This work was supported by a CIHR operating grant (MOP-102624) to Q. Y. We thank Dr Carolyn Harley for helpful discussions throughout the study, Dr. Qinlong Hou, Amin Shakhawat, and Andrea Darby-King for technical support.

Her iki cinsiyette Sprague Dawley sıçan (Charles River) yavrular kullanılır. Litters (doğum PD0 olmak) PD1&#39;in 12 on itlaf edilir. Baraj gıda ve suya ad libitum giriş ile, 12 saat aydınlık / karanlık döngüsünde tutulur. Deneysel prosedürler Memorial Üniversitesi Kurumsal Hayvan Bakım Komitesi tarafından onaylanmıştır. 1. Burun Tak İnşaatı NOT:. Bu prosedür adapte Cummings ve ark modifiye edildi (1997) 35. <ol><li> Kazandırılacak polietilen-20 tüp ve 3-0 ipek sütür konu. </li><li> Yaklaşık olarak 0.8 mm olan polietilen boru 20 küçük bir parça kesin. </li><li> Iplik boru bölümünün her iki yanı üzerinde olduğu bir şekilde hazırlanan boru içinden Konu ipek dikiş ipliği. </li><li> Tıpanın dış ipliğin bir ucunda, ipliğin bir düğüm. </li><li> Iplik düğüm üzerinde aşağı tüp bölümünü çekin. Düğüm boru içine başvurusunda gerekir.</li><li> Ipliğin yaklaşık 2 mm hortumunun bir ucundan dışarı çıkıntı yapan şekilde, ipliğin her iki ucuna da trimerler (Şekil 1A bakınız). </li></ol> Eğitim öncesi 2. Naris Tıkanıklığı <ol><li> Düzenli yatak ile kaplı güvenli bir çanak baraj ve yerden yavru çıkarın. </li><li> Naris üzerine, lokal anestezik jöle,% 2 lidokain kurulamak için bir pamuk uçlu uygulamayı kullanmak tıkanacak. </li><li> Yavru ~ 3 dakika boyunca çanak dinlenmeye bırakın. </li><li> Baskın olmayan elle yavaşça ama güvenli yavru tutun. </li><li> Dominant eli kullanarak, burun fiş almak ve iplik çıkıntı değil hangi ucu etrafında aynı lokal anestezik jel hafifçe sürün. Bu fiş ekleme ile ilgili herhangi bir minör ağrıları için hem bir anestezik olarak hareket edecek ve naris içinde bir yağlayıcı olarak hareket edecektir. NOT: Xylocaine etkisi birkaç dakika içinde başlar ve 20-30 dakika sürer. Xylocaine jöle uygulamadan sonra fiş yerleştirilmesi için genel göstermek iyi tolerans yavrular (myayilan mücadele ve seslendirme). </li><li> Yavaşça sıkıca yavru tutan ve Fişin tamamen takıldığını ve sadece 2 mm diş naris çıkıntı kadar yavaş yavaş çok nazik itişlerle fişi çevirerek burun fişini takın (Şekil 1B bakınız). Bu işlem sırasında ya da herhangi bir kanama naris olmalıdır. Burun fiş yerleştirilmesi sırasında kanama ile yavrular hariç ve barajlar iade edilir </li><li> Hayvan tıpa alıştırmak amacıyla 5 dakika için bu çanak dinlenmeye bırakın. </li><li> Alışma çanak yavru çıkarın ve klima paradigmayı 24 başlar. </li></ol><img alt="Şekil 1" fo:content-width="5in" src="/files/ftp_upload/51808/51808fig1highres.jpg" width="500" /> Bir burun fiş Şekil 1. İnşaat. A) Şemalar bir burun fişi yapma aşamalarını gösteren. Bir iş parçacığı polietilen boru ile çekilir; Bir düğüm yapılan ve pul edilirBunu engellemek için, borunun ortasına içine bırakılır; iki ucu) boru. b üzerinden bir ucunda 2 mm&#39;lik bir tortu ile ön ve bir naris bir burun fişi ile bir sıçan lateral bir görünüşünü kesilir. 3. Kokulu Yatak Hazırlama <ol><li> Yeni eldiven giyen ve koku bulaşmayı önlemek bir plastik torba içine woodchip yatak 500 ml yerleştirmek için bir davlumbaz. </li><li> 0.3 ml nane özü hazırlamak için bir şırınga kullanın ve plastik torba içinde yatak üzerinde bu sprey. </li><li> , Çanta kapalı kravat şiddetle çanta sallayın ve yatak 5 dakika boyunca torbaya dinlenmeye bırakın. </li><li> Kullanmadan önce, 5 dakika boyunca bir çeker ocak içinde ortaya çıkarılan bir net, sığ, akrilik eğitim kutusu (20 x 20 x 5 cm 3, Şekil 2A) olarak kokulu yatak yerleştirin. Yatak hazırlandıktan sonra, bu eldiven atmak ve bu eldivenler hayvanlarla temas etmesine izin vermeyin. </li><li> Özdeş bir şeffaf plastik kutu içinde kokusuz yatak yerleştirin, ve eno kokulu yatak veya kullanılan eldiven ile temas etmediğinden emin olun. </li></ol> 4. Koku Klima Paradigma (Şekil 2A resmi görmek) Yavrular PH 6, ya da birden çok deneme seansları (günde bir seans, PD 3-6) üzerinde, ya tek bir klima oturumu tabi. <ol><li> Kokulu yatak üzerinde habitüe yavru yerleştirin. Koku kontrolü sadece (O / S -) için yavrular, daha sonra 4,5 adıma atlayın, 10 dakika boyunca yatak üzerinde bu yavrular bırakın. Deneysel koku + inme (O / S +) yavrular, bu bölümde aşağıdaki adımlarla devam ediyor. </li><li> İnme 30 saniye boyunca yavru küçük bir fırça kullanarak. Öncelikle yavru arka bölgenin çevresinde hızlı dairesel hareketlerle kullanın. </li><li> Yavru 30 saniye boyunca dinlenmeye bırakın. </li><li> Tekrar 4.2 ve 10 dakika olmak üzere toplam 4,3 (okşayarak + koku yani 10 