Release of amines from acidified stores following accumulation by Transport-P.
1. Transport-P is an uptake process for amines in peptidergic neurones of the hypothalamus. It differs from other uptake processes by its anatomical location in post-synaptic neurones, its functional properties and by the structure of its ligands. Transport-P accumulates amines in intracellular vesicles, derives its energy from the electrochemical proton gradient and is linked to vacuolar-type ATPase (V-ATPase). Transport-P is blocked by antidepressants. We have now studied the release of amines following uptake by Transport-P in a cell line of hypothalamic peptidergic neurones. 2. Release of prazosin was not inhibited by the antidepressant desipramine; as Transport-P is blocked by desipramine, this indicated that amines are released by a mechanism which is independent of Transport-P. 3. Release of prazosin was sensitive to temperature and conformed to the Arrhenius equation. Release was minimal in the range 0-25 degrees C but accelerated exponentially at higher temperatures up to 33 degrees C. The activation energy for the release of prazosin is 83.1 kJ x mol(-1), corresponding to a temperature quotient (Q10) value of 3. 4. Release was accelerated by the organic base chloroquine, the ionophore monensin, bafilomycinA1 which inhibits V-ATPase and by increasing extracellular pH. Thus, retention of prazosin requires an intracellular proton gradient which is generated by V-ATPase. 5. Fluorescence microscopy demonstrated that release of BODIPY FL prazosin was temperature dependent and was accelerated by chloroquine and monensin. 6. Thus, following uptake by Transport-P, amines are accumulated in acidified intracellular stores. Their retention in peptidergic neurones requires intracellular acidity. The amines are released by a temperature-dependent process which is resistant to antidepressants.
Al-Damluji
S
S
Division of Endocrinology, Royal Free and University College Medical School, University College London, London NW3 2PF. damluji@rfhsm.ac.uk
Shen
W B
WB
D000928:Antidepressive Agents / Q000494:pharmacology; D001692:Biological Transport; D001896:Boron Compounds / Q000378:metabolism; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology; D003891:Desipramine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006863:Hydrogen-Ion Concentration; D007031:Hypothalamus / Q000378:metabolism*; D008985:Monensin / Q000494:pharmacology; D011224:Prazosin / Q000378:metabolism*; D013696:Temperature
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D000928:Antidepressive Agents; D001896:Boron Compounds; D002738:Chloroquine; D008985:Monensin; D003891:Desipramine; D011224:Prazosin