Characterization and possible function of adenosine 5'-triphosphate receptors in activated rat microglia.

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1. Purinoceptor agonist-induced currents in untreated (proliferating) and lipopolysaccharide (LPS; 100 ng ml-1)-treated (non-proliferating) rat microglial cells in culture were recorded by the whole-cell patch-clamp technique. These cells have two preferred resting membrane potentials, one at -35 mV and another one at -70 mV. 2. Most experiments were carried out in non-proliferating cells. ATP, ATP-gamma-S and alpha,beta-MeATP (1-1000 microM in all cases) evoked an inward current at a holding potential of -70 mV, followed, in some experiments, by an outward current. At -70 mV 2-methylthio ATP (1-1000 microM) evoked an inward current, whereas at -35 mV it produced an outward current only. 3. When K+ was replaced in the pipette solution by an equimolar concentration of Cs+ (150 mM), the main outward component of the ATP-gamma-S (10 microM) induced response disappeared. Instead, an inward current was obtained. Replacement of K+ by Cs+ did not affect the inward current evoked by 2-methylthio ATP (300 microM). 4-Aminopyridine (1-10 mM), however, almost abolished this current and unmasked a smaller outward current. 4. The rank order of agonist potency was 2-methylthio ATP > ATP > alpha,beta-MeATP. Adenosine and UTP were inactive. Suramin (300 microM) and reactive blue 2 (50 microM) antagonized the effect of 2-methylthio ATP (300 microM). 5. I-V relations were determined by delivering fast voltage ramps before and during the application of 2-methylthio ATP (300 microM). In the presence of extra- (1 mM) and intracellular (150 mM) Cs+, the 2-methylthio ATP-evoked current crossed the zero current level near 0 mV. When both Cs+ (1 mm) and 4-aminopyridine (1 mM) were present in the bath medium, the intersection of the 2-methylthio ATP current with the zero current level was near - 75 mV.6. 2-Methylthio ATP (1-1I000 MicroM) induced the same inward current both in proliferating and nonproliferating microglia. However, the depolarizing response to 2-methylthio ATP (300 MicroM) was larger and longer-lasting in the proliferating cells. When the free Ca2+ concentration in the pipettes was increased from the standard 0.01 to 1 MicroM, the amplitude and duration of this depolarization was increased in non-proliferating cells. 4-Aminopyridine (1 mM) enhanced the duration, but not the amplitude of responses.7. ATP and its structural analogues stimulate microglial purinoceptors of the P2Y-type. This leads to the opening of non-selective cationic channels and potassium channels. Depending on the resting membrane potential, depolarization or hyperpolarization prevails. Although the inward current produced by 2-methylthio ATP is of similar amplitude in proliferating and non-proliferating microglia, the resulting depolarization is smaller in the latter cell type because of the presence of voltage-sensitive, outwardly rectifying potassium channels.