eşleşmeleri) adım. </li><li> , Klima kutusundan yavru çıkarın burun fişini çıkarın ve baraj yavru dönmek. </li></ol> &lt;p class = &quot;jove_title&quot;&gt; 5. Lateralize Koku Tercih Test (Şekil 2B resmi görmek) Test, son antrenman sonra çeşitli zaman noktalarında (örneğin, 24 ya da 48 saat) oluşur. Test 2 cm nötr bölge ile ayrılmış iki eğitim kasası (eğitim kutu 3,4 tarif edilmiştir), üst üste yerleştirilir, paslanmaz çelik bir test bölmesinin (30 x 20 x 18 cm3) içinde gerçekleştirilir. Diğer kutusu temiz, kokusuz yatak içerir iken bir eğitim kutu nane kokulu yataklar içerir. Test bölmesinin tabanı daha sonra plastik örgü çıkarılabilir bir levha (Şekil 2B) ile kaplanmış olan bir metal ızgara vardır. <ol><li> Bir nane ve Bölüm 3 başına bir kokusuz yatak hazırlayın ve 2 cm aralıklarla test odasının, ters taraflarında altında her bir kutu yerleştirin. Test odasının metal ızgaralı zemin üzerine plastik örgü yerleştirin. </li><li> Baraj yavru çıkarın ve naris üzerinde kokusuz silikon gres bir firma dab koyunBu eğitim sırasında tıkanmaktadır. Yeniden başvuruda gerektiği gibi ilk test boyunca gres. NOT: eğitim ve test sırasında Rastgele naris tıkanıklığı önyargı önlemek için kabul edilebilir. </li><li> Test bölmesinin nötr bölgedeki yavru yerleştirin. </li><li> Yavru odasının iki tarafın üzerinden geçirdim ne kadar süre kayıt, yavru 1 dakika odasını keşfetmek için izin (yani nane veya tarafsız kokulu yatak üzerinde). </li><li> Yavru bir kapalı plastik tutma odasının 1 dakika dinlenmeye bırakın. </li><li> Tekrar 10 dakika toplam 5,2 ve 5,3 adım (5 ile kalan denemeleri ile ayrılan 5 testi denemeleri) yönünde tercihlerini kontrol etmek amacıyla oda içindeki yavru bir başlangıç ​​yönlenmesini anahtarlama. </li><li> Testin hemen ardından, naris gelen yağı silin. </li><li> Bölüm 2&#39;deki gibi ters naris, bir polietilen noseplug yerleştirin ve hayvan 10 dakika boyunca dinlenmeye bırakın. </li><li> Olarak bir kez daha test yavru<html> 5.3 -5.6, kapağı çıkarıp, baraja yavru dönmek. Çıkarın ve% 95 etanol ile test odasının plastik örgü temizlemek ve sıvı buharlaşmasına izin verin. Sonraki yavru test etmeden önce geri örgü yerleştirin. NOT: test sırasında ilk naris oklüzyon silikon gres uygulanması, kanama ve burun fiş ekleme ile ilişkili stres şansını engeller. </li></ol><img alt="Şekil 2" fo:content-width="5in" src="/files/ftp_upload/51808/51808fig2highres.jpg" width="500" /> Şekil 2. Erken koku tercihi eğitim ve test. Paradigma okşayarak + koku kullanarak A) Erken koku tercih çalışması. B) bir tarafta nane yatak ile iki seçenek koku tercihini test, karşı tarafta yatak kokusuz kontrol eder. 2 cm nötr bölge arasına yerleştirilir. 6. Tek Naris Oklüzyonu Etkinliğinin Test nt &quot;&gt; Bu deney, tek naris tıkanma koku alma sisteminin lateralizan aktivasyonuna neden olup olmadığını belirlemek için yapılır. <ol><li> Bölüm 2&#39;de tarif edildiği gibi 6 veya PH 7 yavruların tek taraflı naris oklüzyonlarını gerçekleştirin. </li><li> ~ 5 dk alışma sonra, bir kapalı plastik kapta yavru koyun ve 10 dakika boyunca doku bir parça batırılmış 30 ul saf nane yağı maruz kalmaktadır. </li><li> Hemen nane kokusu maruz kaldıktan sonra, periton boşluğu içine (ip) dozunda genel anestezik olarak hidrat (400 mg / kg) ya da pentobarbitolün (150 mg / kg) ile yavru enjekte edilir. </li><li> Tam (baştan kuyruğa veya ayak tutam yanıt vermedikleri) anestezi sonra, transkardiyal 0.1 M fosfat tampon çözeltisi, PBS içinde çözüldü paraformaldehid (% 4, ardından ~ 1 dakika boyunca tuzlu su (% 0.9) içinde buz soğukluğunda çözümlerle yavru, serpmek ). </li><li> Daha sonra, trans, paraformaldehid perfüzyon 10 dakika sonra beyin toplamak ve 4 ° C&#39;de bir gece paraformaldehid koyunEk bir 24 saat boyunca, bir sakaroz çözeltisi (PBS içinde% 20) ile beyin fer. </li><li> Bir kriostat ile 30 mikron kalınlığında koronal beyin dilimleri kesin. OB ve PC dilimleri toplayın ve pCREB antikor 21,25,30 standart immünhistokimya boyama izledi jelatin kaplı slaytlar, üzerine monte. </li></ol> 7. Tek Naris Oklüzyonu reversibilite testi Bloke edici etkisi burun tıkacın çıkarılmasının ardından 24 saat sonra geri olup bu deney testleri. <ol><li> Bölüm 2&#39;de tarif edildiği gibi 6 veya PH 7 yavruların tek taraflı naris oklüzyonlarını gerçekleştirin. </li><li> (Eğitim sırasında tıkanıklığı naris süresine eşdeğer - 5 dk alışma + 10 dk eğitim) 15 dakika sonra, burun kapağı çıkarıp, baraja yavru dönmek. </li><li> 6.2 de tarif edildiği gibi 24 saat sonra, 10 dakika süre ile kapalı bir plastik kap içine nane kokusuna maruz yavru. </li><li> Mezhep aynı adımları izleyin6,3-6,6 iyonlar bulunmaktadır. </li></ol>

