Ho fapana hoa bongata ba Oxytocin Receptor Retentionor in the Nucleus Accumbens ho na le liphello tse sa tšoaneng tsa ho ba le maiketsetso a tšebelisano-'moho le batho ba nang le mekhoa e mengata le likhetho tsa sethepu (2009)

J Neurosci. Mongolo o ngotsoeng ka letsoho; e fumanehang ho PMC Oct 27, 2009.

E hatisitsoe ka mokhoa o qetelle o hlophisitsoeng e le:

PMCID: PMC2768419

NIHMSID: NIHMS93410

Phetolelo ea ho qetela ea sengoliloeng sa sengoliloeng sena e fumaneha mahala ho J Neurosci

Sheba lihlooho tse ling ho PMC hore Qotsa sehlooho se hatisitsoeng.

Eya ho:

inahaneloang

Li-receptor tsa Oxetocin ka har'a li-nucleus bokellitsoeng li bile le tšusumetso ho taolo ea boits'oaro bo kopanetsoeng le ho theha maqhama a kopantsoeng tikolohong ea sechaba se le seng sa mokhosi. Tekanyetso ea "oxytocin" receptor density ho "nucleus accumbens" e lumellana hantle le alloparenting ho bana ba banyenyane le ba baholo ba basali ba prairie voles, 'me mohanyetsi oa oksijene ea kenngoeng kelellong o kenelletse molek'huleng ea methapo e thibelang boitšoaro bona. Ntle le moo, li-voeseree tsa prairie li na le methapo e phahameng ea li-oxytocin receptor ho feta mefuta e seng monogamous rodent, le ho thibela li-receptor tse oelengtocin tse thibelang ho rarahana ho hohelang ho ratoa ha molekane. Mona re sebelisitse phetisetso ea vaerase e amanang le vaerase e amanang le adeno ho lekola kamano ea ts'ebetso lipakeng tsa segokahano sa oxytocin receptor density le boitšoaro ba sechaba sechabeng sa prairie le meadow. Li-prairie tsa basali ba baholo ba hlalosang ka mokhoa o hlakileng litšoantšo tsa "oxytocin" tse bokellaneng tsa nyutlelie li bonts'itse ts'ebetso ea khetho ea molekane e potlakileng kamora ho tloaelana le monna, empa ha ea ka ea bonts'a boitšoaro bo matlafatsang ba ho ratana. Leha ho le joalo, khetho ea molekane ha ea ka ea tataisetsoa libakeng tse se nang monogamous meadow ka ho hlahisa li-oxytocin receptor ka har'a li-nucleus accumbens. Lintlha tsena li tiisa karolo ea karolo ea li-oxytocin receptor ho li-accumbens taolong ea litakatso tsa balekane ho li-voes tsa basali tsa basali, mme li fana ka tlhahiso ea hore polelo ea li-oxytocin receptor ka har'a li-accumbens ha e ea lekana ho nts'etsapele khetho ea molekane ho mefuta e seng monogamous. Lintlha tsena ke tsa pele ho bonts'a kamano e tobileng lipakeng tsa oxytocin receptor density ho li-nucleus bokellanang le phapang ea boitšoaro bo ikamahanyang le sechaba. Kahoo, phapang e fapaneng le e 'ngoe ea polelo ea oxytocin receptor ka har'a striatum e ka kenya letsoho phapang ea tlhaho ho boits'oaro ba sechaba.

Keywords: bo-mme, vaerase, likhetho, neuropeptide, cognition, autoradiography

SELELEKELA

Liroto tsa Microtine li bonts'a phapang e kholo boits'oarong ba sechaba, ho tloha mekhatlong e kopaneng le ea bophelo bo botle, ho isa ho maano a bohlano le a boitšoaro bo bobe.Gruderadams le Getz, 1985). Pholo e monate ea sechaba se le sengMicrotus ochrogaster) lifomatsi tse mamellang kamano ea motho e mong le e mong, kapa likamano tsa botona le botšehali ho ea khahlaneng le ho tlolelana, athe li-voles tse se nang bohloekiMicrotus pennsylvanicus) hangata se ke oa. Litaba tsa basali ba baholo ba basali ba thobalano li bonts'a phapang e kholo boitshwarong ba bona ba ho itšireletsa, kapa boits'oarong, ka halofo e bonts'ang boits'oaro bo joalo ba 'm'a ho bana ho ba bang ha ba setseng ba iphapanyetsa kapa ba hlasela malinyane (Lonstein le De Vries, 1999; Bales le Carter, 2003; Olazabal le Monyane, 2005). Kahoo li-voles li fana ka monyetla o motle oa ho hlahloba mekhoa e amanang le ts'ebetso ea bophelo le tlhokomelo ea setho, hammoho le mekhoa e lebisang karohanong ea boits'oaro bona lipakeng tsa mefuta le pakeng tsa batho ka bomong.

The nonapeptide oxetocin (OT) e bile le tšusumetso ho taolo ea moralo oa khetho ea balekane le boits'oaro ba alloparental libakeng tsa prairie. Ts'oaetso ea mohanyetsi oa li-oxytocin receptor (OTR) ka har'a nucleus accumbens (NAcc), empa eseng ka har'a lithibelo tse haufi tsa caudate putamen li-forming tsa lerato la molekane oa hae, protocol ea laboratori ea sebopeho sa maqhama a mabeli (Young le al., 2001). Litlatsetso tse tšoanang tsa mohanyetsi oa OTR ho NAcc le tsona li thibela boitšoaro bo sa tsotelleng ho basali ba batšehali (Olazabal le Monyane, 2006b). Ka hona ts'ebetso ea li-OTR ho NAcc e thusa ho theha khetho ea balekane le boits'oaro bo botle ba basali libakeng tsa basali tsa praemia.

