Animal model of Wolfram Syndrome in mice: behavioural, biochemical and psychopharmacological characterization
Kuupäev
2013-09-18
Autorid
Ajakirja pealkiri
Ajakirja ISSN
Köite pealkiri
Kirjastaja
Abstrakt
Wolframi sündroom (WS), mida kirjeldasid esmakordselt Saksa arstid Wolfram ja Wagener 1938.a, on haruldane autosomaalne retsessiivne neurodegeneratiivne häire, mida iseloomustavad varajases lapseeas algav suhkurtõbi, progresseeruv nägemisnärvi atroofia, magediabeet ja kurtus. Wolframi sünroomi põhjustavad mutatsioonid Wolframiin 1 (WFS1) geenis. Wolframiin 1 geeni on seotud hirmu ja ärevusega hiirtel ja rottidel ja inimestel on tema polümorfismid on seotud suurenenud meeleoluhäirete riskidega nagu bipolaarseste häirete psühhootilise vormi, skisofreenia, suitsidaarne käitumine ning depressioon.
Käesoleval ajal ei ole veel päris selge, mis moodi WFS1 geeni puudulikkus põhjustab neuropsühhiaatriliste kõrvalekallete ilmnemist. Kuid võttes arvesse WS puhul esinevad psühhiaatrilised sümptomid ning ka Wfs1 geeni ja valgu lokalisatsiooni ajus, võib arvata, et ühetedeks põhjustajateks võivad olla kõrvalekalded GABA- ja dopamiinergilise süsteemi talitluses.
Käesoleva doktoritöö keskseks eesmärgiks oli WS hiire mudeli loomine. Selleks valmistati mutantne hiireliin kustutades Wfs1 geenist 8nda eksoni, kuna WS patsientidel enamik mutatsioone asub just selles eksonis. Selline lähenemisviis aitas luua WS sarnase seisundi, et uurida sündroomi poolt esilekutsutud patoloogilisi muutusi.
Katseloomade iseloomustamiseks kasutati käitumuslikke, biokeemilisi ja psühhofarmakoloogilisi uurimismeetodeid.
Selgus, et Wfs1 mutantsetel hiirtel ei olnud nähtavaid sensoorseid häireid, kuid nad reageerisid rohkem stressi esilekutsuvates keskkondades (testides). Samuti suurenes neil tunduvalt enam, võrreldes metsik-tüüpi hiirtega, korikosterooni tase ringlevas veres peale füsioloogilise lahuse akuutset süstimist ning nad ei suuda kontrollida oma veresuhkru taset glükoosi tolerantsi testis. Wfs1 puudulikkusega hiirtel oli suurenenud tundlikkus GABAA retseptorite agonisti diasepaami ärevusvastase toime suhtes. Diasepaam kõrvaldas ka Wfs1 puudulikksuega hiirtele iseloomulikud häälitsemised stressogeenses situatsioonis. Neil oli vähenenud Gabra1 ja Gabra2 geenide eskpressioon oimusagaras ja otsmikukoores ehk ajustruktuurides, mis on seotud negatiivsete emotsioonide regulatsiooniga. See ekspressiooni langus on ilmselt seotud Wfs1 puudulikkusega hiirtel esineva ülitundlikkusega diasepaami ärevusvastase toime suhtes. Sarnane Gabra1 ja Gabra2 geeniekspressiooni langus esines metsik-tüüpi hiirtel peale nende eksponeerimist pluss puuris (ärevuse mudel), mis viitab, et on olemas seos GABAA retseptorite alatüüpide ekspressiooni languse ja ärevuse vahel.
Lisaks kõigele muule esinesid nendel hiirtel kõrvalekalded dopamiinergilise süsteemi funktsioonis. Nende tundlikkus amfetamiini, kaudse dopamiini agonisti, motoorikat stimuleeriva toime suhtes oli vähenenud ning amfetamiini manustamine ei põhjustanud neil dopamiini ringkäiku muutusi dorsaalses ja ventraalses striatumis võrreldes metsiktüüpi pesakonnakaaslastega. Apomorfiinist tingitud motoorika stimulatsioon oli mõnevõrra tugevam mutantsetel hiirtel, kuid apomorfiin põhjustas kõikidel genotüüpidel ühesugust dopamiini ringkäiku vähenemist, mille alusel võib väita, et Wfs1 geeni puudulikkus ei mõjusta dopamiini metabolismi dopamiini retseptorite vahendusel. Seda kinni¬tavad ka geeni ekspressiooni tulemused, kus dopamiini retseptori (Drd2) ekspressioon oli ühesugune kõikide genotüüpide ventraalses striatumis. Siiski oli nii emastel kui ka isastel Wfs1 puudulikusega hiirtel vähenenud dopamiini transporteri geeni ekspressioon keskajus. See leid koos amfetamiini käitu-muslike ja biokeemiliste toimete vähenemisega viitab dopamiinergilise süsteemi funktsiooni olulisele häirumisele Wfs1 puudulikkusega hiirtel.