<ol>
	<li>Gregory, E. H., Pfaff, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+olfactory-guided+behavior+in+infant+rats.">Development of olfactory-guided behavior in infant rats.</a> Physiol Behav. 6, 573-576 (1971).</li><li>Alberts, J. R., May, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nonnutritive,+thermotactile+induction+of+filial+huddling+in+rat+pups.">Nonnutritive, thermotactile induction of filial huddling in rat pups.</a> Dev Psychobiol. 17, 161-181 (1984).</li><li>Galef, B. G., Kaner, H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Establishment+and+maintenance+of+preference+for+natural+and+artificial+olfactory+stimuli+in+juvenile+rats.">Establishment and maintenance of preference for natural and artificial olfactory stimuli in juvenile rats.</a> J Comp Physiol Psychol. 94, 588-595 (1980).</li><li>Johanson, I. B., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Appetitive+learning+in+1-day-old+rat+pups.">Appetitive learning in 1-day-old rat pups.</a> Science. 205, 419-421 (1979).</li><li>Johanson, I. B., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Appetitive+conditioning+in+neonatal+rats:+conditioned+orientation+to+a+novel+odor.">Appetitive conditioning in neonatal rats: conditioned orientation to a novel odor.</a> Dev Psychobiol. 15, 379-397 (1982).</li><li>Johanson, I. B., Teicher, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classical+conditioning+of+an+odor+preference+in+3-day-old+rats.">Classical conditioning of an odor preference in 3-day-old rats.</a> Behav Neural Biol. 29, 132-136 (1980).</li><li>McLean, J. H., Darby-King, A., Sullivan, R. M., King, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Serotonergic+influence+on+olfactory+learning+in+the+neonate+rat.">Serotonergic influence on olfactory learning in the neonate rat.</a> Behav Neural Biol. 60, 152-162 (1993).</li><li>Moore, C. L., Power, K. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variation+in+maternal+care+and+individual+differences+in+play,+exploration,+and+grooming+of+juvenile+Norway+rat+offspring.">Variation in maternal care and individual differences in play, exploration, and grooming of juvenile Norway rat offspring.</a> Dev Psychobiol. 25, 165-182 (1992).</li><li>Pedersen, P. E., Williams, C. L., Blass, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activation+and+odor+conditioning+of+suckling+behavior+in+3-day-old+albino+rats.">Activation and odor conditioning of suckling behavior in 3-day-old albino rats.</a> J Exp Psychol Anim Behav Process. 8, 329-341 (1982).</li><li>Sullivan, R. M., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reinforcers+in+infancy:+classical+conditioning+using+stroking+or+intra-oral+infusions+of+milk+as+UCS.">Reinforcers in infancy: classical conditioning using stroking or intra-oral infusions of milk as UCS.</a> Dev Psychobiol. 21, 215-223 (1988).</li><li>Sullivan, R. M., Leon, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+olfactory+learning+induces+an+enhanced+olfactory+bulb+response+in+young+rats.">Early olfactory learning induces an enhanced olfactory bulb response in young rats.</a> Brain Res. 392, 278-282 (1986).</li><li>Weldon, D. A., Travis, M. L., Kennedy, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Posttraining+D1+receptor+blockade+impairs+odor+conditioning+in+neonatal+rats.">Posttraining D1 receptor blockade impairs odor conditioning in neonatal rats.</a> Behav Neurosci. 105, 450-458 (1991).</li><li>Sullivan, R. M., Hofer, M. A., Brake, S. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory-guided+orientation+in+neonatal+rats+is+enhanced+by+a+conditioned+change+in+behavioral+state.">Olfactory-guided orientation in neonatal rats is enhanced by a conditioned change in behavioral state.</a> Dev Psychobiol. 19, 615-623 (1986).</li><li>Camp, L. L., Rudy, 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=Changes+in+the+categorization+of+appetitive+and+aversive+events+during+postnatal+development+of+the+rat.">Changes in the categorization of appetitive and aversive events during postnatal development of the rat.</a> Dev Psychobiol. 21, 25-42 (1988).</li><li>Moriceau, S., Wilson, D. A., Levine, S., Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dual+circuitry+for+odor-shock+conditioning+during+infancy:+corticosterone+switches+between+fear+and+attraction+via+amygdala.">Dual circuitry for odor-shock conditioning during infancy: corticosterone switches between fear and attraction via amygdala.</a> J Neurosci. 26, 6737-6748 (2006).</li><li>Roth, T. L., Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endogenous+opioids+and+their+role+in+odor+preference+acquisition+and+consolidation+following+odor-shock+conditioning+in+infant+rats.">Endogenous opioids and their role in odor preference acquisition and consolidation following odor-shock conditioning in infant rats.</a> Dev Psychobiol. 39, 188-198 (2001).</li><li>Roth, T. L., Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Consolidation+and+expression+of+a+shock-induced+odor+preference+in+rat+pups+is+facilitated+by+opioids.">Consolidation and expression of a shock-induced odor preference in rat pups is facilitated by opioids.</a> Physiol Behav. 78, 135-142 (2003).</li><li>Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developing+a+sense+of+safety:+the+neurobiology+of+neonatal+attachment.">Developing a sense of safety: the neurobiology of neonatal attachment.</a> Ann N Y Acad Sci. 1008, 122-131 (2003).</li><li>Wilson, D. A., Sullivan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+associative+conditioning+in+infant+rats+with+brain+stimulation+as+reward.+I.+Neurobehavioral+consequences.">Olfactory associative conditioning in infant rats with brain stimulation as reward. I. Neurobehavioral consequences.</a> Brain Res Dev Brain Res. 53, 215-221 (1990).</li><li>Sullivan, R. M., Wilson, D. A., Leon, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Associative+Processes+in+Early+Olfactory+Preference+Acquisition:+Neural+and+Behavioral+Consequences.">Associative Processes in Early Olfactory Preference Acquisition: Neural and Behavioral Consequences.</a> Psychobiology. , 29-33 (1989).</li><li>McLean, J. H., Harley, C. W., Darby-King, A., Yuan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=pCREB+in+the+neonate+rat+olfactory+bulb+is+selectively+and+transiently+increased+by+odor+preference-conditioned+training.">pCREB in the neonate rat olfactory bulb is selectively and transiently increased by odor preference-conditioned training.</a> Learn Mem. 6, 608-618 (1999).