# Presentation: 258112

## Dopamine Transporter:

** **
**Are Psychostimulants in Your Neighborhood **
**Forcing You to Move?**

## “Brain & Mind” Magazine

- “Brain & Mind” Magazine

## “Brain & Mind” Magazine

- “Brain & Mind” Magazine

## “Brain & Mind” Magazine

- “Brain & Mind” Magazine

## “Brain & Mind” Magazine

- “Brain & Mind” Magazine

## AMPH-STIMULATED DA EFFLUX

- “Brain & Mind” Magazine

**AMPH-STIMULATED DA EFFLUX**

## AMPH-INDUCED 

- “Brain & Mind” Magazine

**AMPH-INDUCED **
**hDAT TRAFFICKING**

## AMPH-INDUCED 

- “Brain & Mind” Magazine

**AMPH-INDUCED **
**hDAT TRAFFICKING**

## AMPH-STIMULATED 

- DA EFFLUX

- AMPH-INDUCED hDAT TRAFFICKING

- REGULATION OF AMPH-INDUCED
- hDAT TRAFFICKING

## UPTAKE 

- OUT

- IN

## EFFLUXES 

- OUT

- IN

## Facilitated Exchange

**OUT**

**IN**

- DA

- AMPH

## AMPH INCREASES [Na]i

- AMPH INCREASES [Na]i

## rest

**rest**

**0 s**

**15 s**

**30 s**

**45 s**

**60 s**

**75 s**

**AMPH**

## AMPEROMETRY

## Slide 16

## Slide 17

## B

- <date/time>

**CTR**

**AMPH**

**COC**

**B**

**A**

**C**

*****

*****

*****

## <footer>

- <date/time>

## AMPH-INDUCED hDAT

- DA EFFLUX

-  TRAFFICKING

## 0

- 255

## <footer>

- <date/time>

**Total**

**Biotin.**

**CTR**

**AMPH**

**Ins+AMPH**

## Slide 23

## Patch-Clamp and Confocal Imaging

**Noise Analysis of Steady-State Currents **

**Transient and Steady-State Current Analysis**

**Transient and Steady-State Current Analysis**

## AMPH-induced hDAT current

- <date/time>

**-160 mV**

**-20 mV**

**-20 mV**

**50 pA**

**250 msec**

**Transient Current**
**(hDAT Number)**

**Steady State Current**
**(hDAT Activity)**

**AMPH-induced hDAT current**

## NT

- <date/time>

**NT**

**Cell-Surface**

**Total**

**CTR   5   10   15   20   30**

**hDAT**

**25 msec**

**-120 mV**

**-20 mV**

**5**

**10**

**15**

**20**

**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**

**100 pA**

## Slide 27

## Slide 28

## Slide 29

## <footer>

- <date/time>

## Is AMPH-Induced 

**Is AMPH-Induced **
**Transporter Activity Responsible for hDAT Trafficking? **

## 20 min

- <date/time>

**Basal**

**10 min**

**20 min**

## WT

**10**

**20**

**30**

**40**

**50**

**60**

**0.5**

**1.0**

**1.5**

**2.0**

**2.5**

**3.0**

**3.5**

**WT**

**CTR**

**AMPH**

**Internal Fluorescence**

**(% of Control)**

**Time (min)**

**0**

**10**

**20**

**30**

**40**

**50**

**60**

**0.5**

**1.0**

**1.5**

**2.0**

**2.5**

**3.0**

**3.5**

**CTR**

**AMPH**

**Y335A**

**Internal Fluorescence**

**(% of Control)**

**Time (min)**

## WT

**10**

**20**

**30**

**40**

**50**

**60**

**0.5**

**1.0**

**1.5**

**2.0**

**2.5**

**3.0**

**3.5**

**WT**

**AMPH + Zn****2+**

**Time (min)**

**0**

**10**

**20**

**30**

**40**

**50**

**60**

**0.5**

**1.0**

**1.5**

**2.0**

**2.5**

**3.0**

**3.5**

**Y335A**

**AMPH + Zn****2+**

**Time (min)**

## AMPH-induced channel like activity supports DA Efflux

**hDAT is active until removed from the plasma membrane**
**The activity of hDAT is required for AMPH-induced trafficking**

## Collaborators

**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