Drosophila melanogaster Ana Filosevic 1 Sabina Al Samarai 1 Josipa Kolobaric 1 and Rozi Andretic Waldowski 1 Laboratory for behavioral genetics Department of biotechnology University of Rijeka ID: 1046031
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1. Locomotor sensitization to cocaine and methamphetamine in Drosophila melanogaster Ana Filosevic1, Sabina Al Samarai1, Josipa Kolobaric1 and Rozi Andretic Waldowski1Laboratory for behavioral geneticsDepartment of biotechnology, University of Rijeka, Rijeka, CROATIA
2. Drosophila melanogaster as a model organism in addictionAddiction related endophenotypesPhenotypes validated with known genesNew genes identified with conserved function in mammalsPower of Drosophila in unbiased gene discoveryHISTORY:Heberlein Lab
3. Drugs induce addiction related endophenotypes COCAINE ETHANOLACUTE Sensitivity SensitivityCHRONIC Behavioral Tolerance sensitization Voluntary consumption Conditioned preference for rewardMcClung C, Hirsh J.Curr Biol. 1999; McClung C, Hirsh J.Curr Biol. 1998; Andretic R et al., Science. 1999; Heberlein U.et al., Neuropharmacol. 2009Bainton RJ. Et al,Curr Biol. 2000Scholz H. et al.,Neuron. 2000 Bainton RJ, et al.Cell. 2005 Rothenfluh A, et al.,Cell. 2006 Eddison M, et al., Neuron. 2011Shohat-Ophir G,et al.,Science. 2012Drosophila melanogaster as a model organism in addiction
4. Drosophila as a model for new gene discoveryJay Hirsh, UVAAndretic et al, Science, 1999.Abarca et al.,PNAS 2002
5. Power of Drosophila genetics in unbiased gene discovery using genetic screensFrom phenotype to genotypeHigh throughput method that is objective, reproducible and specificMUTAGENIZE TEST IDENTIFY STUDYFORWARD GENETICS:
6. FLYBONG - a new, highthroughput platform for measuring drug induced locomotor activityMajor advantages:Minimal fly handlingPrecise dosing of individual fliesObjective quantification of locomotor behaviorPopulation vs. individual analysisHigh throughput
7. Objective and quantitative recording of locomotor activityTriKinetics Drosophila activity monitoring systemAdaptation for volatilized drug delivery
8. 8 min1 min30 min. BEFORE +30 min. AFTERPROTOCOL:FLYBONG - a new, highthroughput platform for measuring drug induced locomotor activity
9. Volatilized COC transiently and dose-dependently increases locomotor activityINDIVIDUAL FLY RESPONSEPOPULATION RESPONSE
10. Repeated exposures to the same COC concentration induce locomotor sensitizationPOPULATIONINDIVIDUAL
11. Validation of COC effects using circadian gene mutants2 X 75 mg COC6 hrs intervalCONTROL2 X warm air6 hrs interval
12. Dopamine transporter is required for development of locomotor sensitizationPOPULATION2 X 75 mg COCINDIVIDUAL2 X 75 mg COCCONTROLSwarm airDAT mutant ishyperactiveDAT mutant issensitive to COCSensitivity is not COC specific
13. Conclusions I:There is individual variation in sensitivity to motor-activating effects of COC- A subset of sensitive flies develops LSFlyBong distinguishes between COC specific and non-specific effects Genes per, Clk, cyc and dopaminergic transporter required for neural plasticity
14. Repeated METH exposures induce locomotor sensitization
15. Functional DAT gene is required for locomotor sensitization to METH POPULATIONINDIVIDUALDAT mutant isnot sensitive to COCDAT is required for locomotor sensitization
16. Locomotor sensitization to METH and COC involve different time interval and circadian genes cd,ecd,eccd,eecece wt ClkJrk cyc01 per01 tim01 pdf01*#TIME INTERVAL 10 HRSCIRC. GENES ClkJrk
17. Is there dual role for circadian genes in locomotor sensitization Some circadian genes REQUIRED for locomotor sensitization - Clk, per and cyc for COC - Clk for METHDoes circadian system (genes) MODULATE sensitivity or locomotor sensitization as a function of time of day?
18. Locomotor sensitization, but not sensitivity varies as a function of the time of day3 DOSES OVER 2 DAYS2 DOSES AT DIFFERENT TIMESSensitivity is stableSensitivity to the 2nd dose changes - LSNeuronal plasticity is under time of day modulation
19. CONCLUSIONS:Distinct parameters of behavioral responses to COC and METHDifferent involvement of circadian genesCircadian effect on modulation of LS vs. requirement of circadian genes in mechanisms of LSNEXT:Genetic screenSelection experimentWhich gene, where in the brain?
20. Research Grant HRZZ br. 4920; Doctoral Support HRZZ 2015University of Rijeka Research SupportSABINAJOSIPAAZRAFUNDING:COLLABORATORS:Ana Filosevic(poster # 189)ACKNOWGLEDGMENTS: