1. Dopamine Transporter: Are Psychostimulants in Your Neighborhood Forcing You to Move?

2. “Brain & Mind” Magazine

6. AMPH-STIMULATED DA EFFLUX

7. “Brain & Mind” Magazine AMPH-INDUCED hDAT TRAFFICKING

8. “Brain & Mind” Magazine AMPH-INDUCED hDAT TRAFFICKING

9. AMPH-STIMULATED DA EFFLUX AMPH-INDUCED hDAT TRAFFICKING REGULATION OF AMPH-INDUCED hDAT TRAFFICKING

10. UPTAKE OUT IN

11. EFFLUXES OUT IN

12. OUT IN DA AMPH Facilitated Exchange

13. OUT IN AMPH Na DA AMPH INCREASES [Na] i

14. rest0 s15 s30 s45 s60 s75 s AMPH

15. AMPEROMETRY Electrode Amperometric 700 mV Patch Electrode Dopamine 2e -

16. 700 mV DA 2e - 200 pA 500 msec 0.1 pA

18. CTR AMPH COC BA 1 2 Patch Clamp C * * *

19. * B -20 mV +40 mV +100 mV * * C STEP 2 sec 0.1 pA Amperometry +700mV A DA 1 2 Patch Clamp * * *

20. AMPH-STIMULATED DA EFFLUX AMPH-INDUCED hDAT TRAFFICKING

21. 0255

22. Total Biotin. CTR AMPHIns+AMPH 0 50 100 150 * % of control AMPH Ins+AMPH

23. AMPH

24. Patch-Clamp and Confocal Imaging Noise Analysis of Steady-State Currents Transient and Steady-State Current Analysis

25. -160 mV -20 mV 50 pA 250 msec Transient Current (hDAT Number) Steady State Current (hDAT Activity) AMPH-induced hDAT current

26. NT Cell-Surface Total CTR 5 10 15 20 30 hDAT 25 msec -120 mV -20 mV 5101520 0.5 1.0 1.5 Biotinylation Transient Charge (pC) 30 Time (min) % of Control -5-10-15-20 -50 -100 -150 -200 r = 0.88 p < 0.001 slope = 8.35 ± 1.5 I (pA) Q (pC) AMPH (min) 100 pA

27. AMPH-induced hDAT current Whole-cell Patch Clamp AMPH FINAL

28. AMPH-induced hDAT current Final Transient Charge Q / Q -160mV -160-120-80-40 -1.5 -0.5 0.0 0.5 1.0 1.5 Initial "ON" Initial "OFF" Membrane Potential (mV) Whole-cell Patch Clamp

29. AMPH

31. Is AMPH-Induced Transporter Activity Responsible for hDAT Trafficking?

32. Basal10 min 20 min Y335A-DAT AMPH 0255 Y335A-DAT AMPH+Zinc

33. 0102030405060 0.5 1.0 1.5 2.0 2.5 3.0 3.5 WT CTR AMPH Internal Fluorescence (% of Control) Time (min) 0102030405060 0.5 1.0 1.5 2.0 2.5 3.0 3.5 CTR AMPH Y335A Internal Fluorescence (% of Control) Time (min)

34. 0102030405060 0.5 1.0 1.5 2.0 2.5 3.0 3.5 WT AMPH + Zn 2+ Internal Fluorescence (% of Control) Time (min) 0102030405060 0.5 1.0 1.5 2.0 2.5 3.0 3.5 Y335A AMPH + Zn 2+ Internal Fluorescence (% of Control) Time (min)

35. AMPH-induced channel like activity supports DA Efflux AMPH-induced channel like activity supports DA Efflux hDAT is active until removed from the plasma membrane hDAT is active until removed from the plasma membrane The activity of hDAT is required for AMPH-induced trafficking The activity of hDAT is required for AMPH-induced trafficking

36. Collaborators Christine Saunders Jonathan Javitch Bill Clarke Kelly Berg NIDA James Lechleiter Toni Shippenberg Brandon Lute Teri Wei Greta Galli Erika Galli Hongwei Wang Kris Kahlig Concetta Dipace Francesca Binda Habibeh Khoshbouei Emily Schwartz Oliver Luo Lyn Daws Charles France Ulrik Gether