Vascular Reactivity by Purinoceptor Activation in Rat Inferior Vena Cava

BACKGROUND: Extracellular ATP, released from platelets and nerve endings, plays significant roles in the regulation of circulation. The effects of ATP depend on the location of the vessels and the species of experimental animals. Until now, studies were limited to arteries, so we compared the effects of ATP in rat vena cava with those in the aorta and attempted to identify the characteristics of their receptors. METHODS: Vascular rings were isolated from the rat inferior vena cava and descending thoracic aorta. Endothelial cells were preserved or removed by gentle rubbing. The isometric contractions were recorded on polygraph using a force transducer. RESULTS: In the vena cava ring precontracted by 100 nM norepinephrine (NE), ATP elicited relaxations in a dose-dependent manner. These effects were abolished by removal of the endothelium or pretreatment with a nitric oxide synthase inhibitor. Relaxations to ATP in the vena cava (EC50 :9.9 microM) were less potent than those in the aorta (1.7 microM). The relative order of potencies was ADP>ATP>AMP>adenosine, but the maximal relaxation to ADP was smaller than to ATP. ATP-induced vasorelaxation was blocked by suramin, a nonselective antagonist for P2 purinoceptor and reactive blue-2, a P2Y blocker. At basal tension, ATP contracted the vena cava dose-dependently and these effects were potentiated by endothelium-removal. Contractions in the vena cava were also less potent than in the aorta, and the order of potencies was alpha, beta-MeATP>UTP>ATP>ADP>AMP=adenosine. ATP-induced vasoconstriction was blocked by suramin and alpha, beta-MeATP, a desensitizing antagonist of P2X purinoceptor, and potentiated by pretreatment with UTP. CONCLUSION: These results suggest that ADP and ATP acts on P2Y1- and P2Y2-purinoceptor in the endothelium, respectively and induces vasorelaxation of the vena cava, which is mediated by nitric oxide. Since ATP and UTP induced vasoconstriction in endothelium-denuded condition, it may be mediated by the activation of the P2X and P2Y4, 6 purinoceptor on smooth muscles, respectively.

Effects of protein kinase C modulation on hepatic hemodynamics and glucoregulation

This study evaluated the effects of PKC activation using phorbol 12-myristate 13-acetate (PMA) and PKC inhibition using the isoquinoline sulfomide derivative H-7 on hemodynamics and glucoregulation in the isolated perfused rat liver. Livers were isolated from fed male Holtzman rats and perfused with Krebs Ringer bicarbonate solution under a constant flow of 50 ml/min at 35degreeC. Portal vein pressure, glucose and lactate concentrations in the medium and oxygen consumption rates were continuously monitored by a Grass polygraph, YSI glucose and lactate monitors, and a YSI oxygen monitor, respectively. PMA at concentration of 2 to 200 nM increased the portal vein pressure, glucose and lactate production, but decreased oxygen consumption rate in a dose-dependent fashion. H-7 (200 micrometer) attenuated PMA (50 nM)-induced vasoconstriction (15.1+/-1.36 vs 10.56+/-1.17 mmHg), glucose production rate (91.3+/-6.15 vs 71.8+/-2.50 micromoles/g/hr), lactate production rate (72.4+/-6.82 vs 53.6+/-4.82 micromoles/g/hr) and oxygen consumption rate (33.7+/-1.41 vs 27.9+/-1.75 microliter/g/min). The effects of PMA were blocked either by addition of verapamil (9 micrometer) or perfusion with Ca2+-free KRB. These results suggest that the hemodynamic and glucoregulatory changes in the perfused rat liver are mediated by protein kinase C activation and require Ca2+ influx from the extracellular fluid.