Põhinedes glükoosi metabolismi uuringute tulemustele ja nende sarnasusele WS on selge, et Wfs1 geeni kaheksanda eksoni puudulikkusega hiir on valiidseks mudeliks WS südnroomile ja ka teistele WFS1 geenimutatsioonidest tingitud häiretele. Sellepärast võib käesolevas uuringus leitud muutusi dopamiini- ja GABA-ergilise süsteemi aktiivsuses Wfs1 puudulikkusega hiirtel laiendada WS patsientidele, et mõista nendel esinevaid Neuropsühhiaatrilisi Sümptomeid.
Wolfram syndrome (WS), first described by German physicians Wolfram and Wagener in 1938, is a rare autosomal reces¬sive neurodegenerative disorder characterised by early juvenile diabetes mellitus, progressive optic nerve atrophy, diabetes insipidus and deafness. Wolfram syndrome is caused by mutations in the wolframin (WFS1) gene. In mice and rats, the wolframin gene has been associated with fear and anxiety; in humans, there is a link between WFS1 polymorphisms and increased risk for mood disorders, such as the psychotic form of bipolar disorder, schizophrenia, suicidal behaviour and depression. At present, it is not yet quite clear, how disruptions in the WFS1 gene cause neuropsychiatric deviations. However, based on the fact that WS is accompanied by psychiatric symptoms and also based on the localisation of the Wfs1 protein in the brain it can be concluded that at least partially it is caused by deviations in the functioning of the GABAergic and dopaminergic systems. The main objective of this dissertation was the creation of a WS mouse model. For that purpose, a mutant mouse line was created by deleting exon 8 in the Wfs1 gene as in WS patients most of the mutations are located in that exon. By this approach we managed to mimic WS in order to study pathological changes induced by the syndrome. To characterise the animals, behavioural, biochemical and psychopharmacological methods were used. Wfs1 mutant mice had no overt sensory deficits, however, they were more sensitive to stress-inducing environments (tests). Also, as compared to wild-type mice, they displayed a much more prominent increase in the level of corticosterone in the circulating blood in response to acute saline injection, and they were unable to keep their blood glucose level under control in the glucose tolerance test. Wfs1-deficient mice had increased sensitivity to the anxiolytic effect of GABA A recep¬tor agonist diazepam. Also, in Wfs1 mice diazepam blocked characteristic vocalisations in a stressful situation. Wfs1-deficient mice had lower expression level of Gabra1 and Gabra2 genes in the temporal and frontal lobes, brain structures involved in the regulation of negative emotions. These decreased expression levels are probably related to hypersensitivity to the anxiolytic effect of diazepam observed in Wfs1-deficient mice. In wild-type mice, a similar drop in the expression levels of Gabra1 and Gabra2 genes was evident after exposure to the plus maze (a model of anxiety), which indicates that there is a link between decreased expression levels of GABA A receptor subtypes and anxiety. Furthermore, mice lacking the wolframin protein had deviations in the function of the dopaminergic system. Their sensitivity to the motor stimulant effect of amphetamine, an indirect agonist of dopamine, had decreased and the administration of amphetamine failed to induce changes in dopamine turnover in the dorsal and ventral striatum as opposed to their wild-type littermates. Apomorphine-induced motor stimulation was somewhat stronger in mutant mice, but in all genotypes apomorphine caused a similar decrease in dopamine turnover. This enables us to conclude that Wfs1 gene deficiency has no effect on dopamine metabolism mediated by dopamine receptors. This is corroborated by the results of gene expression measurements showing that in the ventral striatum the expression level of dopamine D2 receptor (Drd2) was equal in all genotypes. However, both in male and female Wfs1-deficient mice the expression level of the dopamine transporter gene had decreased in the mesencephalon. This finding and also the decreased behavioural and biochemical effects of amphetamine are indicative of serious deviations in the function of the dopaminergic system in Wfs1-deficient mice. Based on the results of glucose metabolism studies and their similarity to WS, it is clear that the mouse, missing exon 8 of the Wfs1 gene, is a valid model of both WS syndrome and other disorders caused by mutations in the WFS1 gene. Therefore, the changes observed in the activity of the dopaminergic and GABAergic systems in Wfs1-deficient mice may also help to explain the neuropsychiatric symptoms of WS patients.