</li><li>Sullivan, R. M., Stackenwalt, G., Nasr, F., Lemon, C., Wilson, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Association+of+an+odor+with+activation+of+olfactory+bulb+noradrenergic+beta-receptors+or+locus+coeruleus+stimulation+is+sufficient+to+produce+learned+approach+responses+to+that+odor+in+neonatal+rats.">Association of an odor with activation of olfactory bulb noradrenergic beta-receptors or locus coeruleus stimulation is sufficient to produce learned approach responses to that odor in neonatal rats.</a> Behav Neurosci. 114, 957-962 (2000).</li><li>Yuan, Q., Harley, C. W., McLean, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mitral+cell+beta1+and+5-HT2A+receptor+colocalization+and+cAMP+coregulation:+a+new+model+of+norepinephrine-induced+learning+in+the+olfactory+bulb.">Mitral cell beta1 and 5-HT2A receptor colocalization and cAMP coregulation: a new model of norepinephrine-induced learning in the olfactory bulb.</a> Learn Mem. 10, 5-15 (2003).</li><li>Fontaine, C. J., Harley, C. W., Yuan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lateralized+odor+preference+training+in+rat+pups+reveals+an+enhanced+network+response+in+anterior+piriform+cortex+to+olfactory+input+that+parallels+extended+memory.">Lateralized odor preference training in rat pups reveals an enhanced network response in anterior piriform cortex to olfactory input that parallels extended memory.</a> J Neurosci. 33, 15126-15131 (2013).</li><li>Morrison, G. L., Fontaine, C. J., Harley, C. W., Yuan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+role+for+the+anterior+piriform+cortex+in+early+odor+preference+learning:+evidence+for+multiple+olfactory+learning+structures+in+the+rat+pup.">A role for the anterior piriform cortex in early odor preference learning: evidence for multiple olfactory learning structures in the rat pup.</a> J Neurophysiol. 110, 141-152 (2013).</li><li>Nakamura, S., Kimura, F., Sakaguchi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postnatal+development+of+electrical+activity+in+the+locus+ceruleus.">Postnatal development of electrical activity in the locus ceruleus.</a> J Neurophysiol. 58, 510-524 (1987).</li><li>Harley, C. W., Darby-King, A., McCann, J., McLean, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beta1-adrenoceptor+or+alpha1-adrenoceptor+activation+initiates+early+odor+preference+learning+in+rat+pups:+support+for+the+mitral+cell/cAMP+model+of+odor+preference+learning.">Beta1-adrenoceptor or alpha1-adrenoceptor activation initiates early odor preference learning in rat pups: support for the mitral cell/cAMP model of odor preference learning.</a> Learn Mem. 13, 8-13 (2006).</li><li>Shakhawat, A. M., Harley, C. W., Yuan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+bulb+alpha2-adrenoceptor+activation+promotes+rat+pup+odor-preference+learning+via+a+cAMP-independent+mechanism.">Olfactory bulb alpha2-adrenoceptor activation promotes rat pup odor-preference learning via a cAMP-independent mechanism.</a> Learn Mem. 19, 499-502 (2012).</li><li>Isaacson, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Odor+representations+in+mammalian+cortical+circuits.">Odor representations in mammalian cortical circuits.</a> Curr Opin Neurobiol. 20, 328-331 (2010).</li><li>Lethbridge, R., Hou, Q., Harley, C. W., Yuan, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+bulb+glomerular+NMDA+receptors+mediate+olfactory+nerve+potentiation+and+odor+preference+learning+in+the+neonate+rat.">Olfactory bulb glomerular NMDA receptors mediate olfactory nerve potentiation and odor preference learning in the neonate rat.</a> PLoS One. 7, e35024 (2012).</li><li>Yuan, Q., Harley, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=What+a+nostril+knows:+olfactory+nerve-evoked+AMPA+responses+increase+while+NMDA+responses+decrease+at+24-h+post-training+for+lateralized+odor+preference+memory+in+neonate+rat.">What a nostril knows: olfactory nerve-evoked AMPA responses increase while NMDA responses decrease at 24-h post-training for lateralized odor preference memory in neonate rat.</a> Learn Mem. 19, 50-53 (2012).</li><li>Schwob, J. E., Price, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+of+axonal+connections+in+the+central+olfactory+system+of+rats.">The development of axonal connections in the central olfactory system of rats.</a> J Comp Neurol. 223, 177-202 (1984).</li><li>Kucharski, D., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+routes+to+early+memories.">New routes to early memories.</a> Science. 238, 786-788 (1987).</li><li>Kucharski, D., Johanson, I. B., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unilateral+olfactory+conditioning+in+6-day-old+rat+pups.">Unilateral olfactory conditioning in 6-day-old rat pups.</a> Behav Neural Biol. 46, 472-490 (1986).</li><li>Cummings, D. M., Henning, H. E., Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+bulb+recovery+after+early+sensory+deprivation.">Olfactory bulb recovery after early sensory deprivation.</a> J Neurosci. 17, 7433-7440 (1997).</li><li>Kucharski, D., Hall, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+change+in+the+access+to+olfactory+memories.">Developmental change in the access to olfactory memories.</a> Behav Neurosci. 102, 340-348 (1988).</li><li>Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unilateral+odor+deprivation:+time+course+of+changes+in+laminar+volume.">Unilateral odor deprivation: time course of changes in laminar volume.</a> Brain Res Bull. 14, 233-237 (1985).</li><li>Kass, M. D., Pottackal, J., Turkel, D. J., McGann, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Changes+in+the+neural+representation+of+odorants+after+olfactory+deprivation+in+the+adult+mouse+olfactory+bulb.">Changes in the neural representation of odorants after olfactory deprivation in the adult mouse olfactory bulb.</a> Chem Senses. 38, 77-89 (2013).</li><li>Kim, H. H., Puche, A. C., Margolis, F. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Odorant+deprivation+reversibly+modulates+transsynaptic+changes+in+the+NR2B-mediated+CREB+pathway+in+mouse+piriform+cortex.">Odorant deprivation reversibly modulates transsynaptic changes in the NR2B-mediated CREB pathway in mouse piriform cortex.</a> J Neurosci. 26, 9548-9559 (2006).</li><li>Korol, D. L., Brunjes, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+changes+in+2-deoxyglucose+uptake+and+amino+acid+incorporation+following+unilateral+odor+deprivation:+a+laminar+analysis.">Rapid changes in 2-deoxyglucose uptake and amino acid incorporation following unilateral odor deprivation: a laminar analysis.</a> Brain Res Dev Brain Res. 52, 75-84 (1990).</li><li>Leung, C. H., Wilson, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trans-neuronal+regulation+of+cortical+apoptosis+in+the+adult+rat+olfactory+system.">Trans-neuronal regulation of cortical apoptosis in the adult rat olfactory system.</a> Brain Res. 984, 182-188 (2003).</li></ol>