Phapang lipakeng tsa tekano ea OTR ho NAcc e fupere maikutlo ho kenya letsoho phapusing ea mefuta ea mokhatlo le boitšoaro bo kopanetsoeng. Li-voes tsa Prairie li na le mathata a phahameng a OTR ho NAcc, athe li-voles tse se nang monogamous, litoeba le likhoto ha li (Insel le Shapiro, 1992; Olazabal le Monyane, 2006a). Li-voiri tsa Prairie li boetse li bonts'a maemo a phahameng a boitšoaro bo fetisang mahlo ho feta li-voles, litoeba kapa likhoto (meadow).Olazabal le Monyane, 2006a). E bapisa kamano ena ea lipakeng tsa mefuta-futa pakeng tsa letsoalo la OTR ho NAcc le boits'oaro ba batsoali, letsoalo la OTR ho NAcc le amana hantle le boits'oaro ba alloparental ho li-voes tsa basali ba baholo le ba baholo ba basali (Olazabal le Monyane, 2006b, a). Phuputsong ena, re sebelisitse mofuta oa adeno o amanang le viral vector (AAV) ho leka ka kotloloho maikutlo a hore phapang pakeng tsa NAcc OTR density e ka kenya letsoho phapusing ea khokahano ea sechaba le boits'oaro ba alloparental. Li-voes tsa basali ba baholo li ile tsa kenngoa ka bonngoe ho NAcc ka mofuta oa AAV o kopantseng mofuta oa mofuta oa VT OTR, oa etsa hore ho be le phallo e kholo ho OTR. Liphoofolo li ile tsa hlahlojoa molemong oa ho etsa liketso le khetho ea molekane. Ka mor'a moo re ile ra leka khopolo-taba ea hore polelo ea OTR ho NAcc e ne e lekane ho tsamaisa khetho ea balekane ka ho kenya li-voales tsa mehend e tšehali ka veter e le 'ngoe. Re ile ra bolela esale pele hore ha ho bapisoa le taolo, li-voes tsa basali tsa OTR tse hlalosang maikutlo ka mokhoa o hlakileng li tla bonts'a boitšoaro bo eketsehileng ba likamano le tšibollo ea khetho ea molekane e potlakileng. Ntle le moo, re boletse esale pele hore basali ba tlisoang ke basali ba hlalosang OTR ho NAcc ba tla etsa khetho ea balekane ho balekane ba banna.

LISEBELISOA LE MEKHOA

Animals

Li-voles tsa Prairie le meadow li ne li bolokiloe ka lihlopha tse ts'oanang tsa thobalano le li-voles / Xage ea 2-3 ho tloha nakong ea ho khoesoa nakong ea matsatsi a 21-23. Bolulo bo ne bo na le mochini o kenelletseng oa 36 × 18 × 19cm Plexiglass e tlatsitsoeng ka mealo ea Bed-ocobbs Laboratoring ea liphoofolo ka tlasa 14: 10 hr leseli / potoloho e lefifi ho 22 ° C le phihlello ea lijo (lera la LabDiet, Richmond, IN) le lipapatso tsa lipapatso tsa metsi. . Li-voes tsa prairie li fumanoe koloneng ea rona ea mosebetsi ea mahlale e neng e simolohile ho li-voale tse hapiloeng ke masimo Illinois. Li-voles tsa Meadow li nkiloe setulong se fumanoeng kolonng e fumanehang Univesithing ea Florida State. Lihlooho e ne e le li-voles tsa basali tsa 2-5 tse neng li le kahare ho thobalano le basali ba mahlale ba basali. Liphoofolo tsa stimulus e ne e le li-voles tsa banna ba baholo ba nang le boiphihlelo. Monna e mong le e mong e ne e le "molekane" le "osele" nakong ea tlhahlobo ea khetho ea molekane (bona ka tlase). Lingoloa li ile tsa abeloa lihlopha tse fapaneng tsa kalafo ho laola ho fapana ka matlung le ka har'a koekoe. Ts'ebetso tsohle li amohetsoe ke Komiti ea Tlhokomelo ea Liphoofolo le Emory University.

Bahlahisi ba likokoana-hloko ba Adeno-Associated

Tatellano ea ho ngola ea OTR e thehiloe ka ho qopitsa tlhaiso ea pele ea lebitso la tlholeho la VT OTR genomic le kenyelletsang libaka tsa pele tse hlano tsa transmembrane (nomoro ea kenyelletso ea Genbank AF079980) le pheletso ea 3 ′ ea OTR e holisitsoe ho prairie vole uterus cDNA. Ts'ebetso ea ho boloka kh'outu e hlophisitsoe bocha ka ho sebelisa PCR ho felisa li-UTR le ho fana ka libaka tse ncha tsa thibelo bakeng sa ho tsamaisa cloning ho AAV vector plasmid. Mofuta o fetotsoeng o ne o kenngoa moqomong oa veine oa AAV2 pakeng tsa morekisi oa 0.6 kb cytomegalovirus (CMV) le sekhechana sa SV40 sa DNA se nang le lets'oao la SV40 t tron ​​le polyA. AAV2-OTR e ne e tšehelitsoe ka bongata ho AAV9 ke phetisetso e tsoang habeli ho lisele tsa AAV-293 (Stratagene, La Jolla, CA) e sebelisa mokhoa o tloaelehileng oa calcium phosphate. Ha ho na kokoana-hloko e thusang. Plasmid ea AAV2-eGFP e ile ea kenngoa ka mokhoa o ts'oanang ho AAV9 e le vector e mpe ea taolo. Ka bokhutšoanyane, vector e 'ngoe le e' ngoe ea AAV plasmid, AAV2 / 9 rep / cap plasmid e fanang ka replicase ea AAV2 le mesebetsi ea capid ea AAV9, le 3rd plasmid encoding Adenovirus assist assist, pHelper (Stratagene), e ile ea fetisetsoa ka har'a lisele tsa 293 ka tekanyo ea molar ea 1: 1: 1. Lisele li ile tsa kotuloa kamora phetiso ea 48 hrs. Li-pellets tsa sele li ile tsa emisoa hape ho DMEM, le likaroloana tsa vaerase tsa intracellular tse lokollotsoeng ka makhetlo a mararo a latellanang a freeze-thaw, tsa lateloa ke centrifugation ho 13,000 rpm bakeng sa 10 min ho centrifuge ea tafoleng. Li-stock tsa vector li ne li bolokiloe ho −80 C, 'me li romelloa ke PCR ea nako ea nnete e sebelisang ABI Prism 7700 Sequence Detection System e tsoang Perkin-Elmer Applied Biosystems (Foster City, CA). Literata li ne li le ka tatellano ea 1012 DRP / ml. (DRP = Dnase Resistant Particles).

