Uptake and Release of Neurotransmitters
- Abstract
- Table of Contents
- Materials
- Figures
- Literature Cited
Abstract
The availability of clonal cell lines for norepinephrine, dopamine, and serotonin transporters allows the characterization of drug interactions with transporter recognition sites using radioligands, as well as the characterization of drug effects on selective transporter?mediated uptake and release of substrate. In addition to clonal cell lines, synaptosomes prepared from specific brain regions can be used to conduct these studies without interference by endogenous transporters or binding proteins that are present in other tissues. This unit presents protocols for uptake and release of tritiated substrates using intact cells (either detached or in suspension) or synaptosomes. An HPLC procedure for electrochemical detection of nonradiolabeled substrates is also provided. Time?dependent release can also be measured in assays involving real?time sampling.
GO TO THE FULL PROTOCOL: PDF or HTML at Wiley Online Library Table of Contents
- Basic Protocol 1: Study of Uptake and Release of Dopamine in Intact Attached Cells Expressing the Recombinant Dopamine Receptor
- Alternate Protocol 1: Study of Dopamine Uptake in Detached Cells
- Basic Protocol 2: Study of Dopamine Uptake in Synaptosomes
- Basic Protocol 3: Study of Dopamine Release from Synaptosomes
- Alternate Protocol 2: Detection of Uptake or Release of Dopamine by HPLC with Electrochemical Detection (HPLC‐EC)
- Alternate Protocol 3: Using a Superfusion Apparatus for Time Sampling
- Basic Protocol 4: Examination of the Dopamine Transporter with Radioligands
- Support Protocol 1: Establishing Initial Binding‐Assay Parameters
- Reagents and Solutions
- Commentary
- Literature Cited
- Figures
- Tables
GO TO THE FULL PROTOCOL: PDF or HTML at Wiley Online Library Materials
Basic Protocol 1: Study of Uptake and Release of Dopamine in Intact Attached Cells Expressing the Recombinant Dopamine Receptor Materials
|
GO TO THE FULL PROTOCOL: PDF or HTML at Wiley Online Library Figures
-
Figure 7.9.1 Uptake of [3 H]dopamine by C6‐hDAT cells. -
Figure 7.9.2 Substrate‐induced release of [3 H]dopamine from COS‐7 cells transfected with cDNA for dopamine transporter. -
Figure 7.9.3 Association (A ) and dissociation (B ) of [125 I]RTI‐55 binding to membranes from COS‐7 cells transiently expressing the dopamine transporter. B e is the amount bound at equilibrium; B t is the amount bound at time t. -
Figure 7.9.4 Binding data analyzed using GraphPad Prism (also see UNIT ).
Videos
Literature Cited
Literature Cited | |
Axelrod, J., Weil‐Malherbe, H., and Tomchick, R. 1959. The physiological disposition of [3H]epinephrine and its metabolite metanephrine. J. Pharmacol. Exp. Ther. 127:251‐256. | |
Berger, P., Elsworth, J., Reith, M., Tanen, D., and Roth, R. 1990. Complex interaction of cocaine with the dopamine uptake carrier. Eur. J. Pharmacol. 176:251‐252. | |
Blakely, R.D., Berson, H., Fremeau, R.T., Caron, M., Peek, M.M., Prince, H.K., and Bradley, C.C. 1991. Cloning and expression of a functional serotonin transporter from rat brain. Nature 354:66‐70. | |
Buck, K.J. and Amara, S.G. 1995. Structural domains of catecholamine transporter chimeras involved in selective inhibition by antidepressants and psychomotor stimulants. Mol. Pharmacol. 48:1030‐1037. | |
Chen, N.‐H. and Reith, M.E.A. 1993. [3H]Dopamine and [3H]serotonin release in vitro induced by electrical stimulation in A9 and A10 dopamine regions of rat brain: Characterization and responsiveness to co |