The authors have nothing to disclose.

Kritik bir zaman penceresi içinde sıçan yavrularında lateralizan koku, öğrenme ve bellek modeli ilk Hall ve arkadaşları tarafından kurulmuştur. Çalışmaların 33,34,36 bir dizi, onlar bir koku tercih bellek sıçan yavrularının PH 6 az bir naris için koku + süt eşleşmeleri tarafından lateralize olabileceğini gösterdi. Tercih bellek aynı naris eğitim ve test sırasında açık olduğunda sağlam, ancak tıkalı naris bloke edilmemiş ve kontrol edilmiş olduğu zaman gözlemlenmemiştir....

Olfaction başarıyla gezinmek veya kendi ortamında hayatta kalmak için mümkün olmaz onsuz kemirgenlerde primer duyu yöntemidir. Bu 1 beslemek için annelerini bulmak için koku alma duyusunu kullanmak için, ilk doğum sonrası hafta boyunca görmeniz ne duymak ne yenidoğan yavrular, için özellikle önemlidir. Sonuç olarak, yenidoğan sıçan yavrular basit deneysel manipülasyonlar ile kokuları tercih klimalı edilebilir. Uyaranların çeşitli iç içe geçme çevre 2,3, süt emzirme 4-6, okşayarak ve dokunma uyarısına 7 de dahil olmak üzere yeni yenidoğanlarda kokulara koşullandırılmış tepkileri (Koşullu uyarı, TH), indükleme koşulsuz uyarıcı (UCS) olarak kullanılmaktadır 12, kuyruk tutam 13, anne tükürük 13, hafif bir ayak şok 14-18, ve intrakranial beyin stimülasyonu 19. Bu çalışmada köklü bir erken koku tercihi paradigma istihdam edildiği bu durumda nane bir koku, bennane, 24 saat sonra 10,11,20 için bir tercih üretilmesi amacıyla dokunma uyarısına birlikte s. Bu kokular hafızası esas olarak koku ampuller (OB) 21-23 ve ön piriform korteks (APC), 24,25 de dahil olmak üzere, sağlam koku devresi bağlıdır. Erken koku tercihi öğrenme Deneysel araştırmalar derinleşti ve bir memeli hafıza moleküler ve fizyolojik temellerinin anlayışımızı genişlettik. Bu memeli model, bellek mekanizmaları okuyan birçok avantajı vardır. İlk olarak, UCS sinyalinin nöral kaynakları tespit edilmiştir. Yukarıda belirtildiği gibi, çeşitli uyarıcılar sırayla 22,27,28 öğrenmeyi desteklemek hücresel ve fizyolojik etkilere neden OB ve aPC&#39;ye birden adrenoreseptörler aktive odağı coeruleus norepinefrin salınımını 26 uyarırlar. İkincisi, bellek destek mekanizmaları iyi tanımlanmış laminer sinir yapıları yer almaktadır.yenidoğan sıçanlarda koku devrenin basitlik sinaptik plastisite ile ilgili karmaşık süreçleri ortaya çıkarmak için birlikte ideal bir çerçeve ile araştırmacılar sağlar. Diğer yapılar 29 arasında OB ve ipsilateral lateral koku yolu (LOT) ile piriform korteks (PC) dönüş projesi çerçevesinde bu mitral / sorguçlu hücrelerin mitral / tüylü hücreler üzerine koku epiteli projede koku alma duyu nöronları (OSN). OSN OB 30,31 sinapsların ve LOT aPC&#39;ye içinde 24,25 sinapsların sinaptik öğrenme destekleyen değişiklikler ve bellek için kritik loci olarak tespit edilmiştir. Üçüncü olarak, sıçanlarda erken yaşta, koku girişler kolayca lateralize edilebilir. Her aPC&#39;ye bu kez beyaz cevher tam post-natal gün 12 (PD12) 32&#39;de oluşturulan ön komissura&#39;da aracılığıyla ikili koku bilgilere erişimi vardır. PD 12 önce, koku girişli tek tıkanıklığı naris 24,25,31,33,34 &lt;aracılığıyla OB ve aPC&#39;nin ipisilateral izole edilebilir/ Sup&gt;. Tek naris tıkanıklığı açık naris gelen koku hafıza oluşumuna olanak ve 12 33 pd önce tıkalı naris aynı bellek önler. Koku hafızası hem OB ve aPC&#39;ye dahil ipsilateral hemisferde izole edilir. Bu nedenle, her sıçan yavru öğrenme ve fizyolojisini destekleyen için kendi kontrol edilebilir. Bu çalışmada, lateralize ilk koku tercih öğrenme protokol verilmektedir. Bu yöntem, böylece gerekli hayvan sayısını ve genel varyasyonu de azaltarak, intra-hayvan kontrolü 24,25,31 sağlayarak koku öğrenmenin temelini nöral mekanizmaları çalışmak için güçlü bir araç olarak hizmet vermektedir. Tıkanıklığı naris gres ya da burun tıkacın uygulanan ve en az stres veya hayvana zarar çıkarılabilir olması ile tersine çevrilebilir. Burada öncelikle, erken koku tercihi eğitim ve test prosedürleri ayrıntılı bir nos tek naris tıkanıklığı kullanarak lateralizan protokolü odaklanarak, tarif edilmektedire fiş. Sonra sonuçlar koku girişini izole ve lateralizan koku belleği üreten tek tıkanıklığı naris etkinliğini göstermek için sunulmaktadır. Son olarak, hem öğrenim hem de destek bellek ifade oluşturmak koku sistemi fizyolojik değişiklikleri incelemek için bu lateralizan öğrenme modelini kullanarak potansiyelleri tartışılmıştır.

<table border="0" cellpadding="0" cellspacing="0" width="580">
	<tbody>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Polythylene 20 tubing</td>
			<td style="width:123px;">Intramedic</td>
			<td style="width:103px;">427406</td>
			<td style="width:205px;">Non radiopaque, Non toxic</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">3-0 silk suture thread</td>
			<td style="width:123px;">Syneture</td>
			<td style="width:103px;">Sofsilk</td>
			<td style="width:205px;">Non absorbant&#160;</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Silicone grease</td>
			<td style="width:123px;">Warner Instrument</td>
			<td style="width:103px;">64-0378</td>
			<td style="width:205px;">Odorless</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">2% xylocaine gel</td>
			<td style="width:123px;">AstraZeneca</td>
			<td style="width:103px;">Prod. No 061</td>
			<td style="width:205px;">Lidocaine hydrochloride jelly,&#160; purchased at local pharmacy</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Paint brush</td>
			<td style="width:123px;">Dynasty</td>
			<td style="width:103px;">206R</td>
			<td style="width:205px;">Similar size/other brands work too</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Peppermint extract</td>
			<td style="width:123px;">Sigma-Aldrich</td>
			<td style="width:103px;">W284807</td>
			<td style="width:205px;">Other brand should be okay too</td>
		</tr>
		<tr height="171">
			<td height="171" style="height:171px;width:149px;">Training box</td>
			<td style="width:123px;">Custom-made</td>
			<td style="width:103px;">N/A</td>
			<td style="width:205px;">Acrylic box (20x20x5cm3), see Figure 2A. Parameters and material for the box are not critical and can be modified. Material used should be odorless and does not absorb odors</td>
		</tr>
		<tr height="182">
			<td height="182" style="height:182px;width:149px;">Testing chamber</td>
			<td style="width:123px;">Custom-made</td>
			<td style="width:103px;">N/A</td>
			<td style="width:205px;">Stainless steel (30x20x18cm3), see Figure 2B. Parameters and material for the chamber are not critical and can be modified. For example, an acrylic chamber instead of a stainless steel one can be used</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">pCREB antibody</td>
			<td style="width:123px;">Cell Signaling</td>
			<td style="width:103px;">9198</td>
			<td style="width:205px;">Ser 133 (87G3) Rabbit mAb</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Chloral hydrate</td>
			<td style="width:123px;">Sigma-Aldrich</td>
			<td style="width:103px;">C8383</td>
			<td style="width:205px;">N/A</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Paraformaldehype</td>
			<td style="width:123px;">Sigma-Aldrich</td>
			<td style="width:103px;">P6148</td>
			<td style="width:205px;">N/A</td>
		</tr>
		<tr height="50">
			<td height="50" style="height:50px;width:149px;">Sucrose</td>
			<td style="width:123px;">Sigma-Aldrich</td>
			<td style="width:103px;">S9378</td>
			<td style="width:205px;">N/A</td>
		</tr>
	</tbody>
</table>

