Biz bal arıları dokunsal öğrenme paradigması bize tanıtmak için Joachim Erber ederim. Bu eser Alman Mükemmeliyet Girişimi çerçevesinde finanse Mükemmellik 277 CITEC arasında Küme tarafından desteklenmiştir.

1. Arılar hazırlanması <ol><li> Bir toplayıcı geziden dönerken Nektarı veya Polen yem bir sükroz besleyici veya doğrudan kovan girişinde ya alanına yakalanmıştır. Her bir arı bir köpük tıkaç ile kapatılmalıdır ve daha işleme için laboratuvar hemen içine alınmış bir cam şişeye yakalanır. </li><li> Onlar hareketsizlik ilk işaretleri göstermek ° C kadar laboratuvarda, yakalanan arılar kısaca 4 de buzdolabında soğutulur. </li><li> Her tek immobilize arı baş ve toraks arasında ve karın üzerinde yapışkan bant ile küçük bir metal tüp monte edilir. Bakım hortum ve antenler serbestçe hareketli olduğu alınmalıdır. </li><li> Vizyon tıkamak için beyaz boya (örneğin solvent içermeyen Tipp-Ex) ile sabit arı bileşik gözleri ve ocelli boyayın. </li><li> Kayıtları sırasında baş hareketleri önlemek için baş ve toraks arasında kasete bunu düzeltmek için arı başının arkasına erimiş mum küçük bir damla ekleyin. </li><li> Daha iyi belirlenmesi için kaset üzerinde bir sayı ile her bir arı işaretleyin ve dehidratasyonu önlemek için nemli bir atmosfere sabit arı ile tüp yerleştirin. </li><li> Bir şırınga ile başvuran 30% sakaroz çözeltisini damlacıkları ile 5 saniye boyunca her bir arı besle ve dokunsal Klima protokolü ile başlamadan önce tüm arılar 30 dk kurtarmanızı sağlar. </li></ol> 2. Dokunsal Şartlandırma <ol><li> Klima önce, her bir arı antenleri uygulanan% 30 sukroz uyarana hortum uzantısı yanıt (PER) için test edilmelidir. Böylece hortum ucu açık alt çeneleri arasına bir sanal çizgi geçmek zorundadır. Sakaroz uyaran bir PER ile yanıt vermeyen bütün arıların atın. </li><li> Dokunsal klima şartlı uyarıcı (CS) gibi bir düz veya bir oyulmuş desen, örneğin, yatay veya 150 mikron dalga boyuna sahip bir ızgara oluşturan dikey oluklar ile bir pirinç küp (örn. 3 x 5 mm), kullanmak için. Fya da koşulsuz uyarıcı (ABD) bir% 30 sukroz çözeltisi (su içinde seyreltilmiş beyaz şeker,) kullanın. </li><li> Pirinç küp (CS) Klima işlem sırasında tam konumlandırma sağlamak için (örneğin, Märzhäuser MM33) bir micromanipulator bir tutucuya yerleştirilir. ABD% 30 sukroz solüsyonu ile dolu bir şırınga ile arı sunulmaktadır. </li><li> Klima prosedür 5 dk arası bir deneme süresi (ITI) ile dokunsal uyarıcı (CS) ve sakaroz çözeltisi (ABD) beş eşleşmeleri oluşur. Monte edilmiş dokunsal uyarıcı (CS) ile mikromanipülatör önünde bir tek arı yerleştirin. Dokunsal uyaran yüzeyinin arının başında (Şekil 1 A ve B) paralel olacak şekilde yavaşça CS, yerleştirin. Hayvan ve dokunsal nesne arasındaki mesafe test arı antennal çalışma yarıçapı aralığında olmalıdır, arı rahat bir pozisyonda w dokunsal uyaran taramak gerekir yaniITH antenler de. Şekil 1&#39;de gösterilen örnekte, mesafe 3 mm idi. Arı 5 sn için dokunsal uyaran (CS) tarama edelim. Ilk olarak 3 saniye sonra,% 30 sakaroz (ABD) hortum altında bir şırınga ile çözelti bir damlacık sunulmaktadır. Nazikçe hortum artırmak için enjektör ucu kullanın. Hortum altında Sakaroz stimülasyon 15 PER koşulsuz seçmektedirler. Arı sakaroz ödül yalamak için izin ver. Klima sırasında tam olarak ne zaman aralıklarını korumak için bir alarm sinyali ile bir kronometre kullanın. </li><li> Klimalı BAŞINA arı öğrenme başarısı için bir ölçü olarak kullanılır. İlk ödüllendirilir eşlendirme sonra, arıların önümüzdeki ödül beklediklerini belirten, kendi burnumun uzantısı CS sunum yanıt başlar. Dokunsal uyaran sunum 3 sn süre penceresi sırasında ve sakkaroz sunum öncesi zaman