Wolfram syndrome (WS), first described by German physicians Wolfram and Wagener in 1938, is a rare autosomal reces¬sive neurodegenerative disorder characterised by early juvenile diabetes mellitus, progressive optic nerve atrophy, diabetes insipidus and deafness. Wolfram syndrome is caused by mutations in the wolframin (WFS1) gene. In mice and rats, the wolframin gene has been associated with fear and anxiety; in humans, there is a link between WFS1 polymorphisms and increased risk for mood disorders, such as the psychotic form of bipolar disorder, schizophrenia, suicidal behaviour and depression. At present, it is not yet quite clear, how disruptions in the WFS1 gene cause neuropsychiatric deviations. However, based on the fact that WS is accompanied by psychiatric symptoms and also based on the localisation of the Wfs1 protein in the brain it can be concluded that at least partially it is caused by deviations in the functioning of the GABAergic and dopaminergic systems. The main objective of this dissertation was the creation of a WS mouse model. For that purpose, a mutant mouse line was created by deleting exon 8 in the Wfs1 gene as in WS patients most of the mutations are located in that exon. By this approach we managed to mimic WS in order to study pathological changes induced by the syndrome. To characterise the animals, behavioural, biochemical and psychopharmacological methods were used. Wfs1 mutant mice had no overt sensory deficits, however, they were more sensitive to stress-inducing environments (tests). Also, as compared to wild-type mice, they displayed a much more prominent increase in the level of corticosterone in the circulating blood in response to acute saline injection, and they were unable to keep their blood glucose level under control in the glucose tolerance test. Wfs1-deficient mice had increased sensitivity to the anxiolytic effect of GABA A recep¬tor agonist diazepam. Also, in Wfs1 mice diazepam blocked characteristic vocalisations in a stressful situation. Wfs1-deficient mice had lower expression level of Gabra1 and Gabra2 genes in the temporal and frontal lobes, brain structures involved in the regulation of negative emotions. These decreased expression levels are probably related to hypersensitivity to the anxiolytic effect of diazepam observed in Wfs1-deficient mice. In wild-type mice, a similar drop in the expression levels of Gabra1 and Gabra2 genes was evident after exposure to the plus maze (a model of anxiety), which indicates that there is a link between decreased expression levels of GABA A receptor subtypes and anxiety. Furthermore, mice lacking the wolframin protein had deviations in the function of the dopaminergic system. Their sensitivity to the motor stimulant effect of amphetamine, an indirect agonist of dopamine, had decreased and the administration of amphetamine failed to induce changes in dopamine turnover in the dorsal and ventral striatum as opposed to their wild-type littermates. Apomorphine-induced motor stimulation was somewhat stronger in mutant mice, but in all genotypes apomorphine caused a similar decrease in dopamine turnover. This enables us to conclude that Wfs1 gene deficiency has no effect on dopamine metabolism mediated by dopamine receptors. This is corroborated by the results of gene expression measurements showing that in the ventral striatum the expression level of dopamine D2 receptor (Drd2) was equal in all genotypes. However, both in male and female Wfs1-deficient mice the expression level of the dopamine transporter gene had decreased in the mesencephalon. This finding and also the decreased behavioural and biochemical effects of amphetamine are indicative of serious deviations in the function of the dopaminergic system in Wfs1-deficient mice. Based on the results of glucose metabolism studies and their similarity to WS, it is clear that the mouse, missing exon 8 of the Wfs1 gene, is a valid model of both WS syndrome and other disorders caused by mutations in the WFS1 gene. Therefore, the changes observed in the activity of the dopaminergic and GABAergic systems in Wfs1-deficient mice may also help to explain the neuropsychiatric symptoms of WS patients.
Kirjeldus
Väitekirja elektrooniline versioon ei sisalda publikatsioone.
Märksõnad
autosoomne retsessiivne pärilikkus, Wolframi sündroom, loomkatsed, kliinilised uuringud, autosomal recessive heredity, Wolfram syndrome, clinical trials