a lateralized odor learning model in neonatal rats for dissecting neural circuitry underpinning memory formation

Rat pups during a critical postnatal period (&#8804; 10 days) readily form a preference for an odor that is associated with stimuli mimicking maternal care. Such a preference memory can last from hours, to days, even life-long, depending on training parameters. Early odor preference learning provides us with a model in which the critical changes for a natural form of learning occur in the olfactory circuitry. An additional feature that makes it a powerful tool for the analysis of memory processes is that early odor preference learning can be lateralized via single naris occlusion within the critical period. This is due to the lack of mature anterior commissural connections of the olfactory hemispheres at this early age. This work outlines behavioral protocols for lateralized odor learning using nose plugs. Acute, reversible naris occlusion minimizes tissue and neuronal damages associated with long-term occlusion and more aggressive methods such as cauterization. The lateralized odor learning model permits within-animal comparison, therefore greatly reducing variance compared to between-animal designs. This method has been used successfully to probe the circuit changes in the olfactory system produced by training. Future directions include exploring molecular underpinnings of odor memory using this lateralized learning model; and correlating physiological change with memory strength and durations.

Burada koku girişini izole ve bir yarımkürede öğrenme tıkanıklığı naris etkinliğini ve bu yöntemin reverzibilitesini göstermek için daha önce kurulmuş sonuçlarının 24 bazı gözden. Erken koku tercihi eğitim sırasında tek naris tıkanıklık lateralizan koku hafızasına 24 yol açar. Bellek korunmuş naris (Şekil 3) ile sınırlıdır. Yavrular eğitimi sırasında olduğu gibi tıkanır aynı naris ile koku tercihi için test ed...

Watch this Scientific Journal Video about A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation at JoVE.com

A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation

This protocol introduces lateralized early odor preference learning in rats using acute single naris occlusion. Lateralized learning permits the examination of behavioral outcomes and underpinning biological mechanisms within the same animals, reducing variance induced by between-animal designs. This protocol can be used to investigate molecular mechanisms underpinning early odor learning.

Sinir Devresi Temelini Bellek Oluşumunda Diseksiyon için Yenidoğan Sıçanlarda bir Lateralize Koku Öğrenme Modeli

Rat pups during a critical postnatal period (&#8804; 10 days) readily form a preference for an odor that is associated with stimuli mimicking maternal ...

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Research

JoVE Journal

Neuroscience

8.9K Görüntüleme.  Faculty of Medicine, Memorial University. This protocol introduces lateralized early odor preference learning in rats using acute single naris occlusion. Lateralized learning permits the examination of behavioral outcomes and underpinning biological mechanisms within the same animals, reducing variance induced by between-animal designs. This protocol can be used to investigate molecular mechanisms underpinning early odor learning.

Video: A Lateralized Odor Learning Model in Neonatal Rats for Dissecting Neural Circuitry Underpinning Memory Formation

C. elegans Positive Butanone Learning, Short-term, and Long-term Associative Memory Assays

The memory of experiences and learned information is critical for organisms to make choices that aid their survival. C. elegans navigates its environment through neuron-specific detection of food and chemical odors1, 2, and can associate nutritive states with chemical odors3, temperature4, and the pathogenicity of a food source5.
Here, we describe assays of C. elegans associative learning and short- and long-term associative memory. We modified an aversive olfactory learning paradigm6 to instead produce a positive response; the assay involves starving ~400 worms, then feeding the worms in the presence of the AWC neuron-sensed volatile chemoattractant butanone at a concentration that elicits a low chemotactic index (similar to Toroyama et al.7). A standard population chemotaxis assay1 tests the worms' attraction to the odorant immediately or minutes to hours after conditioning.
After conditioning, wild-type animals' chemotaxis to butanone increases ~0.6 Chemotaxis Index units, its "Learning Index". Associative learning is dependent on the presence of both food and butanone during training. Pairing food and butanone for a single conditioning period ("massed training") produces short-term associative memory that lasts ~2 hours. Multiple conditioning periods with rest periods between ("spaced training") yields long-term associative memory (&lt;40 hours), and is dependent on the cAMP Response Element Binding protein (CREB),6 a transcription factor required for long-term memory across species.8
Our protocol also includes image analysis methods for quick and accurate determination of chemotaxis indices. High-contrast images of animals on chemotaxis assay plates are captured and analyzed by worm counting software in MatLab. The software corrects for uneven background using a morphological tophat transformation.9 Otsu's method is then used to determine a threshold to separate worms from the background.10 Very small particles are removed automatically and larger non-worm regions (plate edges or agar punches) are removed by manual selection. The software then estimates the size of single worm by ignoring regions that are above a specified maximum size and taking the median size of the remaining regions. The number of worms is then estimated by dividing the total area identified as occupied by worms by the estimated size of a single worm.
We have found that learning and short- and long-term memory can be distinguished, and that these processes share similar key molecules with higher organisms.6,8 Our assays can quickly test novel candidate genes or molecules that affect learning and short- or long-term memory in C. elegans that are relevant across species.

C. elegans Positive Butanone Learning, Short-term, and Long-term Associative Memory Assays

Here we describe methods to test C. elegans associative learning and short- and long-term associative memory. These population assays employ the worms abilities to chemotax toward volatile odorants, and form positive associations upon pairing food with the chemoattractant butanone. Increasing the number of conditioning periods induces long-term memory.

C. elegans Pozitif bütanon Öğrenme, Kısa vadeli ve uzun vadeli ilişkilendirilebilir Bellek Tahliller

The memory of experiences and learned information is critical for organisms to make choices that aid their survival. C. elegans navigates its environment ...