görülen bir tam genişletilmiş hortum olumlu bir yanıt olarak puanlanmıştır. Hayır yanıtı olumsuz sayılır. OCCPER ve urrence deneyci tarafından unutulmamalıdır. CS sunumu sırasında BAŞINA gösteren arıların yüzdesi her deneme için çizilir. </li></ol> 3. Kinematik Kayıtlar <ol><li> Tek bir harnessed arı antennal hareketleri uygun bir makro lens (örneğin TechSpec VZM 200 ile donatılmıştır Basler A602f-2, bir yangın tel bağlantısı yoluyla 50 fps&#39;de işletilen) bir dijital video kamera ile kaydedilmiştir. Kamera yukarıdan aşağıya bir görünümde hayvan (Şekil 1 A) üstünde konumlandırılmıştır. Şekil 1&#39;de, kayıtların uzaysal çözünürlüğü 0.02 mm / piksel. </li><li> Kameranın objektif altında farklı yönlerde 1 mm&#39;lik bir kenar uzunluğu ile 10 x 10 mm dama tek görüntü kaydederken tarafından kamera kalibre. Kalibrasyon kamera kalibrasyon alet kutusu Matlab 16 ile yapılabilir. </li><li> Kamera merceğinin altında bir tek sabit arı yerleştirin. Bir mikromanipülatör sabitlenmiş dokunsal uyaran, sunun, Hayvan için. Dokunsal kurutma için tarif edildiği gibi aynı şekilde devam edin ve arı nesne tararken anten hareketini kaydederler. Bu tam anten çalışma aralığı ve dokunsal uyarı görünür olması önemlidir. Uyaran desen seçenekleri test ve deneme numaraları deneysel tasarım bağlıdır. </li></ol> 4. Veri Analizi <ol><li> Görüntünün arka plan hesaplama Matlab yapılır. İlk olarak, zaman içinde orta gri skala değeri her bir piksel için hesaplanan gerekmektedir. Statik nesneler, arı ve dokunsal uyaran sabit kafa gibi, görüntü arka planı teşkil edecektir. Hareketli nesneler, iki anten gibi, arka planın parçası olmayacaktır. </li><li> Kaydedilen video her bir ardışık kare yüklü olması gerekir, ve akım çerçeve ve resim arka planı arasındaki fark hesaplanarak gerekir. Bu çıkarma sonucu kareden diğerine hareket ediyor görüntünün bölümleri vurgular. Kimlikeally, arı antenleri sıfır değerleri (Şekil 1C) tek alanlardır. </li><li> Daha fazla işlem için, sıfır olmayan piksel değerleri ile en büyük iki alanda antenler olduğu varsayılır. Her iki anten için bir ikili maskesi piksel anten ve aksi değeri 0 aitse, her piksel için bir değer 1 içeren oluşturulan olmaktır. Bu maske daha sonra anten ipuçları yerelleştirilmesi için bir temel olarak hizmet vermektedir. Tüm görüntü için ön maske kazandırmak farkı görüntünün her pikselin gri değer önceden tanımlanmış eşik karşılaştırılır. Biz gürültü icabına beri sonradan bu eşiğin oldukça düşük olduğu için tercih edilir. Öncelikle, görüntü gürültü küçük bölgeler nedeniyle halen maskesi parçası vardır: ön maskesi sonra hala hatalar iki çeşit taşır. İkincisi, anten ait alanlar mutlaka tam bağlanmamış olabilir. Arka plan yaklaşık olarak anten ile aynı ışıldama varsa ikincisi çoğunlukla oluşur. Bu eserler ortadan kaldırmak için, standartgörüntü işleme morfolojik işlemi uygulanır, standart Matlab fonksiyonları imerode ve (daha fazla açıklama için 17 pp 158-205 bakınız) sırasıyla imdilate güvenerek, görüntü erozyon ve dilatasyon bir arada yani. Denoising işleminden sonra antenler hipotezler hala aynı maske içinde yer alır. Bu nedenle, bir sonraki adım olarak ikili maske standart Matlab fonksiyonuyla bwlabel (görüntü bölütleme algoritması hakkında daha fazla bilgi için 17 pp 40-48 bakınız) kullanarak ayrık alanlar için kümelenmiştir. </li><li> Küme başına piksel sayısı sayılır ve en büyük iki kümesi seçilmektedir. Ağırlık merkezi sol ve sağ anten (Şekil 1B) arasındaki fark olarak hesaplanmıştır. </li><li> Her anten için anten ucu proksimal-to-distal yönde (Şekil 1D bakınız) en yüksek değere sahip bir kümede piksel olarak tanımlanabilir. </li></ol>

