Title The effect of methamphetamine on the redox status of Drosophila melanogaster
Title (croatian) Utjecaj metamfetamina na redoks status kod Drosophila melanogaster
Author Valentina Dukić
Mentor Rozi Andretić Waldowski (mentor)
Mentor Ana Filošević (komentor)
Committee member Ivana Munitić (predsjednik povjerenstva)
Committee member Elitza Petkova Markova-Car (član povjerenstva)
Granter University of Rijeka (Faculty of Biotechnology and Drug Development) Rijeka
Defense date and country 2020-09-08, Croatia
Scientific / art field, discipline and subdiscipline BIOTECHNICAL SCIENCES Biotechnology
Abstract Methamphetamine (METH) is a psychostimulant that affects central nervous system by increasing the concentration of the free dopamine, whose oxidation leads to the permanent damage in the structure and function of neurons, as the consequence of the increased production and accumulation of the radical oxygen species (ROS).
By using Drosophila melanogaster as a model organism we have tested impact of an acute dose of volatilized METH (vMETH) on the concentration of hydrogen peroxide (H2O2), in the brain, as one of the ROS and redox status representative. Individual flies of wild type strain CantonS (wt) used in the experiment were exposed to vMETH using the FlyBong method. The concentration of H2O2 was measured in the head tissue extracts, 10 hours after the administration, using dihydroetidium florescent dye. The results have shown that the acute exposure to vMETH does not lead to the change in the concentration of H2O2 in the wt flies compared to the control group.
Considering that research have shown connection between administration of the METH and increased expression of period (per) gene with enzyme activity of tyrosine hydroxylase, included in the synthesis of dopamine, I tested whether there is a difference in the concentration of H2O2 after the acute exposure to vMETH between different circadian mutants and wt flies. Circadian mutants of genes per, timeless (tim), Clock (Clk) and cycle (cyc) with alleles per01, tim01, ClkJrk and cyc01 were used in the experiment. per01 and tim01 flies showed the decreased concentration of H2O2 compared to the controls while ClkJrk and cyc01 flies had higher concentration of H2O2 after the acute exposure to vMETH. The concentration of H2O2 is significantly lower in tim01 flies compared to wt flies after the exposure to vMETH while cyc01 flies had a significantly higher concentration than wt flies.
Based on the results of this thesis, we suggest the influence of circadian genes on the endogenous concentration of H2O2, which is in the absence of per or tim increased, and in the absence of Clk or cyc decreased compared to wt. The administration of vMETH potentiates the production of H2O2 in the absence of cyc while the absence of tim gene attenuates that effect. The lack of difference in wt flies is surprising and possible mistakes in the method or its use should be considered. By establishing the molecular pathways involving H2O2 and circadian genes new methods for treatment of addiction could be developed.
Abstract (croatian) Metamfetamin (METH) je psihostimulans koji utječe na središnji živčani sustav povećavajući koncentraciju slobodnog dopamina, čija oksidacija uzrokuje trajna oštećenja strukture i funkcije neurona kao posljedicu povećane proizvodnje i akumulacije slobodnih kisikovih radikala (ROS-a).
Korištenjem Drosophile melanogaster kao modelnog organizma istražen je utjecaj akutne doze volatiliziranog METH-a (vMETH) na koncentraciju vodikovog peroksida (H2O2), u mozgu, kao jednog od predstavnika ROS-a i redoks statusa. U eksperimentu su korištene individualne mušice divljeg tipa CantonS (wt) izložene vMETH–u korištenjem FlyBong metode. Koncentracija H2O2 je mjerena u ekstraktima tkiva glava mušica 10 sati nakon administracije upotrebom dihidroetidij fluorescentne boje. Rezultati su pokazali da akutno izlaganje vMETH-u ne dovodi do promjene u koncentraciji H2O2 kod wt mušica u odnosu na kontrolu.
S obzirom na literaturno poznatu povezanost administracije METH-a s povećanom ekspresijom period (per) gena i aktivnosti enzima uključenog u sintezu dopamina, tirozin hidroksilaze, testirano je ima li razlike u koncentraciji H2O2 nakon akutne izloženosti vMETH-u između različitih cirkadialnih mutanata i wt mušica. Za eksperimente su korišteni cirkadialni mutanti u genima per , timeless (tim), Clock (Clk) i cycle (cyc) sa sljedećim alelima: per01, tim01, ClkJrk i cyc01. per01 i tim01 mušice su pokazale smanjenu koncentraciju H2O2 u odnosu na svoje kontrole, dok su ClkJrk i cyc01 mušice imale veću koncentraciju H2O2 nakon akutnog izlaganja vMETH-u. Koncentracija H2O2 je značajno niža kod tim01 mušica u usporedbi s wt mušicama nakon izlaganja vMETH-u, dok cyc01 mušice imaju značajno veću koncentraciju nego wt mušice.
S obzirom na rezultate ovoga rada predlažemo utjecaj cirkadialnih gena na endogenu koncentraciju H2O2 koja je u odsustvu per ili tim povećana, a u odsustvu Clk ili cyc smanjena u odnosu na wt. Administracija vMETH-a potiče proizvodnju H2O2 u odsustvu cyc dok odsustvo tim gena sprječava taj efekt. Nedostatak razlike kod wt mušica je iznenađujući te je potrebno razmotriti moguće pogreške u metodi ili njezinom provođenju. Utvrđivanjem molekularnih putova koji uključuju H2O2 i cirkadialne gene mogle bi se razviti nove metode liječenja ovisnosti o METH-u.
Keywords
Drosophila melanogaster
methamphetamine
circadian genes
hydrogen peroxide
Keywords (croatian)
Drosophila melanogaster
metamfetamin
cirkadijalni geni
vodikov peroksid
Language english
URN:NBN urn:nbn:hr:193:434929
Study programme Title: Biotechnology and drug research Study programme type: university Study level: undergraduate Academic / professional title: sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) biotehnologije i istraživanja lijekova (sveučilišni/a prvostupnik/prvostupnica (baccalaureus/baccalaurea) biotehnologije i istraživanja lijekova)
Type of resource Text
File origin Born digital
Access conditions Open access
Terms of use
Repository Repository of the University of Rijeka, Faculty of Biotechnology and Drug Development
Created on 2020-09-10 08:49:43