Nörobilim

Assaying Locomotor, Learning, and Memory Deficits in Drosophila Models of Neurodegeneration

Advances in genetic methods have enabled the study of genes involved in human neurodegenerative diseases using Drosophila as a model system1. Most of these diseases, including Alzheimer's, Parkinson's and Huntington's disease are characterized by age-dependent deterioration in learning and memory functions and movement coordination2. Here we use behavioral assays, including the negative geotaxis assay3 and the aversive phototaxic suppression assay (APS assay)4,5, to show that some of the behavior characteristics associated with human neurodegeneration can be recapitulated in flies. In the negative geotaxis assay, the natural tendency of flies to move against gravity when agitated is utilized to study genes or conditions that may hinder locomotor capacities. In the APS assay, the learning and memory functions are tested in positively-phototactic flies trained to associate light with aversive bitter taste and hence avoid this otherwise natural tendency to move toward light. Testing these trained flies 6 hours post-training is used to assess memory functions. Using these assays, the contribution of any genetic or environmental factors toward developing neurodegeneration can be easily studied in flies.

Assaying Locomotor, Learning, and Memory Deficits in Drosophila Models of Neurodegeneration

Behavior assays for measuring locomotor functions, learning, and memory abilities in Drosophila.

Assaying Lokomotor, Öğrenme ve Hafıza Açıkları. Drosophila Nörodejenerasyon, modeller

Advances in genetic methods have enabled the study of genes involved in human neurodegenerative diseases using Drosophila as a model system1. Most of ...

Morris Water Maze Test for Learning and Memory Deficits in Alzheimer&#39;s Disease Model Mice

The Morris Water Maze (MWM) was first established by neuroscientist Richard G. Morris in 1981 in order to test hippocampal-dependent learning, including acquisition of spatial memoryand long-term spatial memory 1. The MWM is a relatively simple procedure typically consisting of six day trials, the main advantage being the differentiation between the spatial (hidden-platform) and non-spatial (visible platform) conditions 2-4. In addition, the MWM testing environment reduces odor trail interference 5. This has led the task to be used extensively in the study of the neurobiology and neuropharmacology of spatial learning and memory. The MWM plays an important role in the validation of rodent models for neurocognitive disorders such as Alzheimer&#x2019;s Disease 6, 7. In this protocol we discussed the typical procedure of MWM for testing learning and memory and data analysis commonly used in Alzheimer&#x2019;s disease transgenic model mice.

Morris Water Maze Test for Learning and Memory Deficits in Alzheimer&#039;s Disease Model Mice

The Morris Water Maze is a behavioral task to test hippocampal-dependent learning and memory. It has been widely used in the study of neurobiology, neuropharmacology and neurocognitive disorders in rodent models.

Alzheimer Hastalığı Model Fareler Öğrenme ve Hafıza Açıkları Morris Su Labirent Testi

The Morris Water Maze (MWM) was first established by neuroscientist Richard G. Morris in 1981 in order to test hippocampal-dependent learning, including ...

Optical Recording of Suprathreshold Neural Activity with Single-cell and Single-spike Resolution

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also propagation of suprathreshold spiking activity involves populations of neurons. Empirical studies addressing cortical function directly thus require recordings from populations of neurons with high resolution. Here we describe an optical method and a deconvolution algorithm to record neural activity from up to 100 neurons with single-cell and single-spike resolution. This method relies on detection of the transient increases in intracellular somatic calcium concentration associated with suprathreshold electrical spikes (action potentials) in cortical neurons. High temporal resolution of the optical recordings is achieved by a fast random-access scanning technique using acousto-optical deflectors (AODs)1. Two-photon excitation of the calcium-sensitive dye results in high spatial resolution in opaque brain tissue2. Reconstruction of spikes from the fluorescence calcium recordings is achieved by a maximum-likelihood method. Simultaneous electrophysiological and optical recordings indicate that our method reliably detects spikes (&gt;97% spike detection efficiency), has a low rate of false positive spike detection (&lt; 0.003 spikes/sec), and a high temporal precision (about 3 msec) 3. This optical method of spike detection can be used to record neural activity in vitro and in anesthetized animals in vivo3,4.

Understanding the function of the vertebrate central nervous system requires recordings from many neurons because cortical function arises on the level of populations of neurons. Here we describe an optical method to record suprathreshold neural activity with single-cell and single-spike resolution, dithered random-access scanning. This method records somatic fluorescence calcium signals from up to 100 neurons with high temporal resolution. A maximum-likelihood algorithm deconvolves the underlying suprathreshold neural activity from the somatic fluorescence calcium signals. This method reliably detects spikes with high detection efficiency and a low rate of false positives and can be used to study neural populations in vitro and in vivo.

Tek hücreli ve tek başak çözünürlüğe sahip supratreshold nöral aktivitenin Optik kaydı

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also ...

Viral Tracing of Genetically Defined Neural Circuitry

Classical methods for studying neuronal circuits are fairly low throughput. Transsynaptic viruses, particularly the pseudorabies (PRV) and rabies virus (RABV), and more recently vesicular stomatitis virus (VSV), for studying circuitry, is becoming increasingly popular. These higher throughput methods use viruses that transmit between neurons in either the anterograde or retrograde direction.
Recently, a modified RABV for monosynaptic retrograde tracing was developed. (Figure 1A). In this method, the glycoprotein (G) gene is deleted from the viral genome, and resupplied only in targeted neurons. Infection specificity is achieved by substituting a chimeric G, composed of the extracellular domain of the ASLV-A glycoprotein and the cytoplasmic domain of the RABV-G (A/RG), for the normal RABV-G1. This chimeric G specifically infects cells expressing the TVA receptor1. The gene encoding TVA can been delivered by various methods2-8. Following RABV-G infection of a TVA-expressing neuron, the RABV can transmit to other, synaptically connected neurons in a retrograde direction by nature of its own G which was co-delivered with the TVA receptor. This technique labels a relatively large number of inputs (5-10%)2 onto a defined cell type, providing a sampling of all of the inputs onto a defined starter cell type.
We recently modified this technique to use VSV as a transsynaptic tracer9. VSV has several advantages, including the rapidity of gene expression. Here we detail a new viral tracing system using VSV useful for probing microcircuitry with increased resolution. While the original published strategies by Wickersham et al.4 and Beier et al.9 permit labeling of any neurons that project onto initially-infected TVA-expressing-cells, here VSV was engineered to transmit only to TVA-expressing cells (Figure 1B). The virus is first pseudotyped with RABV-G to permit infection of neurons downstream of TVA-expressing neurons. After infecting this first population of cells, the virus released can only infect TVA-expressing cells. Because the transsynaptic viral spread is limited to TVA-expressing cells, presence of absence of connectivity from defined cell types can be explored with high resolution. An experimental flow chart of these experiments is shown in Figure 2. Here we show a model circuit, that of direction-selectivity in the mouse retina. We examine the connectivity of starburst amacrine cells (SACs) to retinal ganglion cells (RGCs).