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Çıkar çatışması ilan etti.

Arı hazırlanması: Toplama ve arı hayvan düşük gerilme düzeyini sağlamak için, hızlı bir şekilde yapılmalıdır sabitleme. Stres PER-yanıt üzerinde bir etkisi vardır ve bu nedenle, arılar 19, 20 öğrenme performansı üzerinde dolaylı bir etkisi olabilir. Stres düzeyini hızlı bir şekilde hareketsiz alındıktan hemen sonra doğrudan buz üzerinde arılar ile cam şişe yerleştirerek azaltılabilir. Arılara anestezi öncesi dönemde olduğu uzun kurtarmak için daha fa...

<table border="1"><tbody><tr><td> Reaktif / Malzeme Adı </td><td> Şirket </td><td> Katalog Numarası </td><td> Yorumlar </td></tr><tr><td> Polen toplayıcılar, Apis mellifera L. </td><td></td><td></td><td> Özel kurdeşen </td></tr><tr><td> Cam şişe (22 x 45 mm) </td><td> Carl Roth GmbH &amp; Co </td><td> X655.1 </td><td></td></tr><tr><td> Pirinç borular, 10 x 30 mm </td><td></td><td></td><td> Kendinden üretilen </td></tr><tr><td> Pirinç küp, 3 x 5 mm </td><td></td><td></td><td> Kendinden üretilen </td></tr><tr><td> Tipp-Ex (solventsiz) </td><td> Tipp-Ex GmbH &amp; Co KG </td><td></td><td></td></tr><tr><td> Sukroz çözeltisi (% 30) </td><td></td><td></td><td> Ev suger su ile seyreltilir </td></tr><tr><td> Etanol (% 70) </td><td> Carl Roth GmbH &amp; Co </td><td> 9065,3 &lt;/ Td&gt; <td></td></tr><tr><td> Şırınga (1 ml) ve iğne (0.6 x 25) </td><td> Carl Roth GmbH &amp; Co </td><td> 59.1 </td><td></td></tr><tr><td> İzle Durdur </td><td> Carl Roth GmbH &amp; Co </td><td> L423.1 </td><td></td></tr><tr><td> Mikromanipülatör </td><td> Märzhäuser MM33 </td><td> 00-42-101-0000 </td><td></td></tr><tr><td> Dijital video kamera </td><td> Basler A602f-2 </td><td></td><td></td></tr><tr><td> Kamera makro objektif </td><td> TechSpec VZM 200 </td><td></td><td></td></tr><tr><td> Matlab R2009b </td><td> MathWorks </td><td></td><td></td></tr></tbody></table>

tactile conditioning and movement analysis of antennal sampling strategies in honey bees (apis mellifera l.)

