記録匂い物質誘発性応答

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

Appetitive Associative Olfactory Learning in Drosophila Larvae

In the following we describe the methodological details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with genetic interference, provides a handle to analyze the neuronal and molecular fundamentals of specifically associative learning in a simple larval brain.
Organisms can use past experience to adjust present behavior. Such acquisition of behavioral potential can be defined as learning, and the physical bases of these potentials as memory traces1-4. Neuroscientists try to understand how these processes are organized in terms of molecular and neuronal changes in the brain by using a variety of methods in model organisms ranging from insects to vertebrates5,6. For such endeavors it is helpful to use model systems that are simple and experimentally accessible. The Drosophila larva has turned out to satisfy these demands based on the availability of robust behavioral assays, the existence of a variety of transgenic techniques and the elementary organization of the nervous system comprising only about 10,000 neurons (albeit with some concessions: cognitive limitations, few behavioral options, and richness of experience questionable)7-10.
Drosophila larvae can form associations between odors and appetitive gustatory reinforcement like sugar11-14. In a standard assay, established in the lab of B. Gerber, animals receive a two-odor reciprocal training: A first group of larvae is exposed to an odor A together with a gustatory reinforcer (sugar reward) and is subsequently exposed to an odor B without reinforcement 9. Meanwhile a second group of larvae receives reciprocal training while experiencing odor A without reinforcement and subsequently being exposed to odor B with reinforcement (sugar reward). In the following both groups are tested for their preference between the two odors. Relatively higher preferences for the rewarded odor reflect associative learning - presented as a performance index (PI). The conclusion regarding the associative nature of the performance index is compelling, because apart from the contingency between odors and tastants, other parameters, such as odor and reward exposure, passage of time and handling do not differ between the two groups9.

Appetitive Associative Olfactory Learning in Drosophila Larvae

Drosophila larvae are able to associate odor stimuli with gustatory reward. Here we describe a simple behavioral paradigm that allows the analysis of appetitive associative olfactory learning.

で食欲連想嗅覚学習<em&gt;ショウジョウバエ</em&gt;幼虫

In the following we describe the methodological  details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with ...

In Vivo Electrophysiological Measurements on Mouse Sciatic Nerves

Electrophysiological studies allow a rational classification of various neuromuscular diseases and are of help, together with neuropathological techniques, in the understanding of the underlying pathophysiology1. Here we describe a method to perform electrophysiological studies on mouse sciatic nerves in vivo.
The animals are anesthetized with isoflurane in order to ensure analgesia for the tested mice and undisturbed working environment during the measurements that take about 30 min/animal. A constant body temperature of 37 &#176;C is maintained by a heating plate and continuously measured by a rectal thermo probe2. Additionally, an electrocardiogram (ECG) is routinely recorded during the measurements in order to continuously monitor the physiological state of the investigated animals.
Electrophysiological recordings are performed on the sciatic nerve, the largest nerve of the peripheral nervous system (PNS), supplying the mouse hind limb with both motoric and sensory fiber tracts. In our protocol, sciatic nerves remain in situ and therefore do not have to be extracted or exposed, allowing measurements without any adverse nerve irritations along with actual recordings. Using appropriate needle electrodes3 we perform both proximal and distal nerve stimulations, registering the transmitted potentials with sensing electrodes at gastrocnemius muscles. After data processing, reliable and highly consistent values for the nerve conduction velocity (NCV) and the compound motor action potential (CMAP), the key parameters for quantification of gross peripheral nerve functioning, can be achieved.

In Vivo Electrophysiological Measurements on Mouse Sciatic Nerves

Measurements of nerve conduction properties in vivo exemplify a powerful tool to characterize various animal models of neuromuscular diseases. Here, we present an easy and reliable protocol by which electrophysiological analysis on sciatic nerves of anesthetized mice can be performed.

マウス坐骨神経に生体内電気生理学的測定値の

Electrophysiological studies allow a rational classification of various neuromuscular diseases and are of help, together with neuropathological ...

Electrophysiological Recording From Drosophila Labellar Taste Sensilla

The peripheral taste response of insects can be powerfully investigated with electrophysiological techniques. The method described here allows the researcher to measure gustatory responses directly and quantitatively, reflecting the sensory input that the insect nervous system receives from taste stimuli in its environment. This protocol outlines all key steps in performing this technique. The critical steps in assembling an electrophysiology rig, such as selection of necessary equipment and a suitable environment for recording, are delineated. We also describe how to prepare for recording by making appropriate reference and recording electrodes, and tastant solutions. We describe in detail the method used for preparing the insect by insertion of a glass reference electrode into the fly in order to immobilize the proboscis. We show traces of the electrical impulses fired by taste neurons in response to a sugar and a bitter compound. Aspects of the protocol are technically challenging and we include an extensive description of some common technical challenges that may be encountered, such as lack of signal or excessive noise in the system, and potential solutions. The technique has limitations, such as the inability to deliver temporally complex stimuli, observe background firing immediately prior to stimulus delivery, or use water-insoluble taste compounds conveniently. Despite these limitations, this technique (including minor variations referenced in the protocol) is a standard, broadly accepted procedure for recording Drosophila neuronal responses to taste compounds.