Ts'oaetso ea vaerase ea vaerase

Stereotaxic infusions e entsoe ka tlasa anoflurane anesthesia ho Kopf stereotax e kentsoeng ka Ultra Micro Pump II (Lisebelisoa tsa Lefatše tsa Precision, Sarasota, FL) le serope sa 26-gauge Hamilton. Basali ba ile ba kenngoa ka kotloloho mokokotlong oa NAcc (li-prairie voles: AP + 1.7mm, ML .9mm, DV −4.5mm, meadow voles: AP + 1.6mm, ML .9mm, DV −4.3mm) e nang le 750nl ea vAole. oxytocin receptor (CMV-OTR, N = 12), kapa control eGFP e hlahisang vector (CMV-GFP, N = 16). Virusi e ile ea kenngoa ka tekanyo ea 93.8nl / min. Syringe e ne e siiloe sebakeng sa 5 min kamora ho ts'oaroa ho nyenyefatsa ts'oaro ea vector ho fihlela pina ea nale. Liphoofolo tse tsamaeang le Sham (N = 9) li ne li sa sebetsoa 'me li ne li e-na le letlalo la tsona le hloekile. Kamora ho etsa opereishene, liphoofolo li ne li lula malapeng ho fihlela nako ea tlhahlobo ea khetho ea balekane. Lithuto tsa pele li bonts'a hore polelo ea OTR setsing sa ente e ne e tsitsitse kamora matsatsi a 10.

Teko ea Tsoalo ea Alloparental

Khoeli e le 'ngoe kamora ho entoa, li-prairie voles li ile tsa lekoa bakeng sa boits'oaro ba batsoali. Teko e etsahetse lipakeng tsa 0800hr le 1800hr. Liphoofolo tsa liteko li ile tsa beoa ka mokoting o moholo o hloekileng (45.5 × 24 × 20cm) mme tsa lumelloa ho tloaelana le 15min. Malinyane a mabeli (matsatsi a 2-5 a lilemo) a behiloe ntlheng e le 'ngoe ea cage. Ho ile ha hatisoa leqhubu la ho atamela malinyane, palo ea liphoofolo tse hlaselang malinyane, le nako e sebelisitsoeng ho itlhopha, ho tsoka le ho khutlisa malinyane. Ho lateloa metsotsoana e 900 ho liphoofolo tse sa kang tsa atamela malinyane nakong ea tlhahlobo ea metsotso ea 15 molemong oa tlhahlobo ea lipalo. Liteko li ile tsa emisoa hanghang haeba e tšehali e hlasela malinyane. Liphoofolo li ne li arotsoe ka bongata haeba li ka qeta metsotso e 30 li nyeka malinyane ntle le ho li hlasela. Ho ipapisitsoe le liphetho tsa liteko tsa li-prairie voles, meadow voles ha ea ka ea lekoa bakeng sa boits'oaro ba batsoali.

Teko ea Tlhahlobo ea Li-Partner (Prairie Vole)

Khoeling e le 'ngoe e latelang tlhahlobo ea boitšoaro ea alloparental,' me likhoeli tse peli kamora ts'ebetso ea ts'oaetso ea AAV, basali ba ile ba fuoa 4 μg ea estradiol benzoate (EB; Fisher, Pittsburgh, PA) e qhibilihile ho 0.1ml ea sesame oli IP letsatsi le leng le le leng bakeng sa matsatsi a 3 pele ho ho nyaloa ho hohela boits'oaro ba thobalano. . Lihora tsa 16 ka mor'a hore liphoofolo tsa ho qetela tsa ente ea EB li kentsoe ka lekoteng le hloekileng (28 × 17 × 12cm) le monna e moholo ea nang le thobalano ea nang le thobalano bakeng sa lihora tsa 6 ebe o khutlela ntlong ea sehlopha. Boitšoaro ba mating bo tlalehiloe nakong ea ts'ebelisano ea hora ea 6 ea hora. Moralo oa morao-rao oa ho kenella kahare le palo ea linako tsa ho hola ka hora ea pele li pakiloe. Latency ea metsotsoana ea 3600 e fuoe liphoofolo tse sa kopaneng nakong ea hora ea 1 molemong oa tlhahlobo ea lipalo. Hoseng ho latelang (14 hrs kamora ho ts'oaroa), liphoofolo li ile tsa lekoa bakeng sa khetho ea molekane. Tlhahlobong ea moratuoa oa mosali, mosali ea hlahlobang o kenngoa ka setsing se se nang lehlakore sa lisebelisoa tse kamoreng e meraro, moo monna e mong a kenelletseng kamoreng e le 'ngoe' me monna e mong ea sa tsebeng letho a kenngoe kahare ho e 'ngoe.Williams et al., 1992). Phoofolo ea teko e na le bolokolohi ba ho tsamaea ka likamoreng tsohle 'me nako e sebelisitsoeng haufi le monna e mong le e mong e tla rekisoa ho sebelisoa sistimi ea boikhathollo ea beamo (Curtis le Wang, 2005b, a; Lim et al., 2007).

Letsatsi hamorao, tse tšehali li ile tsa kopanngoa hape le molekane a le mong bakeng sa lihora tse ling tsa 12 tse eketsehileng tsa ho phomola. Basali ba ile ba lekoa hape bakeng sa khetho ea molekane (18 hora ea nako eohle ea ho ikopanya). Morero ona oa mekhahlelo e 'meli o sebelisitsoe ho eketsa tlhahlobo ea rona ea tlhahlobo ea tlhahlobo ea khetho ea balekane, hobane ho na le phapang nakong ea nako e hlokahalang ho theha khetho ea molekane. Liphoofolo li ne li nkuoa li na le boikhethelo ba molekane haeba li ka qeta nako e ngata li atamelane haufi le molekane ha li bapisoa le moeti.

Teko ea Tšehetso ea Partner (Meadow vole)

Ka tloaelo mehato e tsoang mefuteng ea rona ea kolone e ke ke ea theha setlamo kamora ho pepesetsoa maqhubu a molekane oa 24hrs. Ka hona bakeng sa teko ena, liteko tsa khetho ea molekane li ile tsa etsoa kamora ho qobelloa ha 24 le palo e eketsehileng ea 48 hrs (kakaretso ea 72 hrs). Mekhoa eohle e meng ea kalafo le liteko e tšoana le e kaholimo.