Bal arıları (Apis mellifera L.) eusocial böcekler ve iyi işçilik ve çağrışımsal öğrenme yeteneği 1, 2 kendi karmaşık bir bölümü için bilinir. Onlar larva hemşirelik veya düzenli altıgen yemek için tarak (örneğin polen veya nektar) ve yavru 3 bina nerede işçi arılar, karanlık kovan içinde kendi yaşamının ilk yarısını harcamak. Antenler olağanüstü multisensory duyarga ve kovan bina 5 ve örüntü tanıma 6 dahil olmak üzere çeşitli dokunsal aracılı görevleri 4, çok önemli bir rol oynamaktadır. Daha sonra hayatta, her bir arı yiyecek aramak için kovanı terk eder. Sonra arı, karlı gıda kaynakları ayrımcılık konumlarını ezberlemek ve 7 yuvasını arkadaşları için iletişim öğrenmek zorundadır. Arı besin kaynağının multisensory hafıza oluşturmak üzere petal yüzey 9 renk ve koku veren 7, 8 gibi başka çiçek sinyalleri, ama aynı zamanda dokunsal işaretlerini kullanır. Laboratuvar işbirliği altındaşulları, arılar gibi kenarlarını ya da anten 10, 11, 12, 13 ile oluklar gibi dokunsal nesnenin özellikleri, ayrımcılık bir iştahlı öğrenme paradigması içinde eğitilmiş olabilir. Bu öğrenme paradigma yakından koşulmuş arı 14 hortum uzantı yanıt (başına) klasik koku klima ilgilidir. Laboratuarda dokunsal öğrenme paradigması avantajı anten hareket model analizi de dahil olmak üzere çeşitli fizyolojik ölçümleri ile ilgili öğrenme davranışsal deney birleştirme ihtimalidir.

Aşağıdaki deneyde görmeyenlerin dokunsal öğrenme bal arıları antennal örnekleme davranışını nasıl etkilediği incelenmiştir. Bunun için, biz önce naif ve klimalı arılar anten ucu ve dokunsal uyaran sunum sırasında hareketi izlenir. İlk olarak, polen yem bir grup kendiliğinden hareket anten (N = 42) 1 dakika boyunca kaydedildi. Arıların yarısı (N = 21) daha sonra beş kez% 30 sakaroz ödül ile dokunsal uyaran eşleştirme tarafından şartlandırılmış oldu. Bu k...

Watch this Scientific Journal Video about Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L. at JoVE.com

Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L.

Bu protokolü biz durumu dokunsal uyaranlara karşı bal arıları koşulmuş ve ince çaplı anten örnekleme deseni kinematik analiz için bir 2B hareket yakalama tekniği tanıtmak nasıl gösterir.

Dokunsal İklimlendirme Ve Bal Arısı yılında antennal Örnekleme Stratejileri Hareket Analizi (<em&gt; Apis mellifera</em&gt; L.)

Honey bees (Apis mellifera L.) are eusocial insects and well known for their complex division of labor and associative learning capability1, 2. The worker ...

tactile-conditioning-movement-analysis-antennal-sampling-strategies

Research

JoVE Journal

Neuroscience

Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees (Apis mellifera L.)

10.5K Görüntüleme.  Bielefeld University. Bu protokolü biz durumu dokunsal uyaranlara karşı bal arıları koşulmuş ve ince çaplı anten örnekleme deseni kinematik analiz için bir 2B hareket yakalama tekniği tanıtmak nasıl gösterir.

Video: Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L.