A method of tracing synaptically connected neurons is described. We use TVA specificity of an upstream cell to probe whether a cell population of interest receives synaptic input from genetically defined cell types.

Genetiği Tanımlı Nöral Devre arasında Tracing Viral

Classical methods for studying neuronal circuits are fairly low throughput. Transsynaptic viruses, particularly the pseudorabies (PRV) and rabies virus ...

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution has been a long sought tool for neuroscientists in the systems-level search for the neural circuitry governing complex behavioral states. Optogenetics is a cutting-edge tool that is revolutionizing the field of neuroscience and represents one of the first systematic approaches to enable causal testing regarding the relation between neural signaling events and behavior. By combining optical and genetic approaches, neural signaling can be bi-directionally controlled through expression of light-sensitive ion channels (opsins) in mammalian cells. The current protocol describes delivery of specific wavelengths of light to opsin-expressing cells in deep brain structures of awake, freely-moving rodents for neural circuit modulation. Theoretical principles of light transmission as an experimental consideration are discussed in the context of performing in vivo optogenetic stimulation. The protocol details the design and construction of both simple and complex laser configurations and describes tethering strategies to permit simultaneous stimulation of multiple animals for high-throughput behavioral testing.

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

Optogenetics has become a powerful tool for use in behavioral neuroscience experiments. This protocol offers a step-by-step guide to the design and set-up of laser systems, and provides a full protocol for carrying out multiple and simultaneous in vivo optogenetic stimulations compatible with most rodent behavioral testing paradigms.

Kemirgen Santral Sinir Sistemi vivo optogenetic Stimülasyon içinde

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution ...

Assessing Spatial Learning and Memory in Small Squamate Reptiles

Clinical research has leveraged a variety of paradigms to assess cognitive decline, commonly targeting spatial learning and memory abilities. However, interest in the cognitive processes of nonmodel species, typically within an ecological context, has also become an emerging field of study. In particular, interest in the cognitive processes in reptiles is growing although experimental studies on reptilian cognition are sparse. The few reptilian studies that have experimentally tested for spatial learning and memory have used rodent paradigms modified for use in reptiles. However, ecologically important aspects of the physiology and behavior of this taxonomic group must be taken into account when testing for spatially based cognition. Here, we describe modifications of the dry land Barnes maze and associated testing protocol that can improve performance when probing for spatial learning and memory ability in small squamate reptiles. The described paradigm and procedures were successfully used with male side-blotched lizards (Uta stansburiana), demonstrating that spatial learning and memory can be assessed in this taxonomic group with an ecologically relevant apparatus and protocol.

This paper describes a modification of the Barnes maze, a standard rodent paradigm used to assess spatial memory and learning, for use in small squamate reptiles.

Küçük pul pul Sürüngenler Mekansal Öğrenme ve Bellek değerlendirilmesi

Clinical research has leveraged a variety of paradigms to assess cognitive decline, commonly targeting spatial learning and memory abilities. However, ...

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such as epilepsy and neurodegenerative disorders. However, it is technically difficult to obtain EEG recordings in neonates as it requires specialized handling and great care. Here, we present a novel method to record EEG in neonatal rat pups (P8-P15). We designed a simple and reliable electrode using computer pin loci; it can be easily implanted into the skull of a rat pup to record high-quality EEG signals in the normal and epileptic brain. Pups were given an intraperitoneal (i.p.) injection of the neurotoxin kainic acid (KA) to induce epileptic seizures. The surgical implantation performed in this procedure is less expensive than other EEG procedures for neonates. This method allows one to record high-quality and stable EEG signals for more than 1 week. Furthermore, this procedure can also be applied to adult rats and mice to study epilepsy or other neurological disorders.

Here, we introduce a novel technique designed to record electroencephalography (EEG) in freely moving neonatal epileptic pups and describe its procedures, features, and applications. This method allows one to record EEG for more than 1 week.

Yeni Bir Yöntemi Kullanarak Serbest Dolaşan Yenidoğan Sıçanlarında EEG Kaydı

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

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 ...

In Vitro Wedge Slice Preparation for Mimicking In Vivo Neuronal Circuit Connectivity

In vitro slice electrophysiology techniques measure single-cell activity with precise electrical and temporal resolution. Brain slices must be relatively thin to properly visualize and access neurons for patch-clamping or imaging, and in vitro examination of brain circuitry is limited to only what is physically present in the acute slice. To maintain the benefits of in vitro slice experimentation while preserving a larger portion of presynaptic nuclei, we developed a novel slice preparation. This &#8220;wedge slice&#8221; was designed for patch-clamp electrophysiology recordings to characterize the diverse monaural, sound-driven inputs to medial olivocochlear (MOC) neurons in the brainstem. These neurons receive their primary afferent excitatory and inhibitory inputs from neurons activated by stimuli in the contralateral ear and corresponding cochlear nucleus (CN). An asymmetrical brain slice was designed which is thickest in the rostro-caudal domain at the lateral edge of one hemisphere and then thins towards the lateral edge of the opposite hemisphere. This slice contains, on the thick side, the auditory nerve root conveying information about auditory stimuli to the brain, the intrinsic CN circuitry, and both the disynaptic excitatory and trisynaptic inhibitory afferent pathways that converge on contralateral MOC neurons. Recording is performed from MOC neurons on the thin side of the slice, where they are visualized using DIC optics for typical patch-clamp experiments. Direct stimulation of the auditory nerve is performed as it enters the auditory brainstem, allowing for intrinsic CN circuit activity and synaptic plasticity to occur at synapses upstream of MOC neurons. With this technique, one can mimic in vivo circuit activation as closely as possible within the slice. This wedge slice preparation is applicable to other brain circuits where circuit analyses would benefit from preservation of upstream connectivity and long-range inputs, in combination with the technical advantages of in vitro slice physiology.

Integration of diverse synaptic inputs to neurons is best measured in a preparation that preserves all pre-synaptic nuclei for natural timing and circuit plasticity, but brain slices typically sever many connections. We developed a modified brain slice to mimic in vivo circuit activity while maintaining in vitro experimentation capability.

In Vitro Kama Dilimi In Vivo Nöronal Devre Bağlantısı Nın TaklitI Için Hazırlık

In vitro slice electrophysiology techniques measure single-cell activity with precise electrical and temporal resolution. Brain slices must be relatively ...