Methylene blue mukutsvakurudza zebrafish: chinowedzerwa pakuchengetwa uye chinokanganisa kuedza

Mvura yemazai ezebrafish inowanzova nebhuruu risina kunyanya kusimba, uye vanhu vashoma vanonyora kuti nei zvakadaro. Dhayi yacho imethylene blue, inowedzerwa kudzivirira chakuvhe, uye nzira zhinji dzebasa dzinoiona sechinhu chiripo chete chisina basa pakuedza. Kufunga kwakadaro hakuna kururama pano. Zviri nani kuiona sechinhu chinokanganisa kuedza chisina kunyorwa: chinhu chiripo mumidziyo ine zviri kuedzwa asi chisipo mumapoka ekuenzanisa, chine mashandiro emushonga pahuwandu hunoshandiswa, uye chinokwanisa kuchinja zviratidzo zviri kuyerwa nekuedza kwacho.

Musiyano iwoyo ndiwo unoronga zvose zviri pasi apa. Kushandiswa nguva dzose kwechinodzivirira chakuvhe pakuchengetwa inyaya imwe. Kuti chinowedzerwa ichocho hachichinji here bvunzo yako inyaya yakasiyana. Zvinyorwa zvitsva zvekutsvakurudza zebrafish zvinopa mhinduro inonyaradza panyaya yokutanga, asi inonetsa pane yechipiri.

Zvinonyatsorayirwa nemirayiridzo

Hapana mwero mumwe chete unotevedzwa nevose, uye ichocho chikamu chedambudziko. Mirayiridzo yeCanadian Council on Animal Care inobvumira 0.5 mg/L, kana 0.00005%, mumushonga wemvura unoshandiswa kutakura mazai netuvana twehove kuderedza kukura kwechakuvhe. Mirayiridzo yeEurope yemhuka dzemurabhoritari inotsanangura mazai ari kukura achichengetwa mumvura inowanzova ne0.00005% imwe chete iyoyo. Nzira yeCold Spring Harbor yekugadzira E3 inorayira huwandu hwakapetwa kaviri, 1 mg/L kana 0.0001%, sechimwe chezvinoumba mvura yacho. Asi pane chimwe chinosiyiwa pachena: mirayiridzo yeOECD yebvunzo yekukuvadza kunokonzerwa nemakemikari kumazai ehove ari kukura inorayira mvura yekuderedza kusimba kwemakemikari yakagadzirwazve uye kemikari iri kuedzwa, pasina kurudziro yemethylene blue mushanduro yayo ya2013 kana mushanduro yakazogadziridzwa. Saka imwe rabhoritari inokurisa mazai ezebrafish ari kukura mumvura yebhuruu ine methylene blue kubva pamaawa okutanga oupenyu, imwe ichiita bvunzo yemazai ari kukura inodiwa nemitemo mumvura isina ruvara, uye dzose dzichiti mvura yadzo ndiyo yakajairika. Ani naani anoenzanisa zvabuda mumarabhoritari akasiyana, kana kubatanidza zvakabuda mumapoka ekuenzanisa ezvidzidzo zvekare, anotora kusawirirana ikoko.

Kuti huwandu uhu hunzwisisike, 0.00005% ingangoita 1.56 micromolar uye 0.0001% ingangoita 3.13 micromolar. Idzi inhamba diki, uye zviri nyore kufunga kuti idiki zvakanyanya kuti dziite shanduko. Zvidzidzo zviri pasi apa zvinoedza pfungwa iyoyo chaiyo, pahuwandu ihwohwo chaihwo.

Vaverengi vanobva munyaya dzekuchengeta hove panzvimbo perabhoritari vanosangana nemorekuru rimwe chete iri munzvimbo yakasiyana, inotsanangurwa mugwaro remethylene blue muhove dzemuaquarium. Kemisitiri yacho yakafanana; tsananguro yokuti dhayi inotamisa sei maerekitironi mukati memasero inobatsira kunzwisisa kuti nei chinhu chinoshanda mukupa nekugamuchira maerekitironi chichiramba chichionekwa muzvinyorwa zvemetabolism. Zvinotevera zvakanangana nemvura yemazai ezebrafish ine methylene blue.

Metabolism: shanduko yacho inodzokera kune rimwe divi zvichienderana nedanho

Chidzidzo chakapa ruzivo rwakanyanya chakaisa zebrafish dzaikura zvakajairika mumethylene blue kubva pazuva rokutanga roupenyu kusvika pamazuva 5 mushure mekusangana kwezai nourume, chikayera metabolism inoshandisa okisijeni padanho rimwe nerimwe, sezvinotsanangurwa mumushumo wakabudiswa wekuedza metabolism yemazai ari kukura. Maitiro ezvakabuda anofanira kutsanangurwa nokungwarira nokuti shanduko yacho inodzokera kune rimwe divi.

Pazuva 1 mushure mekusangana kwezai nourume, kushandiswa kweokisijeni pakuzorora kwakakwira mumazai ari kukura akaiswa mudhayi pahuwandu hwose huviri hunoshandiswa pakuchengetwa. Pamazuva 2, musiyano wemagetsi pamembrane yemitochondria uri murukoko rwekunze rwetuvana twehove wakakwira pakuiswa mu0.00005%, 0.0001%, uye 0.0003%. Pakazosvika mazuva 4 kusvika 5, mushure mekuchechenywa, shanduko yakanga yadzokera kune rimwe divi: kushandiswa kwesimba kunobva mukushandisa okisijeni kwakaderera, zvisinei nokuti dhayi yakanga iripo nguva dzose kubva pakusangana kwezai nourume kana kuti yaingovapo kwenguva pfupi kubva pazuva 3 kusvika pazuva 4 chete. Panguva imwe chete, zvinyorwa zveRNA zvezvikamu zvemaumbirwo ekufema emitochondria zvakaderera pamazuva 1 ne4, pamwe nezvinyorwa zveRNA zvine chokuita nekugadzirwa kwemahomoni esteroid uye kugadzirwa kweglucose kubva kune zvisiri makabhohaidhiretsi.

Zvimwe zvakaonekwa zviviri zvinojekesa nyaya yacho. Kuchechenywa kwakaitika nokukurumidza zvishoma pahuwandu hwakakwirira hunoshandiswa pakuchengetwa, mazai akaiswa mudhayi achitungamira pamazuva 2 uye boka rekuenzanisa rozobata pazuva 3. Uye hurongwa hwemuviri hunopindura kushushikana hwairatidzika kushanda zvakanaka: tuvana twehove twemazuva 5 patwakaiswa muchinhu chinokonzera kushushikana kuburikidza nekutenderedzwa kwemvura, cortisol mumuviri wose yakakwira zvakajairika mumapoka ose ari maviri. Saka pahuwandu uhu dhayi haisi chepfu inokanganisa kukura kwose, uye haideredzi mhinduro dzose dzemashandiro emuviri. Inochinja metabolism yesimba zvichienderana nedanho, ichikwira kuchechenywa kusati kwaitika uye ichiderera pashure pacho.

Maitiro: nei zvidzidzo zviviri zvisingawirirani, uye nei zvose zvichigona kuva zvakarurama

Zvinyorwa zvemaitiro zvinoita sezvinopesana kusvikira nzira dzekuisa dhayi dzaenzaniswa. Chimwe chidzidzo chekuchengeteka pakukura nemaitiro chakaisa mazai ari kukura mudhayi kubva pamaawa angangoita 6 mushure mekusangana kwezai nourume pahuwandu hunosvika 10 micromolar, chikatsiva mvura ine dhayi zuva nezuva kusvika pazuva 4, ndokuzoendesa tuvana twehove mumvura isina dhayi pamazuva 5 ne6 uye chikaedza kufamba mubvunzo yechiedza nerima pazuva 6. Zvakabuda: hapana shanduko yakakosha pakufa, kuchechenywa, kana chimiro chemuviri, uye hapana shanduko yekufamba yakakonzerwa nekuiswa mudhayi. Vanyori vakakurudzira zvine musoro 10 micromolar semuganhu wepamusoro pasi pemamiriro echidzidzo chavo.

Rimwe boka, raiva nevamwe vanyori vaivawo muchidzidzo chemetabolism, rakaita kuisa dhayi sezvinoitwa pakuchengetwa chaiko: dhayi yaivapo kubva pamaawa 2 kusvika 3 mushure mekusangana kwezai nourume, chikamu chemvura chichitsiviwa zuva nezuva, uye maitiro achiyerwa dhayi ichiripo kana kuti ichangobva kubviswa. Pahuwandu humwe chete ihwohwo hunoshandiswa nguva dzose, tuvana twehove twakaratidza kufamba kushoma mubvunzo yemaawa 24, zvakabuda izvi zvichidzokororwa mumapoka ezebrafish ane nhaka yemajini yakasiyana uye mumarabhoritari akasiyana. Chidzidzo chekukanganiswa kwezvabuda pamaitiro chakabatanidza shanduko yacho nemwero wekushanda kweserotonin, zvichienderana nekudzivisa monoamine oxidase kunozivikanwa kuitwa nedhayi. Chinonyanya kuratidza nyaya yacho ndechokuti mvura yemazai ari kukura yaiva nemethylene blue yakavanza mhinduro yekufamba yaitarisirwa pamushonga weSSRI unonzi fluoxetine. Saka bvunzo yekuongorora inoitwa mumvura yebhuruu inogona kutadza kuona shanduko inokonzerwa nemushonga iyo mvura isina ruvara yaizoratidza.

Hapana kupesana kwechokwadi. Chinhu chinogona kusasiya shanduko inoonekwa mazuva maviri mushure mekubviswa mubvunzo imwe, asi chichiri kushanda pachinenge chiripo mune imwe. Pakushandisa zvakabuda izvi, kusawanikwa kweshando kunotsigira muganhu wepamusoro wehuwandu, kwete pfungwa yokuti chinhu chacho hachichinji zvinoyerwa. Ukuwo, kuwanikwa kweshando kunotsanangura njodzi inokosha: dhayi inoshanda iri kuvapo panguva yokuyera, munzira chaiyo inoshandiswa nemarabhoritari mazhinji.

Dhayi yeredox iri mukati mebvunzo yeredox

Pane mumwezve musungo unofanirwa kuva nendima yawo nokuti unokanganisa nzira yokuyera panzvimbo pemhuka. Bvunzo yeAlamar Blue yekushandiswa kwesimba pachayo inoshanda nekemisitiri yeredox: resazurin inogamuchira maerekitironi ichishanduka kuva resorufin inobudisa chiedza chefluorescence. Methylene blue inoshandawo nekemisitiri yeredox. Mumapoka ekuenzanisa asina masero, huwandu hwose huviri hunoshandiswa pakuchengetwa hwakaderedza fluorescence yeAlamar Blue zvishoma asi pamwero wakakosha maererano nenhamba mushure memaawa 24. Mukuedza pamhuka, mazai ari kukura akaiswa mudhayi akagezwa kanenge katatu mumvura isina dhayi bvunzo isati yaitwa, uye shanduko yemuviri mutuvana twehove twakanga twachechenywa yakapfuura kukanganisa kwakaonekwa pasina masero. Saka zvinhu zvose zviri zviviri ndezvechokwadi: mhuka dzakagezwa dzinenge dzichine shanduko chaiyo yemetabolism, uye dhayi isina kubviswa nekugeza inowedzera kukanganisa kwakananga kunobva kukemisitiri. Mapoka ekuenzanisa anofanira kusiyanisa zvinhu izvi zviviri, kana kuti hapana nhamba yadzo ine zvainoreva. Mirayiridzo yakajairika pamusoro pekududzira zvabuda mubvunzo dzinoshandisa dhayi ine kukosha kwakanyanya pano.

Nguva dzematanho ekukura

Ichi chinhu chepakutanga chakagadzirirwa chinyorwa chino. Huwandu hwacho ndihwo huviri hunowanzo shandiswa pakuchengetwa, 0.00005% ne0.0001%, kunze kwepanotaurwa zvimwe.

Chinofanira kuyeukwa kukwira kwekutanga uye kuderera kwepashure kwezviratidzo zvemetabolism, pamwe neshanduko dzemaitiro dziripo dhayi ichiripo asi dzisingachaoneki mushure mokubviswa kwayo mubvunzo imwe chete yakaongorora izvi.