Behavioural Pharmacology in Classical Conditioning of the Proboscis Extension Response in Honeybees (Apis mellifera)

Honeybees (Apis mellifera) are well known for their communication and orientation skills and for their impressive learning capability1,2. Because the survival of a honeybee colony depends on the exploitation of food sources, forager bees learn and memorize variable flower sites as well as their profitability. Forager bees can be easily trained in natural settings where they forage at a feeding site and learn the related signals such as odor or color. Appetitive associative learning can also be studied under controlled conditions in the laboratory by conditioning the proboscis extension response (PER) of individually harnessed honeybees3,4. This learning paradigm enables the study of the neuronal and molecular mechanisms that underlie learning and memory formation in a simple and highly reliable way5-12. A behavioral pharmacology approach is used to study molecular mechanisms. Drugs are injected systemically to interfere with the function of specific molecules during or after learning and memory formation13-16.
Here we demonstrate how to train harnessed honeybees in PER conditioning and how to apply drugs systemically by injection into the bee flight muscle.

Behavioural Pharmacology in Classical Conditioning of the Proboscis Extension Response in Honeybees Apis mellifera

We demonstrate how to implement a behavioral pharmacology method in an appetitive olfactory conditioning paradigm in honeybees (Apis mellifera) by systemic application of drugs. This method allows investigation of the mechanisms underlying learning and memory formation in a simple and reliable way.

Bal arısı burnumun Extension Tepki Klasik Koşullanma Davranış Farmakoloji ( Apis mellifera)

Honeybees (Apis mellifera) are well known for their communication and orientation skills and for their impressive learning capability1,2. Because the ...

Nörobilim

Automated Sholl Analysis of Digitized Neuronal Morphology at Multiple Scales

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3. Specifically, it affects the ability of neurons to receive inputs from other cells2 and contributes to the propagation of action potentials4,5. The morphology of the neurites also affects how information is processed. The diversity of dendrite morphologies facilitate local and long range signaling and allow individual neurons or groups of neurons to carry out specialized functions within the neuronal network6,7. Alterations in dendrite morphology, including fragmentation of dendrites and changes in branching patterns, have been observed in a number of disease states, including Alzheimer's disease8, schizophrenia9,10, and mental retardation11. The ability to both understand the factors that shape dendrite morphologies and to identify changes in dendrite morphologies is essential in the understanding of nervous system function and dysfunction.
Neurite morphology is often analyzed by Sholl analysis and by counting the number of neurites and the number of branch tips. This analysis is generally applied to dendrites, but it can also be applied to axons. Performing this analysis by hand is both time consuming and inevitably introduces variability due to experimenter bias and inconsistency. The Bonfire program is a semi-automated approach to the analysis of dendrite and axon morphology that builds upon available open-source morphological analysis tools. Our program enables the detection of local changes in dendrite and axon branching behaviors by performing Sholl analysis on subregions of the neuritic arbor. For example, Sholl analysis is performed on both the neuron as a whole as well as on each subset of processes (primary, secondary, terminal, root, etc.) Dendrite and axon patterning is influenced by a number of intracellular and extracellular factors, many acting locally. Thus, the resulting arbor morphology is a result of specific processes acting on specific neurites, making it necessary to perform morphological analysis on a smaller scale in order to observe these local variations12.
The Bonfire program requires the use of two open-source analysis tools, the NeuronJ plugin to ImageJ and NeuronStudio. Neurons are traced in ImageJ, and NeuronStudio is used to define the connectivity between neurites. Bonfire contains a number of custom scripts written in MATLAB (MathWorks) that are used to convert the data into the appropriate format for further analysis, check for user errors, and ultimately perform Sholl analysis. Finally, data are exported into Excel for statistical analysis. A flow chart of the Bonfire program is shown in Figure 1.

We have developed a computer program to analyze neuronal morphology. In combination with two existing open source analysis tools, our program performs Sholl analysis and determines the number of neurites, branch points, and neurite tips. The analyses are performed so that local changes in neurite morphology can be observed.

Çoklu Ölçeklerde Sayısallaştırılmış Nöronal Morfoloji Otomatik Sholl Analizi

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3.  Specifically, it affects the ability of neurons to ...