Ho bokella lisele le ho sebetsa

Kamora boithuto ba boits'oaro ba liphoofolo bo ile ba khethoa kamora ho anesthetization e tebileng le isofluorane. Ka mor'a moo, ho ile ha bokelloa likoto tsa boea 'me tsa hoamisoa leqhoeng le omileng le nang le leqhoa. Mehopolo e ne e arotsoe ka NAcc ka 6 letotong la 20μm, ka kristallara, ho li-slide tsa Fisher Frost-plus. Li-Slide li ne li bolokiloe ho −80C ho fihlela li sebelisoa ho autoradiography.

Autoradiography ea OTR

OTR receptor autoradiography e sebelisitsoe ho lekola ho tlama ha OTR ho liphoofolo tse lemetseng tsa AAV. Autoradiography e entsoe joalo ka ha ho hlalositsoe pejana ka liphetoho tse nyane (Insel et al., 1991; Wang le Bacha, 1997). Likarolo li tlositsoe sebakeng sa polokelo ea motlakase sa 80C, se lumelletsoe moeeng, se kenngoe ka 0.1% paraformaldehyde ho phosphate-buffered saline (pH 7.4), 'me sa ts'oaroa habeli ho 50 mM Tris buffer (pH 7.4) ho tlosa endo native OT. Ka mor'a moo lithane li ile tsa kenngoa ho 50 pM 125I-OVTA (NEX 254050UC PerkinElmer, Waltham, MA) bakeng sa hora e le 'ngoe. Radioligand e sa fetoheng e tlositsoe ke ho hlatsuoa tse 'ne ho 50mM Tris le 2% MgCl2 (pH 7.4) ebe e kenngoa ka har'a dH20 le moea li omelletsoe ke moea o pholileng. Hang ha e omme, li-slide li ile tsa pepesetsoa filimi ea BioMax MR (Kodak, Rochester, NY) bakeng sa lihora tsa 72. Liphoofolo tse peli tsa CMV-OTR le e 'ngoe ea CMV-GFP ha lia kenyelletsoa tlhahlobisong ea boitšoaro ka lebaka la liphoso tsa ente.

GFP Immunohistochemistry

Setsi sa liphoofolo tse tlase (N = 5) se kentsoeng CMV-GFP, se ile sa tšeloa oli ka bongata ka 50 ml ea PBS, sa lateloa ke 50 ml ea 4% paraformaldehyde ho 0.1 M phosphate buffer e nang le 2.5% acrolein (Polysciences, Warrington, PA). Hang ka mor'a ho etsoa ka litlolo, methapo e ile ea tlosoa 'me ea bolokoa ho 4 ° C ka tharollo ea 30% ho fihlela e aroloa. Boreli bo ne bo arotsoe likarolo tsa coron tsa 25μm ka microtome e sa tsitsang 'me e bolokiloe e sa phalle ka tharollo ea cryoprotectant ka −20 ° C ho fihlela ts'ebetso ea immunohistochemical.

1: Letoto la 6 ka har'a rostrocaudal axis ea boko bo bong le bo bong le ile la sebetsoa bakeng sa GFP. Ka bokhutšoanyane, likarolo li tlositsoe tharollong ea "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" "" ("mas" (s) penshene). Likarolo li ile tsa kenngoa tharollong ea antibody ea mantlha e lebisitsoeng khahlanong le GFP ho saline ea potasiamo phosphate-lubffered (KPBS) e nang le 7.4% Triton-X bakeng sa hora ea 15 mochesong oa kamore o lateloang ke lihora tsa 1 ho 0.1 ° C. Lisele tse nang le GFP li ile tsa bonts'oa ka ho sebelisa 'mele oa anti-GFP anticlone (Cat. No. A1, Attitrogen, Carlsbad, CA) moo ho nang le 48: 4. Kamora ho kenella kahare ho antibody, lithane li ile tsa phatloloa KPBS, e fuoe hora ea 6455 ho poli ea biotinylated ea poli ea IgG (Jackson Immunoresearch, West Grove, PA) kopanong ea 1: 100,000, le rinsed ho KPBS, ea lateloa ke 1- incubation ea hora ho avidin-biotin peroxidase tata (ABC Elite Kit PK-1 Vector, Burlingame, CA) kopanong ea 600: 1. Kamora ho hlapoloha ho KPBS le Tris buffer (pH 6100), GFP e ile ea bonoa e le sehlahisoa sa karabelo e sootho ka ho sebelisa 1′-diaminobenzidine e nang le 200% hydrogen peroxide ho Tris buffer. Sehlahisoa sa karabelo se ile sa emisoa kamora metsotso e ka bang 7.2 ka ho hlaba ka Tris buffer. Likarolo li ne li kentsoe ka letsoai ho li-slide tse kenngoeng ka li-gelatin, tse omisitsoeng ke moea, tse se nang metsi ka bongata letotong la li-alcohols tse halikiloeng, li hloekisitsoe ho Histoclear (National Diagnostics, Atlanta, GA), 'me e koaetsoe ka Krystalon (EMD Chemicals, Gibbstown, NJ).

Tlhahlobo ea Litekanyetso

Lintlha li hlahisoa joalo ka phoso ± phoso e tloaelehileng ea se boleloang (SEM). Litsela tse peli tsa RM ANOVA li ne li sebelisoa ka nako e sebelisitsoeng le phoofolo e 'ngoe le e' ngoe e susumetsang joalo ka phapano e itšetlehileng ka eona, se kahare ho lihlooho e le molekane kapa motho osele, le sehlopha sa kalafo e le sona se pakeng tsa taba. Teko ea Holm-Sidak e ne e sebelisetsoa papiso ea post hoc ka bobeli ha ho ne ho fumanoa tšusumetso e kholo ea tšebelisano. Tsela e 'ngoe ea li-ANOVA e ne e sebelisoa ka boits'oaro ba batho bohle le ba ho nyalanya li-voles tsa thota. Boitšoaro bo neng bo sa fihlelle litekanyetso tsa maemo a tloaelehileng bo ile ba hlahlojoa ho sebelisoa Kruskal-Wallis One Way Analysis of Variance on Ranks. Teko ea 'nete ea Fisher e sebelisitsoe ho tseba phapang ea sehlopha palong ea liphoofolo tse bonts'ang boits'oaro ba bongoana. Boitšoaro ba Meadow vole mating bo ile ba hlahlojoa ho sebelisoa t-Test le Mann-Whitney Rank Sum Test ha tlhahlobo e tloaelehileng e hloleha.

