Reaktion #11846

Reaktionsgleichung

[3H]Dopamine

nicotine

→

Dopamine

Reaktionsbedingungen

Aufarbeitung

1. 1
   workup.WAITthe suspension was incubated at 37° C. for another 10 minutes
2. 2
   workup.WAITPerfusion buffer (room temperature) was pumped into the chambers at a rate of 3 ml/min for a wash period of 8 minutes
3. 3
   SonstigeFractions (12 seconds each) were continuously collected from each chamber throughout the experiment
4. 4
   SonstigeThe perfusate was collected directly into scintillation vials, to which scintillation fluid
5. 5
   workup.ADDITIONwas added

Vorschrift

The synaptosomal suspension was incubated for 10 minutes at 37° C. to restore metabolic activity. [3H]Dopamine ([3H]DA, specific activity=28.0 Ci/mmol, NEN Research Products) was added at a final concentration of 0.1 μM and the suspension was incubated at 37° C. for another 10 minutes. 50 μL aliquots of tissue +100 μL perfusion buffer were loaded into the suprafusion chambers of a Brandel Suprafusion System (series 2500, Gaithersburg, Md.). Perfusion buffer (room temperature) was pumped into the chambers at a rate of 3 ml/min for a wash period of 8 minutes. Test compound (10 μM) or nicotine (10 μM) was then applied in the perfusion stream for 40 seconds. Fractions (12 seconds each) were continuously collected from each chamber throughout the experiment to capture basal release, agonist-induced peak release and to re-establish the baseline after the agonist application. The perfusate was collected directly into scintillation vials, to which scintillation fluid was added. [3H]DA released was quantified by scintillation counting. For each chamber, the integrated area of the peak was normalized to its baseline.

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