The Culture of Primary Motor and Sensory Neurons in Defined Media on Electrospun Poly-L-lactide Nanofiber Scaffolds

Electrospinning is a technique for producing micro- to nano-scale fibers. Fibers can be electrospun with varying degrees of alignment, from highly aligned to completely random. In addition, fibers can be spun from a variety of materials, including biodegradable polymers such as poly-L-lactic acid (PLLA). These characteristics make electrospun fibers suitable for a variety of scaffolding applications in tissue engineering. Our focus is on the use of aligned electrospun fibers for nerve regeneration. We have previously shown that aligned electrospun PLLA fibers direct the outgrowth of both primary sensory and motor neurons in vitro. We maintain that the use of a primary cell culture system is essential when evaluating biomaterials to model real neurons found in vivo as closely as possible. Here, we describe techniques used in our laboratory to electrospin fibrous scaffolds and culture dorsal root ganglia explants, as well as dissociated sensory and motor neurons, on electrospun scaffolds. However, the electrospinning and/or culture techniques presented here are easily adapted for use in other applications.

Aligned electrospun fibers direct the growth of neurons in vitro and are a potential component of nerve regeneration scaffolds. We describe a procedure for preparing electrospun fiber substrates and the serum-free culture of primary rat E15 sensory (DRG) and motor neurons. Visualization of neurons by immunocytochemistry is also included.

Kültür İlköğretim Motor ve Electrospun Poli-L-lactide Nanofiber Yapı iskeleleri Tanımlı Medya Duyu Nöronlar

Electrospinning is a technique for producing micro- to nano-scale fibers. Fibers can be electrospun with varying degrees of alignment, from highly aligned ...

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory bulb1-5. Olfactory information in the external world is transmitted to the antennal lobe by olfactory sensory neurons (OSNs), which segregate to distinct regions of neuropil called glomeruli according to the specific olfactory receptor they express. Here, OSN axons synapse with both local interneurons (LNs), whose processes can innervate many different glomeruli, and projection neurons (PNs), which convey olfactory information to higher olfactory brain regions.
Optical imaging of the activity of OSNs, LNs and PNs in the antennal lobe - traditionally using synthetic calcium indicators (e.g. calcium green, FURA-2) or voltage-sensitive dyes (e.g. RH414) - has long been an important technique to understand how olfactory stimuli are represented as spatial and temporal patterns of glomerular activity in many species of insects6-10. Development of genetically-encoded neural activity reporters, such as the fluorescent calcium indicators G-CaMP11,12 and Cameleon13, the bioluminescent calcium indicator GFP-aequorin14,15, or a reporter of synaptic transmission, synapto-pHluorin16 has made the olfactory system of the fruitfly, Drosophila melanogaster, particularly accessible to neurophysiological imaging, complementing its comprehensively-described molecular, electrophysiological and neuroanatomical properties2,4,17. These reporters can be selectively expressed via binary transcriptional control systems (e.g. GAL4/UAS18, LexA/LexAop19,20, Q system21) in defined populations of neurons within the olfactory circuitry to dissect with high spatial and temporal resolution how odor-evoked neural activity is represented, modulated and transformed22-24.
Here we describe the preparation and analysis methods to measure odor-evoked responses in the Drosophila antennal lobe using G-CaMP25-27. The animal preparation is minimally invasive and can be adapted to imaging using wide-field fluorescence, confocal and two-photon microscopes.

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

We describe an established technique to measure and analyze odor-evoked calcium responses in the antennal lobe of living Drosophila melanogaster.

Koku-uyarılmış Cevaplarının Kalsiyum Görüntüleme Drosophila Antennal Lob

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory ...

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.