LIPOTSO

Boitšoaro ba Alloparental in Prairie Voles

Karolo ea li-voes tsa basali tsa li-prairie tsa CMV-OTR tse entšitsoeng tse bonts'ang boits'oaro ba batho bohle (3/9) li ne li sa fapana le taolo (4/15) kapa sham voles (3/10) (p> 0.5; Setšoantšo sa 1A). Li-CMV-OTR tse peli, taolo e 'ne ea CMV-GFP, le tse tšehali tse peli tsa sham, tse neng li khethollotsoe ka hore ha li sebetse, li ile tsa hlasela malinyane. Latency ho atamela malinyane (Setšoantšo sa 1B) le nako e sebelisitsoeng ho nyeka / ho itlhopha le eona ha ea ka ea fapana haholo lipakeng tsa lihlopha (H = 0.31, P> 0.8 bakeng sa latency, H = 0.40, P> 0.8 bakeng sa boitlhopho). Ha ho ne ho bapisoa feela liphoofolo tse fihletseng litekanyetso tsa ho ba batho bohle, latency ea ho atamela malinyane bakeng sa basali ba CMV-OTR (metsotsoana e 140.2 ± 65.2) e ne e sa fapana le CMV-GFP (metsotsoana e 62.5 ± 39.3) kapa sham (metsotsoana e 169.4 ± 51.7) basali (F (2,7) = 1.27, p> 0.3). Ho ne ho boetse ho se na phapang palong ea nako eo basali ba CMV-OTR ba sebelisang bana ba nyekang ha ba bapisoa le ba CMV-GFP kapa basali ba sham (F (2,7) = 1.94, p> 0.2; Setšoantšo sa 1C). Nako eohle eo sehlopha ka seng se qetileng ho e nyeka / ho itlhopha, ho tsamaisa le ho tsamaisa malinyane le eona e ne e sa fapane lipakeng tsa alloparental CMV-OTR (metsotsoana e 695.2 ± 77.1) CMV-GFP (387.5 ± 132.7 metsotsoana) kapa sham (metsotsoana e 503.8 ± 82.8 liphoofolo (F (2,7) = 1.98, p> 0.2).

Setšoantšo sa 1  

Boitšoaro ba alloparental sham, CMV-GFP le CMV-OTR basali prairie voles. A) Ha ho na phello ea kalafo ka tekanyo ea basali sehlopheng se seng le se seng sa kalafo se bonts'ang boitšoaro bo kopanetsoeng. Mekoallo e lefifi e emela liperesente tsa liphoofolo tseo ...

Ho Ts'oarella Litekong tsa Li-Prairie Voles

Boitšoaro ba thobalano bo ne bo sa angoa haholo ke kalafo ea basali ea kalafo. Ho lieha ho kenella pele ho ne ho sa fapana haholo ho banna ba kopantsoeng le basali ba CMV-OTR ha ba bapisoa le ba batona ba nyalaneng le CMV-GFP kapa basali ba sham (H = 5.043, p = .08; Setšoantšo sa 2A). Le ha sehlopha sa CMV-OTR se ne se tloaetse ho nyalana kapele ho feta lihlopha tse ling, se ne se sa kopane hangata. Palo ea likhahla tsa ho nyalana ka hora ea pele ha ea ka ea fapana ka lipara tse nang le basali ba CMV-OTR ho tloha lipara tse nang le CMV-GFP kapa basali ba sham (F (2,31) = 0.46, p> 0.6; Setšoantšo sa 2B).

Setšoantšo sa 2  

Litloaelo tsa batho ba ratang ho ratana le balekane ho sham, CMV-GFP, le CMV-OTR. Boemo ba morao-rao ba ho kenella pele (A) le palo ea likhahla tsa mating (B) li ne li sa fapana haholo pakeng tsa lihlopha. C) Kamora nako ea cohabitation ea hora ea 6, ...

Boitšoaro ba Boikhethelo ba Molekane ho Prairie Voles

Kamora nako ea boiketlo ba 6 hr, ha ho le se seng sa lihlopha se bonts'itseng khetho ea bohlokoa ea molekane (Setšoantšo sa 2C). Ho ne ho se na phello ea mantlha ea kalafo (F (2,31) = 0.56, p> 0.5) kapa nako e sebelisitsoeng le molekane ho fapana le motho osele (F (1,31) = 0.46, p> 0.5). Kamora ho qeta lihora tse 12 tsa ho lula hammoho, ho ne ho se na tšusumetso e kholo ea kalafo (F (2,29) = .78, p = 0.5) kapa nako e sebelisitsoeng le molekane le motho eo a sa mo tsebeng (F (1,29) = 3.71 , leq = 0.06). Leha ho le joalo, ho bile le tšusumetso e kholo ea tšebelisano (F (2,29) = 5.56, p = 0.009). Teko ea post-hoc e senotse hore basali ba CMV-OTR ba sebelisitse nako e ngata haholo ba le haufi le molekane ho feta mojaki (p <0.001; Setšoantšo sa 2D). Leha ho le joalo, basali ba CMV-GFP le ba sham ba hlotsoe ho bonts'a khetho ea balekane kamora nako ena (p> 0.05; Setšoantšo sa 2D). Ntle le moo, 80% ea li-voles tsa CMV-OTR tse kentsoeng e fihletse litekanyetso tsa ho ratoa ke molekane, ke hore, ho qeta nako e ngata le molekane ho feta moeti, ha ho bapisoa le 31% feela ea CMV-GFP e kenelletseng le 44% ea liphoofolo tse sham (Setšoantšo sa 2E, F).

Pontšo ea OTR le GFP ho NAcc ea Prairie Voles

Autoradiography e entsoe ho tseba ho beoa ha ente ea AAV le ho netefatsa hore vector ea CMV-OTR e hlahisitse tlamo ea OTR e ngata ha e bapisoa le taolo. Joalokaha ho tlalehiloe pejana, ho bile le phapano ea bohlokoa ea motho ka mong ho OTR e tlamang NAcc ea taolo ea CMV-GFP prairie voles (Setšoantšo sa 3A, B). Ho phahamisitsoe ka mokhoa o pharalletseng litlamong tsa OTR ho fumanoe ho NAcc ea CMV-OTR injected prairie voles (Setšoantšo sa 3C). Ntle le moo, immunohistochemistry ea GFP e ile ea etsoa likarolong tsa boko ba liphoofolo tsa CMV-GFP ho fumana tekanyo ea polelo ho li-neurons tsa NAcc. Ho ngotsoe ka mokhoa o hlakileng oa lisele tse nang le litšobotsi tsa neuronal ho fumanoe, ho tiisa hore li-veter tsa CMV AAV li tsamaisitse polelo ho li-neurons (Setšoantšo sa 3E, F).

Setšoantšo sa 3  

Tlhahlobo ea polelo ea OTR le GFP ho li-CMV-OTR le CMV-GFP basali prairie voles. Sekhahla sa OTR se ne se ikemiselitsoe ho sebelisa li-receptor autoradiography. A, B) Ho bile le phapang e kholo boteng ba OTR e tlamehang ho NAcc ea basali ea CMV-GFP. Sena se fapane ...

Ho Ts'oarella Litekong Me Meheleng e Mebele

Ha ho na phapang e kholo ea boits'oaro ba botona le botšehali e bonoeng lipakeng tsa lihlopha tsa kalafo ea mead voles. Ts'ebetso ea ho kenella ka lekhetlo la pele e ne e sa fapana ho banna ba kopantsoeng le CMV-OTR (1245.1 ± 1465.9 metsotsoana) basali ha ba bapisoa le ba batona ho laola CMV-GFP (metsotsoana e 566.6 ± 989.8) ea basali (T = 128.0, p> 0.2). Palo ea likhahla tsa ho nyalana ka hora ea pele le eona ha ea ka ea fapana ka lipara tse nang le basali ba CMV-OTR (6.8 ± 5.1) ho tloha lipara tse nang le basali ba CMV-GFP (9.5 ± 4.4) (t = 1.42, p> 0.1).

Khetho ea Molekane ho Meadow Voles

Kamora nako ea boiketlo ba 24hr, ha ho sehlopha se bonts'itseng khetho ea bohlokoa ea molekane (Setšoantšo sa 4A). Ho ne ho se na phello ea mantlha ea kalafo (F (1,20) = 0.19, p> 0.6) kapa nako e sebelisitsoeng le molekane ho fapana le motho eo a sa mo tsebeng (F (1,20) = 0.09, p> 0.7). Kamora lihora tse ling tsa 48 tsa ho lula hammoho, ho ile ha hlola ho e-ba le tšusumetso e kholo ea kalafo (F (1,19) =. 65, p> 0.4) kapa nako e sebelisitsoeng le basali (F (1,19) = 0.62, p> .4; Setšoantšo sa 4B).

Setšoantšo sa 4  

Boitšoaro bo ikhethileng ba li-voales tsa basali tsa kamore e tšehelitsoeng ka mor'a ho kopana ha 24 hr (A) le kamora ts'ebelisano ea hora ea 72 hora e kopanetsoeng (B). CMV-OTR le CMV-GFP tse tšehali basali ha lia ka tsa bontša khetho ea molekane. Ba qetile nako e lekanang le molekane eo ...

Tlhaloso ea OTR ho NAcc ea Meadow Voles

CMV-GFP e kentseng li-vooses tsa mehend li ne li e-na le OTR e fokolang kapa e ne e sa tlame letho ho NAcc (Figure5A). Ka lehlakoreng le leng, li-voles tsa CMV-OTR li kentse li-voles tsa mehend li ne li tlama OTR ea bohlokoa ho NAcc, e bapisoang le li-voes tsa prairie (Setšoantšo sa 5B).

Setšoantšo sa 5  

Receptor autoradiography e bonts'a letsoalo le tlamang la OTR ho NAcc ea CMV-OTR (A) le CMV-GFP (B) li-meadow voles. Hlokomela hore li-voales tsa mehendi tsa basali tsa CMV-GFP li ne li e-na le OTR e fokolang kapa e ne e sa tlame letho ho NAcc. Leha ho le joalo, OTR e tlamang ho NAcc e ile ea phahamisoa haholo ...

TŠOHLOA

Mefuta ea phapano ea bongata ba OTR ho NAcc e amahantsoe le mefuta e fapaneng ea leano la ho emisa (morogamy vs polygamy) le boits'oaro bo bonts'ang maikutlo, a fana ka maikutlo a hore phapang ea polelo ea li-receptor tsa OTR e ka baka phapang ea mefuta ea mokhatlo le boits'oaro (Insel et al., 1991; Bacha le Wang, 2004; Olazabal le Monyane, 2006b). Li-voes tsa monogamous prairie tsa sechaba li na le mathata a phahameng a OTR ho NAcc ho feta li-meadow le li-montane voles, le pharmacological blockade ea li-receptor tse thibelang ho ratoa ha balekane (Young le al., 2001; Olazabal le Monyane, 2006b). Ho na le phapang e fapaneng e ikemetseng le bongateng ba OTR ho NAcc kahare ho libaka tse rorisang (Setšoantšo sa 3A, B). Sekhahla sa OTR tikolohong ena se tsamaisana hantle le phapang e fapaneng ea boits'oaro ba batho ba baholo ba nang le thobalano ea basali ba nang le thobalano (E monyenyane, 1999; Olazabal 2006) le tsamaiso ea mohanyetsi oa OTR ho NAcc e felisa boitšoaro bona (Olazabal le Monyane, 2006a). Ka hona, thutong ena re batlile ho leka ka kotloloho kamano pakeng tsa letsoalo la receptor ho NAcc le boitšoaro bo amanang le li-voles, ka ho fetisetsa mofuta oa AAV.

