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DA Transporter Currents: A Reason for Excitement

DA Transporter Currents: A Reason for Excitement. Susan L. Ingram WSU-Vancouver, Vancouver WA. Neurotransmitter transporters: Transport is electrogenic. Neurotransmitter. DA, 5HT, NE, GABA, Glycine . +. 1. -. 3 Na. -. 1Cl. Substrate-gated uncoupled conductance. cytoplasm.

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DA Transporter Currents: A Reason for Excitement

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  1. DA Transporter Currents: AReason for Excitement Susan L. Ingram WSU-Vancouver, Vancouver WA

  2. Neurotransmitter transporters: Transport is electrogenic Neurotransmitter DA, 5HT, NE, GABA, Glycine + 1 - 3Na - 1Cl Substrate-gated uncoupled conductance cytoplasm extracellular space

  3. Questions • Does activation of DAT produce measurable currents in DA neurons? • What ions are responsible? • Is the current large enough to change excitability of the DA neurons? • Does the current play a role in neurotransmitter release?

  4. Presynaptic cell MGluR GIRK D2 X X Amph COC D2 D1 a1 Postsynaptic cell Dopaminergic Synapse DA DA

  5. Raclopride Sulpiride Prazosin TTX biocytin Whole-cell patch clamp recordings from DA neurons in culture

  6. Does activation of DAT produce measurable currents?

  7. DAT substrates activate an inward current

  8. 50 Current (pA) 10-9 10-9 10-5 10-5 [DA] 0.1 Uptake (fmol/cell/s) [DA] DAT currents are uncoupled from transport

  9. What ions are responsible?

  10. DAT currents are carried by chloride ions control nitrate Normalized current Voltage (mV) -100 pNO3/pCl = 14 * Permeability to nitrate is 10-fold greater than GABAA-mediated chloride conductance (pNO3/pCl = 2.5)

  11. Currents are larger using the gramicidin perforated patch technique

  12. Is the current large enough to change excitability of the DA neurons?

  13. DA stimulates firing by a D2-independent mechanism * Shi, et al. (2000) Dual effects of amphetamine on dopamine neurons mediated by dopamine and nondopamine receptors. J Neurosci 20(9):3504-3511.

  14. Low DA concentrations stimulate firing via a DAT-dependent mechanism

  15. Muscimol - control DA - control Vrest Vrest Normalized current Normalized current Voltage (mV) Voltage (mV) GABAA-mediated chloride conductance is inhibitory * Gramicidin perforated-patch recordings DAT-mediated chloride conductance is excitatory

  16. Is the DAT-mediated excitation sufficient to stimulate neurotransmitter release?

  17. DA neurons make glutamatergic autapses in culture

  18. Amphetamine increases the AMPA-mediated EPSC

  19. 32 ± 8%

  20. Conclusions • DAT mediates an uncoupled chloride conductance. • DAT currents in neurons are activated at much lower DA concentrations than the D2-receptor or transport. • The DAT-mediated chloride conductance excites DA neurons. • Activation of DAT-mediated currents is a potential • mechanism for stimulating DA release in the SN.

  21. Susan Amara Krishna Prasad Spencer Watts Mark Sonders Kevin Poth Ole Mortensen Chialin Cheng Bill Freeman Andreia Fontana Jonathan Morris Alessandra Cecchini Wendy Fairman Barb Leighton Mary Chalmers Debra Peth Jean Yang Bozena Kutyba Nikki Lind

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