Elektrofizyolojik Ölçümler ve İnsan Bebeklerde nosisepsiyon Analizi

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

Rapid Determination of the Thermal Nociceptive Threshold in Diabetic Rats

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity to normally innocuous stimuli1. Hyperalgesia and allodynia have been studied in many different rodent models of diabetes mellitus2. However, as stated by B&ouml;lcskei et al, determination of "pain" in animal models is challenging due to its subjective nature3. Moreover, the traditional methods used to determine behavioral responses to noxious thermal stimuli usually lack reproducibility and pharmacological sensitivity3. For instance, by using the hot-plate method of Ankier4, flinch, withdrawal and/or licking of either hind- and/or fore-paws is quantified as reflex latencies at constant high thermal stimuli (52-55 &deg;C). However, animals that are hyperalgesic to thermal stimulus do not reproducibly show differences in reflex latencies using those supra-threshold temperatures3,5. As the recently described method of B&ouml;lcskei et al.6, the procedures described here allows for the rapid, sensitive and reproducible determination of thermal nociceptive thresholds (TNTs) in mice and rats. The method uses slowly increasing thermal stimulus applied mostly to the skin of mouse/rat plantar surface. The method is particularly sensitive to study anti-nociception during hyperalgesic states such as PDN. The procedures described bellow are based on the ones published in detail by Alm&aacute;si et al 5 and B&ouml;lcskei et al 3. The procedures described here have been approved the Laboratory Animal Care and Use Committee (LACUC), Wright State University.

Here, we describe a rapid reliable and simple procedure to determine the lowest temperature at which rats or mice show nocifensive behavior, i.e. the thermal nociceptive threshold (TNT). This method applies a slowly increasing thermal stimulus allowing precise and reproducible estimation of TNTs with minimum, if any, stress to the animals.

Diyabetik Sıçanlarda Termal Nosiseptif Eşik Hızlı Belirlenmesinde

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity ...

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees (Apis mellifera)

This video demonstrates novel techniques of RNA interference (RNAi) which downregulate two genes simultaneously in honey bees using double-stranded RNA (dsRNA) injections. It also presents a protocol of proboscis extension response (PER) assay for measuring gustatory perception.	
	RNAi-mediated gene knockdown is an effective technique downregulating target gene expression. This technique is usually used for single gene manipulation, but it has limitations to detect interactions and joint effects between genes. In the first part of this video, we present two strategies to simultaneously knock down two genes (called double gene knockdown). We show both strategies are able to effectively suppress two genes, vitellogenin (vg) and ultraspiracle (usp), which are in a regulatory feedback loop. This double gene knockdown approach can be used to dissect interrelationships between genes and can be readily applied in different insect species.	
	The second part of this video is a demonstration of proboscis extension response (PER) assay in honey bees after the treatment of double gene knockdown. The PER assay is a standard test for measuring gustatory perception in honey bees, which is a key predictor for how fast a honey bee's behavioral maturation is. Greater gustatory perception of nest bees indicates increased behavioral development which is often associated with an earlier age at onset of foraging and foraging specialization in pollen. In addition, PER assay can be applied to identify metabolic states of satiation or hunger in honey bees. Finally, PER assay combined with pairing different odor stimuli for conditioning the bees is also widely used for learning and memory studies in honey bees.

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees Apis mellifera

In this protocol, we describe two strategies that simultaneously suppress two genes (double gene knockdown) in honey bees. Then we present how to use the proboscis extension response (PER) assay to study the effect of double gene knockdown on honey bee gustatory perception.

Bal Arısı RNAi-aracılı Çift Gen demonte ve tat Algı Ölçüm (<em&gt; Apis mellifera</em&gt;)

This video demonstrates novel techniques of RNA  interference (RNAi) which downregulate two genes simultaneously in honey bees  using double-stranded RNA ...

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize and to study. Therefore, bees have long been a favored model amongst neurobiologists and neuroethologists for studying the neural basis of social and natural behavior. It is important, however, that the experimental techniques used to study bees do not interfere with the behaviors being studied. Because of this, it has been necessary to develop a range of techniques for pharmacological manipulation of honey bees. In this paper we demonstrate methods for treating restrained or free-flying honey bees with a wide range of pharmacological agents. These include both noninvasive methods such as oral and topical treatments, as well as more invasive methods that allow for precise drug delivery in either systemic or localized fashion. Finally, we discuss the advantages and disadvantages of each method and&#160;describe common hurdles and how to best overcome them. We conclude with a discussion on the importance of adapting the experimental method to the biological questions rather than the other way around.

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

This manuscript describes several protocols for administering pharmacological agents to honey bees, including simple noninvasive methods for free-flying bees, as well as more invasive variants that allow precise localized treatment of restrained bees.