Liphetho tsa rona li bonts'a hore, joalo ka ha ho boletsoe esale pele, li-voes tsa basali tsa basali tse nang le maemo a phahameng a OTR ponong ea NAcc e potlakileng ea khetho ea molekane ha e bapisoa le basali ba nang le letsoalo le tlase la OTR (CMV-GFP kapa shams). Cka lebaka la boitlamo ba rona, ho ne ho sena phapang boits'oarong bo kopanetsoeng pakeng tsa lihlopha, ho fana ka maikutlo a hore na OTR e ikemetseng e laola mofuta oa khetho ea molekane joang le boitšoaro ba ho ratana. Liphetho tsa rona li tšehetsa khopolo-taba ea hore liphapang tsa motho ka mong ho polelo ea OTR li kenya letsoho phanong ea mefuta e meng ea likarolo tse ling tsa boitšoaro bo kopanetsoeng. Leha ho le joalo, polelo e ntseng e eketseha ea OTR ho NAcc e ne e sa lekana ho susumetsa khetho ea balekane ho li-mehend tsa mehend, le kamora ho qeta lihora tsa 72, ho fana ka maikutlo a hore mefuta ea mefuta ea OTR feela ha e ea lekana ho hlalosa mefuta ea mefuta ka bokhoni ba ho theha molekane. likhetho. Ho bohlokoa ho hlokomela hore ho ne ho se na phapang pakeng tsa boits'oaro ba mating ho efe kapa efe ea lipalo tse hlahlojoang. Ka hona, khetho e ntlafalitsoeng ea moratuoa libakeng tsa liteko tsa teko e ke ke ea hlahisoa ka ho eketseha ha thobalano nakong ea thobalano ea pele.

Li-voalee tsa basali ba baholo li bontša phapang e ikhethang ea motho ka mong pontšong ea bona ea tlhokomelo e atamelaneng. Hoo e ka bang 50% ea basali ba nang le botona ba botona le botšehali e tla fumana, e lick / monyali le ho hula malinyane ao ba a tliselitsoeng (Lonstein le De Vries, 1999; Bales le Carter, 2003; Olazabal le Monyane, 2005). Ho na le phapang e fapaneng e ikemetseng le bongateng ba OTR ho NAcc har'a li-voes tsa prairie (E monyenyane, 1999). Basali ba nang le maemo a phahameng a OTR ho NAcc ba khona ho bonts'a karabelo e matla ho feta liphoofolo tse nang le OTR e tlase sebakeng sena. Ka hona ho ile ha makatsa hore ho phahamisa letsoalo la OTR ho NAcc ea li-voes tsa prairie ha hoa ka ha ntlafatsa boitšoaro bo bonts'ang maikutlo. Taba ea hore liphoofolo tsena li bonts'itse ts'ebetso ea khetho ea molekane e potlakileng e fana ka maikutlo a hore OTR e tsoang ho viral vector transgene e ne e kopantsoe ka nepo ho supa lits'ebetso tsa phetisetso ho NAcc.

Re kholoa hore phapang ea ts'ebetso ea OTR ho NAcc nakong ea ntlafatso e ka bapala karolo ea bohlokoa ho hlahiseng phapang ea mefuta ea tlhaho le boits'oaro bo kopanetsoeng. Kamano e ntle mathateng a OTR ho NAcc le boits'oaro bo botle li bontšitsoe libakeng tsa bacha tsa prairie le ho batho ba baholo (Olazabal le Monyane, 2006b, a). Boitšoaro ba alloparental libakeng tsa prairie bo bonahala bo le bohlokoa haholo ho lits'oaetso nakong ea kholo. Mohlala, ho pepeseha hoa ts'ebetso ea "OT" e fetolang boitšoaro ba motho ka linako tsohle ho li-voes tsa basali tsa basali (Bales et al., 2007). Ka hona, batho ba nang le mathata a phahameng a OTR nakong ea nts'etsopele ba ka ba le mats'oao a ho eketseha a OTR ho NAcc, ea baka phetoho ea nako e telele ea methapo e eketsang monyetla oa ho bonts'a mekhoa ea alloparental ha ba eba batho ba baholo. Haeba khopolo-taba ena e nepahetse, re ka bolela esale pele hore polelo ea ho eketsa OTR ho NAcc ka neonatally e tla eketsa khafetsa ea boits'oaro.

Tlhaloso e 'ngoe bakeng sa ho sitoa ho ntlafatsa polelo ea OTR ho NAcc ho eketsa boitšoaro bo kopanetsoeng ke hore phapang ea polelo ea OTR libakeng tse ngata tsa boko e ka ba ea hlokahala ho hlahisa phapano e lebelletsoeng ea boits'oaro. Mohlala, polelo ea OTR ho septum ea morao e amanang hantle le boitšoaro bo amanang le alloparental (Olazabal le Monyane, 2006a). Kamora nako, ho ka etsahala hore phapang ea ho pepesetsoa hoa li-hormone kapa boiphihlelo ba sechaba e susumetse tsitsipano ea OTR ho NAcc ntle le ho fetola boits'oaro ba alloparental. Mohlala, ho ba teng ha ntate kapa ho ba sieo ha ntate nakong ea nts'etsopele kapa tšusumetso ea kelello ea lihormone tsa steroid ho ka susumetsa ponaletso ea karabelo e matla ho li-voes tsa basali tsa prairie (Roberts le al., 1996; Roberts le al., 1998; Lonstein le De Vries, 2000).

Li-voes tsa prairie tsa banna, vasopressin e bapala karolo ea bohlokoa ho bonts'a boitšoaro ba bo-ntate hammoho le sebopeho sa khetho ea molekane, e tsamaisana le karolo ea oxytocin ho basali (Winslow et al., 1993; Wang et al., 1994). Boithuto bo ikhethileng ba bongaka ba saense bo bontša hore V1aR ka har'a ventral pallidum, tlhahiso e kholo ea NAcc, e bohlokoa bakeng sa ho thehoa ha maqhama a mabeli (Lim le lenyane, 2004). Ho eketsa V1aR ka har'a pallidum ea ventral e sebelisang phetiso ea vaerase ea viral e potlakisang ts'ebetso ea lebelo ho potlakisa kaho ea khetho ea molekane.Pitkow et al., 2001), phumano e tšoanang le thuto ea hona joale. Le ha V1aR e bua ka mokhoa o fetelletseng oa li-voallary pallidum tsa li-mehend tsa banna e ne e sa lekana ho etsa boitšoaro ba ntate, e khothalelitse khetho ea molekane (Lim et al., 2004); ho fapana le hoo, thutong ea hona joale ea OTR ho NaCC ha ea ka ea hlohlelletsa khetho ea molekane maemong a basali a makhulo.