Calcium Antagonist-like Action of Antidepressants Including Sertraline in PCl2 Cells / 신경정신의학

It has been known that antidepressants have calcium antagonist-like action in neuronal tissues. However, their mechanisms are still obscure. For the study of neurochemical machanism of antidepressants, the authors examined the effects of antidepressants(1-100 microM ) on the intracellular Ca2+ concentration ([Ca2+]i) and the membrane potential in PCl2 cells using fluorescent dyes, fura-2/AM and bisoxonol, respectively. The results were as follows : 1) Sertraline, a selective serotonin reuptake inhibitor (SSRI), inhibited the increment of [Ca2+]i induced by high 60 mM KCI and 100 microM ATP with an IC50 value of 2.5 microM and 5.4 microM, respectively. 2) SSRIs(sertraline, paroxetine and fluoxetine) and tricyclic antidepressants(imipramine and amitriptyline) had strong effects on the inhibition of both voltage-dependent Ca2+ channel and receptor-dependent Ca2+ channel, whereas atypical antidepressant(trazodone) and MAO inhibitor(moclobemide) had lisle effects. 3) Sertraline itself depolarized the membrane potential in a sustained manner depending on its own concentration and it also increased the basal level of [Ca2+]i. 4) The increment of [Ca2+]i might be induced partly by the release from the intracellular calcium store, but mostly induced by the calcium transport through membrane. 5) Among those antidepressants tested, sertraline was the most potent one. Other SSRIs(paroxetine and fluoxetine) and tricyclic antidepressants(imipramine and amitriptyline) were moderately potent. Atypical antidepressant(trazodone) had little effects, and MAO inhibitor (moclobemide) had no effect on the depolarization. 6) External application of ATP induced temporary depolarization. This effect was blocked by prior treatment with sertraline with an IC50 value of 30 microM. 7) The increment of [Ca2+]i through voltage-dependent Ca2+ channel was almost inhibited by a selective calcium channel blocker(nimodipine). However, the ATP-induced increment of [Ca2+]i was partially inhibited by nimodipine. These inhibitory effects were potentiated by the addition of sertraline. In the light of these results, it is likely that SSRIs and tricyclic antidepressants could show the blocking effects on both voltage-dependent and receptor-dependent calcium channel by depolarizing neuronal cell membrane potential in a sustained manner and by increasing intracellular free calcium level.

Apoptosis induced by adenosine 5'-trisphosphate in mouse leukemic cells

Extracellular ATP elicits various biological responses and plays a significant role in physiological regulation. Recently, ATP-induced growth inhibitions were reported in some tumor cell lines, but these effects and mechanisms are not well known. This study was conducted to investigate ATP-induced growth inhibition in mouse leukemic (P388D,) cells. ATP inhibited cell growth in a dose-dependent manner as analyzed by MTS assay (IC50: 33.1 muM). Nucleotides other than ATP, such as ADP (37.5 muM) and AMP (33.2 muM) had the same effects as ATP, but adenosine (57.8 muM) showed less effect than ATP. ATP attenuated the cells in G0/G1 and G2/M phases, but increased those in S phase in flow cytometric analysis. Hypodiploid cells (A0), the presumptive findings of apoptosis, were found among the ATP-treated cells. ATP induced DNA fragmentation into 180 ~ 200 bps as measured by electrophoresis. Some apoptotic cells were stained by TUNEL method. ATP increased the intracellular free Ca++ concentration ((Ca++)i) and the increment of (Ca++)i was caused by influx from the extracellular space. These results suggest that extracellular ATP induces growth inhibition through apoptosis.

Hemodynamic characteristics of extracellular UTP in the perfused rat liver

Uridine 5'-triphosphate (UTP) is stored in the granules of cells such as platelets and is released into the extracellular space upon cell stimulation. Extracellular UTP is known to influence many biological processes. We investigated the hemodynamic effects of UTP on the perfused rat liver and characterized its receptors. Liver perfusions were performed in a recirculation system under constant pressure (28 cmH2O). The perfusion flow and oxygen consumption rate were measured at 30 second intervals. UTP decreased the perfusion flow and the oxygen consumption rate, dose-dependently. UTP-induced changes were transient and disappeared in about 10 minutes. Suramin (P2-purinergic antagonist, 100 uM) and indomethacin (cyclooxygenase inhibitor, 20 uM) blocked UTP-induced hemodynamic changes significantly. The effects of UTP were also inhibited when Kupffer cells were damaged with treatment of gadolinium chloride (10 mg/kg iv). L-NAME (1 mM), a potent inhibitor of nitric oxide synthase, markedly enhanced and prolonged the contractile response of UTP in the hepatic vessel. These results suggest that UTP acts mainly on suramin-sensitive UTP receptors on the Kupffer cell through prostanoid synthesis. The nitric oxide systems in the endothelium seem to counteract the vasoconstrictile action of UTP in the hepatic circulation.