ATP-regulated neuronal catecholamine uptake: A new mechanism
Abstract
Uptake of the catecholamines (CA), dopamine (DA) and norepinephrine (NE) into synaptosomes prepared from rat and bovine brains was potentiated by ATP (from 0.1 to 5.0 mM) in a dose-dependent manner. Other nucleotides, particularly the nonhydrolyzable ATP analogs β, γ-imidoadenosine-5′-triphosphate (AMP-PNP) and β, γ-methyladenosine-5′-triphosphate (AMP-PCP) also potentiated [3H]DA and [3H]NE uptake. Several endogenous 5′-nucleotide triphosphates (e.g. GTP, UTP and CTP) potentiated [3H]CA uptake, but were less effective than ATP. Among the ATP metabolites, only ADP potentiated uptake whereas AMP and adenosine did not.
[3H] Dopamine uptake measured in Krebs bicarbonate buffer had a Km of 2.1 μM and a Vmax of 163.9 pmol/mg prot./min. In presence of ATP, [3H]DA uptake had much higher affinity (Km = 0.56 μM) and larger capacity (Vmax = 333 pmol/mg prot./min) than uptake in absence of added ATP. Furthermore, [3H]DA uptake in presence of ATP had faster rate of uptake, and was independent of temperature while in absence of added ATP it was temperature-dependent. This ATP-dependent [3H]DA uptake was retained by synaptosomal ghosts that were obtained after lysing the striatal synaptosomes and removing their contents of synaptic vesicles and mitochondria.
It is proposed that, in addition to the carrier-mediated (neuronal) uptake of CA, there is neuronal uptake that is regulated by ATP and inhibited by cocaine, which may be more relevant for terminating the synaptic action of CA because of its faster rate of uptake and larger capacity.
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Cation requirements of basal and ATP-regulated dopamine transport in rat pheochromocytoma cells
1996, NeuroscienceThe transport of dopamine into presynaptic nerve terminals is the primary mechanism for the termination of dopaminergic neurotransmission. This transport process has recently been found to be composed of two components, a basal dopamine transport pathway which exists in the absence of extracellular ATP and an ATP-regulated moiety which comprises approximately 66% of the total transport system [ Cao C.J. et al. (1990) Biochem. Pharmac.39, R9–R14; Cao C.J. et al. (1989) Biochemistry8, 207–220; Dunigan C.D. and Shamoo A.E. (1995) Neuroscience65, 1–4; Eshleman A. et al. (1995) Life Sci.56, 1613–1621]. Using a rat pheochromocytoma cell line and a Krebs bicarbonate buffering system, the present study examined the effect of several cations on both basal and ATP-regulated dopamine transport. In the absence of extracellular ATP, dopamine transport had an absolute dependence on the presence of Na+, but exhibited no requirement for Mg2+. Kinetically, the addition of 120 mM NaCl increased the Vmax of basal dopamine transport by approximately 150%. In contrast, the ATP-regulated dopamine transport pathway displayed a different sensitivity to Na+ and was completely dependent upon the presence of Mg2+. The addition of 1.2 mM MgSO4 increased the Vmax of transport in the presence of 0.7 mM extracellular ATP by 222%. Both basal and ATP-regulated transport were unaffected by the removal of either Ca2+ or K+ from the assay buffer. When the effects of ouabain, a potent inhibitor of Na+, K+-ATPase, were tested in the rat pheochromocytoma cell model, it was found that concentrations of ouabain as high as 1 mM were ineffective at inhibiting either the basal or ATP-regulated dopamine transport components. These results imply that the Na+ gradient supplied by Na+, K+-ATPase is not the sole provider of energy needed to drive either transport process.
The ionic requirements of the basal and ATP-regulated dopamine transport pathways demonstrate the distinction between the two transport processes. In addition, the ionic dependency profile of the ATP-regulated moiety has provided some mechanistic insights into ATP-regulated catecholamine uptake, as the absolute Mg2+ requirement and the ineffectiveness of Ca2+ argues against the involvement of either purinergic receptors or a Ca2+-dependent, Mg2+-independent ectokinase in the ATP-regulated transport system. Copyright © 1996 IBRO. Published by Elsevier Science Ltd.
ATP enhances catecholamine uptake into PC12 cells
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Li<sup>+</sup> stimulates ATP-regulated dopamine uptake in PC12 cells
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The neuronal transport mechanism for noradrenaline is not influenced by the ATP analogue ATPγS
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