Ho na le litlhaloso tse 'maloa bakeng sa ho hloleha ha li-voales tsa mehends tsa basali tse nang le OTR e phahameng ho NAcc ho theha khetho ea molekane. Taba ea pele, mefuta ea mefuta ea ho lokolloa ha OT kahara NAcc nakong ea ho tsofala e kanna ea fapana lipakeng tsa basali ba libaka tsa basali le maqhubu a mehendi. Le ha tikoloho ea OTR e fapana lipakeng tsa mefuta, li-voes tsa prairie le li-voint tsa meadow li na le likhoele tse sebetsang tsa OT ho NAcc (Ross and Young, data e sa phatlalatsoang). Ka vivo liteko tsa tlhahlobo ea microsalal li bonts'itse hore ho hanana ho matlafatsa tokollo ea OT ho NAcc ea li-voes tsa basali tsa basali, empa lithuto ha li e-so ka li etsoa ho li-voalk voles (CD Cole and Young, data e sa phatlalatsoang). Ho ka etsahala hore ho kenella ha OT ho CMV-OTR liphoofolo tse tšoaroang ho ka nolofalletsa mookameli oa khetho ea molekane ho li-voales tsa mehendi. Leha ho le joalo, ho lokela ho hlokomeloe hore ho likhoto le linku, ho hlohlelletsa vaginocervical ho eketsa ho tsoa ha OT ea mantlha (Kendrick et al., 1986; Sansone et al., 2002). Ke ka hona ho ka etsahalang hore li-voles tsa mehend li se li ntse li le teng ka lebaka la ho phahama ha palo ea batho ba tlotsitsoeng ka marang-rang. Tlhaloso e 'ngoe bakeng sa ho hloleha ha polelo ea NAcc OTR ho susumetsa khetho ea molekane ho li-voales tsa mehend ea basali ke hore phapang ea mefuta ea polelo ea OTR libakeng tse ling tsa boko le eona e hlokahala bakeng sa ho kopanya. Mohlala, letsoalo la OTR le lona le phahame kahara cortex ea pele le li-amygdala tsa morao-rao libakeng tsa prairie ha li bapisoa le mefuta e seng ea ammonogamous vole (Insel et al., 1991; Young le al., 1996; Smeltzer et al., 2006). Ka hona, ho phahamisa polelo ea OTR libakeng tse ngata tsa marang-rang e kanna ea hlokahala bakeng sa ho susumetsa likhetho tsa balekane ba ikemiselitseng ho hola ka li-voales vooses. Kamora nako, ho a khonahala hore phapang e pakeng tsa methapo pakeng tsa mefuta e ikarabella bakeng sa phapang ea mefuta (mohlala, dopamine, corticotropin releading factor) (Gingrich et al., 2000; Liu le Wang, 2003; Smeltzer et al., 2006; Lim et al., 2007).

Ho na le likheo tse 'maloa tsa bohlokoa tsa mokhoa oa rona oa teko oa puisano o netefatsang. Taba ea mantlha, CMV-OTR e kanna ea eketsa polelo ea OTR mefuteng eohle ea sele e sebakeng se lemetseng, ho kenyelletsa le palo ea methapo ea kutlo eo hangata e sa arabeleng OT. Leha ho le joalo, ho ka etsahala hore transgenic OTR e hlahise ho hooa ho hoholo ha li-neuron tse atisang ho hlalosa OTR ea nako e telele ka lebaka la ho matlafatsa maikutlo ho OT e lokolotsoeng li-neuron tseo, kapa boteng ba limolek'hule tse loketseng tse theohelang. Taba ea bobeli, sebaka sa poleloana ea transgene e hlahisitsoeng ke vaerase ea virator ha sea ka sa hlahisoa ka mokhoa o ts'oanang ka boholo ba rostral-caudal kapa bohareng ba NAcc. Ho basali ba CMV-OTR, polelo ea OTR e ne e tsitsitse ka mokhoa o tšoanang maemong a mantlha a NAcc. Leha ho le joalo, ho lokela ho hlokomeloe hore khetla ea NAcc e bile le tšusumetso e matla ho taolo ea moralo oa khetho ea balekane le letsoalo la OTR sebakeng sena se lumellanang haholo le boitšoaro ba alloparental ho feta sebakeng sa mantlha.

Ena ke thuto ea pele ho bontša ka ho hlakileng hore ho feto-fetoha ha polelo ea OTR bokong ho ka kenya letsoho phapusing ea boitšoaro ba sechaba. OT e kentse letsoho khafetsa taolong ea boits'oaro bo fapaneng, ho kenyelletsa ts'ebetso ea tlhahisoleseling ea litaba tsa sechaba le mohopolo, khetho ea molekane, ho holisa le ho hokahanya basali (Dantzer et al., 1987; Kendrick, 2000; Ferguson et al., 2001; Kavaliers et al., 2003). Hona joale ho na le bopaki bo hlakileng ba hore OT e hlophisa kutloisiso ea motho ea boiketlo ba batho, ho kenyelletsa ho ts'epahalla motho ka mong, ho shebella ka mahlo, ho hopola lintho ka sefahleho le ho lemoha maikutlo (Kosfeld et al., 2005; Domes et al., 2007; Guastella et al., 2008; Savaskan et al., 2008). Tsamaiso ea OT e eketsa ho boloka kelello ea sechaba ka mosebetsi oa ho khothatsa ka lentsoe ho lithuto tsa autistic (Hollander et al., 2007), le lithuto tse 'maloa tsa liphatsa tsa lefutso li tlalehile litloaelano tse itekanetseng lipakeng tsa li-polymorphisms tse sa ngoliseng mofuta oa OTR gene le autism spectrum disorder (Wu et al., 2005; Jacob et al., 2007; Lerer et al., 2007; Yrigollen et al., 2008). Ho na le tlhaiso-leseling e pharalletseng mabapi le phepelo ea OTR, 'me ha ho letho le tsejoang mabapi le phapang ea motho ka mong khong ea OTR bokong ba motho. Liphetho tsa rona li fana ka maikutlo a hore phapang ea bongata ba OTR libakeng tse ikhethang tsa boko e ka kenya letsoho phapang e fapaneng ea tšebetso ea kelello ea motho ho batho.

Liteboho

Bangoli ba batla ho leboha Lorra Mathews ka mosebetsi oa hae o motle oa ho tsamaisa kolone ea rona. Phuputso ena e tšehelitsoe ke lithuso tsa NIH MH064692 ho LJY, RR00165 ho Jerkes National Primates Research Center, le NSF STC IBN-9876754.

LITLHAHISO

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