Rondedzero yekuongorora kurongwa kwemapoka ekuenzanisa

Ichi ndicho chinhu chechipiri chepakutanga chakagadzirirwa chinyorwa chino. Chishandise apo nzira dzekushandisa methylene blue muzebrafish dzinosangana nekuedza.

1. Nyora dhayi sechinhu chinoshandiswa mukuedza, kwete semvura. Taura huwandu hwacho muzvikamu zvezana uye mumicromolar, nguva yokutanga neyokumira mumaawa kana mazuva mushure mekusangana kwezai nourume, hurongwa hwekutsiva mvura, uye nguva yekubviswa kwedhayi isati yaitwa kuyerwa kumwe nokumwe.
2. Ronga mapoka ekuenzanisa maererano nemubvunzo. Chidzidzo chekemikari iri kuedzwa chinoda mapoka ekuenzanisa asina dhayi; chidzidzo chekuchengetwa chinoda mapoka asina dhayi achienzaniswa neane dhayi, pachinyorwa zvakabuda pakudzivirira chakuvhe, nokuti hapana pazvidzidzo zviviri zvataurwa chakaera kushanda kwekudzivirira chakuvhe.
3. Geza bvunzo dzeredox dzisati dzaitwa, wobva wasimbisa. Geza kanenge katatu mumvura isina dhayi uye ita pamwe chete bvunzo yekuenzanisa isina masero ine dhayi pamwe nechinhu chinoshandiswa mubvunzo, nokuti dhayi inoderedza chiratidzo cheAlamar Blue zvakananga.
4. Ona maitiro nezviratidzo zvemonoamine sezviri panjodzi huru yekukanganiswa. Zvidzidzo zvekufamba, kuvhunduka, maitiro akafanana nekuzvidya mwoyo, mishonga yekuderedza kuora mwoyo, uye zvinodzivisa MAO zvinofanira kudzivisa dhayi panguva yokuyera kana kuratidza kuti haichinji chiratidzo ichocho; kubviswa kwedhayi kwemazuva rimwe kusvika maviri kwakabvisa imwe shanduko yekufamba asi hakuna kusimbiswa kune dzimwe nzira dzekuedza.
5. Ona zviratidzo zvemetabolism nezvemitochondria sezviri panjodzi huru yekukanganiswa padanho rose. Kushandiswa kweokisijeni, kushandiswa kwesimba, musiyano wemagetsi pamembrane, uye zvinyorwa zveRNA zvezvikamu zvinofambisa kufema zvinofamba pahuwandu hunoshandiswa pakuchengetwa, zvichienda kumativi anopesana kuchechenywa kusati kwaitika uye pashure pacho.
6. Usafungidzira kuti kusachinja zvinoyerwa kunowanikwa mune imwe nzira yekuedza kunoshanda mune imwe. Kusawanikwa kweshando mushure mekubviswa kwedhayi hakubatanidzi mamiriro ayo dhayi ichiripo; kusawanikwa kweshando pachimiro chemuviri hakubatanidzi metabolism kana kudyidzana nemushonga; zvakabuda muboka rimwe rine nhaka yemajini yakati, pane imwe tembiricha, kana panguva yakati yebvunzo hazvibatanidzi mamwe mamiriro.
7. Taura mirayiridzo yawakatevera neimwe nzira iripo. Doma zvakajeka huwandu hweCCAC, hweEurope, kana hweCold Spring Harbor, uye ratidza apo bvunzo yeOECD yemazai ari kukura isingabatanidzi dhayi, kuitira kuti muverengi anzwisise mamiriro akashandiswa pamhuka dzako.

Hapana pane izvi chinopikisa kushandisa dhayi kuchengetedza mazai ari mapenyu. Zvinopikisa kurega sarudzo yekuchengetwa ichipinda muzvabuda pasina kuongororwa. Bhuruu riri mundiro imorekuru rine mashandiro emushonga, uye kuedza kunonyanya kunge kuchikanganiswa ndidzo bvunzo dzemetabolism uye dzemaitiro ane chokuita netsinga dzinoongorora zvakawanda nokukurumidza, dziri kunyanya kusarudzirwa zebrafish diki mazuva ano. Nyora kushandiswa kwayo, ronga mapoka ekuienzanisa, ibvise nekugeza usati wayera, uye ongorora bvunzo yainogona kunge ichibatanidzwa mairi chinyararire.