Ölçülü ve Serbest uçan Bal Arısı Nörofarmakolojik Manipülasyon,<em&gt; Apis mellifera</em

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize ...

The Use of Trace Eyeblink Classical Conditioning to Assess Hippocampal Dysfunction in a Rat Model of Fetal Alcohol Spectrum Disorders

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain undergoes rapid organizational change and is similar to accelerated brain changes that occur during the third trimester in humans. This model of fetal alcohol spectrum disorders (FASDs) produces severe brain damage, mimicking the amount and pattern of binge-drinking that occurs in some pregnant alcoholic mothers. We describe the use of trace eyeblink classical conditioning (ECC), a higher-order variant of associative learning, to assess long-term hippocampal dysfunction that is typically seen in alcohol-exposed adult offspring. At 90 days of age, rodents were surgically prepared with recording and stimulating electrodes, which measured electromyographic (EMG) blink activity from the left eyelid muscle and delivered mild shock posterior to the left eye, respectively. After a 5 day recovery period, they underwent 6 sessions of trace ECC to determine associative learning differences between alcohol-exposed and control rats. Trace ECC is one of many possible ECC procedures that can be easily modified using the same equipment and software, so that different neural systems can be assessed. ECC procedures in general, can be used as diagnostic tools for detecting neural pathology in different brain systems and different conditions that insult the brain.

Trace eyeblink classical conditioning (ECC) was used to assess hippocampal-dependent associative learning in adult rats that were administered a high concentration (11.9% v/v) of alcohol during early neonatal brain development. In general, ECC procedures are sound diagnostic tools for detecting brain dysfunction across many psychological and biomedical settings.

Hipokampal disfonksiyon Fetal alkol spektrum bozuklukları, fare modeli değerlendirmek için izleme Eyeblink Klasik koşullanma kullanımı

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain ...

Drosophila Courtship Conditioning As a Measure of Learning and Memory

Many insights into the molecular mechanisms underlying learning and memory have been elucidated through the use of simple behavioral assays in model organisms such as the fruit fly, Drosophila melanogaster. Drosophila is useful for understanding the basic neurobiology underlying cognitive deficits resulting from mutations in genes associated with human cognitive disorders, such as intellectual disability (ID) and autism. This work describes a methodology for testing learning and memory using a classic paradigm in Drosophila known as courtship conditioning. Male flies court females using a distinct pattern of easily recognizable behaviors. Premated females are not receptive to mating and will reject the male&#39;s copulation attempts. In response to this rejection, male flies reduce their courtship behavior. This learned reduction in courtship behavior is measured over time, serving as an indicator of learning and memory. The basic numerical output of this assay is the courtship index (CI), which is defined as the percentage of time that a male spends courting during a 10 min interval. The learning index (LI) is the relative reduction of CI in flies that have been exposed to a premated female compared to na&#239;ve flies with no previous social encounters. For the statistical comparison of LIs between genotypes, a randomization test with bootstrapping is used. To illustrate how the assay can be used to address the role of a gene relating to learning and memory, the pan-neuronal knockdown of Dihydroxyacetone phosphate acyltransferase (Dhap-at) was characterized here. The human ortholog of Dhap-at, glyceronephosphate O-acyltransferase (GNPT), is involved in rhizomelic chondrodysplasia punctata type 2, an autosomal-recessive syndrome characterized by severe ID. Using the courtship conditioning assay, it was determined that Dhap-at is required for long-term memory, but not for short-term memory. This result serves as a basis for further investigation of the underlying molecular mechanisms.

Drosophila Courtship Conditioning As a Measure of Learning and Memory

This protocol describes a Drosophila learning and memory assay called courtship conditioning. This classic assay is based on a reduction of male courtship behavior after sexual rejection by a non-receptive premated female. This natural form of behavioral plasticity can be used to test learning, short-term memory, and long-term memory.

Öğrenme ve Bellek Tedrisi Olan Drosophila Kurbanlığı İklimlendirme

Many insights into the molecular mechanisms underlying learning and memory have been elucidated through the use of simple behavioral assays in model ...