Electrophysiological Recording From Drosophila Labellar Taste Sensilla

This protocol describes extracellular recording of the action potential responses fired by labellar taste neurons in Drosophila.

から電気生理学的記録<em&gt;ショウジョウバエ</em&gt; Labellar味感覚器

The peripheral taste response of insects can be powerfully investigated with electrophysiological techniques. The method described here allows the ...

The Olfactory System as a Model to Study Axonal Growth Patterns and Morphology In Vivo

The olfactory system has the unusual capacity to generate new neurons throughout the lifetime of an organism. Olfactory stem cells in the basal portion of the olfactory epithelium continuously give rise to new sensory neurons that extend their axons into the olfactory bulb, where they face the challenge to integrate into existing circuitry. Because of this particular feature, the olfactory system represents a unique opportunity to monitor axonal wiring and guidance, and to investigate synapse formation. Here we describe a procedure for in vivo labeling of sensory neurons and subsequent visualization of axons in the olfactory system of larvae of the amphibian Xenopus laevis. To stain sensory neurons in the olfactory organ we adopt the electroporation technique. In vivo electroporation is an established technique for delivering fluorophore-coupled dextrans or other macromolecules into living cells. Stained sensory neurons and their axonal processes can then be monitored in the living animal either using confocal laser-scanning or multiphoton microscopy. By reducing the number of labeled cells to few or single cells per animal, single axons can be tracked into the olfactory bulb and their morphological changes can be monitored over weeks by conducting series of in vivo time lapse imaging experiments. While the described protocol exemplifies the labeling and monitoring of olfactory sensory neurons, it can also be adopted to other cell types within the olfactory and other systems.

The Olfactory System as a Model to Study Axonal Growth Patterns and Morphology In Vivo

We describe a protocol for in vivo labeling of olfactory sensory neurons by electroporation and subsequent confocal laser-scanning or multiphoton microscopy to visualize neuronal morphology and its development over time.

モデルとしての嗅覚系は、軸索の成長パターンと形態を研究する<em&gt;インビボ</em

The olfactory system has the unusual capacity to generate new neurons throughout the lifetime of an organism. Olfactory stem cells in the basal portion of ...

Whole Mount Labeling of Cilia in the Main Olfactory System of Mice

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single dendrite to the surface of the epithelium, ending in a structure called dendritic knob. Cilia emanate from this knob into the mucus covering the epithelial surface. The proteins of the olfactory signal transduction cascade are mainly localized in the ciliary membrane, being in direct contact with volatile substances in the environment. For a detailed understanding of olfactory signal transduction, one important aspect is the exact morphological analysis of signaling protein distribution. Using light microscopical approaches in conventional cryosections, protein localization in olfactory cilia is difficult to determine due to the density of ciliary structures. To overcome this problem, we optimized an approach for whole mount labeling of cilia, leading to improved visualization of their morphology and the distribution of signaling proteins. We demonstrate the power of this approach by comparing whole mount and conventional cryosection labeling of Kirrel2. This axon-guidance adhesion molecule is known to localize in a subset of sensory neurons and their axons in an activity-dependent manner. Whole mount cilia labeling revealed an additional and novel picture of the localization of this protein.

Cilia of olfactory sensory neurons contain proteins of the signal transduction cascade, but a detailed spatial analysis of their distribution is difficult in cryosections. We describe here an optimized approach for whole mount labeling and en face visualization of ciliary proteins.

マウスの主嗅覚系における繊毛のホールマウントのラベリング

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single ...

A Free-breathing fMRI Method to Study Human Olfactory Function

The study of human olfaction is a highly complex and valuable field with applications ranging from biomedical research to clinical evaluation. Currently, evaluation of the functions of the human central olfactory system with functional magnetic resonance imaging (fMRI) is still a challenge because of several technical difficulties. There are some significant variables to take into account when considering an effective method for mapping the function of the central olfactory system using fMRI, including proper odorant selection, the interaction between odor presentation and respiration, and potential anticipation of or habituation to odorants. An event-related, respiration-triggered olfactory fMRI technique can accurately administer odorants to stimulate the olfactory system while minimizing potential interference. It can effectively capture the precise onsets of fMRI signals in the primary olfactory cortex using our data post-processing method. The technique presented here provides an efficient and practical means for generating reliable olfactory fMRI results. Such a technique can ultimately be applied in the clinical realm as a diagnostic tool for diseases associated with olfactory degeneration, including Alzheimer&#39;s and Parkinson&#39;s disease, as we begin to further understand the complexities of the human olfactory system.

We present the technical challenges and solutions for obtaining reliable functional magnetic resonance imaging (fMRI) data from the human central olfactory system. This includes special considerations in olfactory fMRI paradigm design, descriptions of fMRI data acquisition with an MRI-compatible olfactometer, odorant selection, and a special software tool for data post-processing.

ヒトの嗅覚機能を研究する自由呼吸fMRI法

The study of human olfaction is a highly complex and valuable field with applications ranging from biomedical research to clinical evaluation. Currently, ...