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Am J Physiol Heart Circ Physiol 294: H1716-H1723, 2008. First published February 8, 2008; doi:10.1152/ajpheart.00945.2007
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A positive inotropic effect of ATP in the human cardiac atrium

Ulrich Gergs,1 Peter Boknik,2 Wilhelm Schmitz,2 Andreas Simm,3 Rolf-Edgar Silber,3 and Joachim Neumann1

1Institut für Pharmakologie und Toxikologie and 2Institut für Pharmakologie und Toxikologie, Universitätsklinikum, Westfälische Wilhelms-Universität, Münster; and 3Klinik für Herz- und Thoraxchirugie, Medizinische Fakultät, Martin-Luther-Universität Halle-Wittenberg, Halle, Germany

Submitted 15 August 2007 ; accepted in final form 4 February 2008


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
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We studied contractile effects in isolated electrically driven (1 Hz) atrial preparations from patients undergoing cardiac bypass surgery. ATP concentration dependently (10, 30, and 100 µM) and rapidly decreased force of contraction (negative inotropic effect, NIE) and thereafter more slowly increased force of contraction. The maximum positive inotropic effect (PIE) at 100 µM ATP amounted to 152% of the predrug value (n = 9) and was stable and could be washed out fast and completely. The PIE did not affect time parameters of contraction (time to peak tension and time of relaxation). Moreover, a similar NIE and PIE were noted with adenosine 5'-O-(2-thiotriphosphate) (100 µM). In contrast 2-methyl-thio-ATP did not exert a NIE but only a PIE. In a second set of experiments, preparations were first incubated for 30 min with purinoreceptor antagonists and, in their continuous presence, 100 µM ATP was applied. However, the PIE and NIE of ATP could neither be blocked with suramin (100 and 500 µM), pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (50 µM), nor reactive blue 2 (30, 100, and 500 µM), which are known blockers for subtypes of P2 receptors, or 1,3-dipropyl-cyclopentvl-xanthine (1 and 10 µM), a subtype (A1 adenosine) P1 receptor blocker. Likewise, the inhibitor of phospholipase C (PLC) activity (U-73122) and the inhibitor of adenylate cyclase activity (SQ-022563) (10 µM each) failed to affect the NIE and the PIE of ATP. We tentatively suggest that the PIE of ATP might be mediated via P2X4-like receptors. In summary, we describe a novel biphasic effect of ATP on force contraction in the isolated human atrium. It is conceivable that ATP plays a physiological role in the human heart, for instance, after cardiac injury to sustain contractility.

human heart; right atrium; adenosine 5'-triphosphate; purinoceptor; inotropy

SOON AFTER DISCOVERY of ATP, Drury and Szent-Györgyi reported on cardiac effects of the compound: a negative chronotropic effect and a decrease in blood pressure (19). Berne suggested that the vasodilatory effect of ATP may be mediated by its degradation product adenosine (5). This led to extensive studies on the effects of adenosine in the cardiovascular system. Only later on was interest in ATP itself renewed. This was in part driven by the suggestion that ATP may be a cotransmitter in nonadrenergic and noncholinergic nerves (13). In fact, ATP can be released from nerve terminals as a cotransmitter with norepinephrine and acetylcholine (14).

Extracellular ATP was found to exert many effects in the cardiovascular system. These include negative (NIE) and positive (PIE) inotropic effects (species dependent), negative chronotropic and dromotropic as well as antihypertrophic effects (for review, see Ref. 62). A number of additional physiological effects of ATP in the heart have been observed, like inhibition of glucose transport (22), involvement in preconditioning, and release (47, 68).

Large amounts of ATP can be released from ischemic cardiomyocytes (5) and, e.g., from activated platelets (18). Levels of ATP in the coronary system are usually quite low (e.g., 1 nM; see Ref. 10) because ATP is rapidly degraded to ADP, AMP, and adenosine by soluble and membrane-bound ATPases (33, 65). In the interstitium of the heart, however, higher levels of ATP (~40 nM) can be measured (39).

ATP can act via P1 and P2 receptors (15). The latter receptors are further divided in P2X and P2Y receptors (24). In rat atrium (25), an initial NIE of ATP followed by a PIE was reported. In rat and guinea pig atrium, the PIE of ATP was antagonized by suramin or reactive blue 2 (RB2), respectively (25, 43). The NIE of ATP in the rat and in the guinea pig atrium was P1 adenosine receptor mediated because it was sensitive to 1,3-dipropyl-cyclopentvl-xanthine (DPCPX), a selective inhibitor of the A1 receptor subtype of the P1 receptor (25, 43). In the rat, 2-methyl-thio-ATP only exerted a NIE (25) while in chicken cardiomyocytes or mouse cardiomyocytes it increased contractility (44, 50). In the ventricle of the rat, no NIE but only a small PIE of ATP is consistently noticed (41, 54), which is accompanied by an enhanced current through L-type-calcium channels and a resultant increase in free intracellular Ca2+ that augments contractility (29, 54). ATP does not sensitize the contractile proteins against Ca2+ (21) but can alkalinize the cytosol, which may contribute to its PIE (51). Clinically, ATP is used to treat supraventricular arrhythmias, especially in children. Because ATP is rapidly degraded to adenosine in serum, ATP blocks the arteriovenous node of patients because the resultant adenosine activates A1 adenosine receptors (and leads to vagal activation) and this does not involve P2 receptors (53). Inotropic effects of ATP in the human heart (atrium or ventricle) have hitherto not been reported. In the present work, we tested the hypothesis that ATP affects contractility in the human atrium like it does in the rat atrium, i.e., a DPCPX-sensitive NIE followed by a suramin-sensitive PIE. However, although we observed a biphasic response, the antagonists that worked in the rat atrium were ineffective in the human heart, indicative of mediation by different receptors or subtypes. Part of this work has been presented in abstract form (26).


   MATERIALS AND METHODS
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 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
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Contraction experiments were performed as described previously (61, 69). In brief, trabeculae of right atrium from patients with coronary heart disease undergoing cardiac surgery (diameter, <1 mm; length, 5–8 mm) were dissected in gassed bathing solution (see below) at 4°C. The age of the patients ranged from 61 to 81 yr (mean, 71 yr). All patients were in New York Heart Association class II-III. Medical treatment consisted of angiotensin-converting enzyme inhibitors, β-adrenoceptor blockers, diuretics, antilipemic agents, cholinergic agents, antithyroid agents, hypoglycemic agents, gout suppressants, antibiotics, {alpha}-adrenoceptor blockers, parasympatholytics, calcium channel blockers, proton pump inhibitors, and analgetics (see Table 1). These studies were approved by the local ethical committee (hm-bü 04.08.2005), and patients gave informed consent. In some experiments, mouse left atrial preparations (from 3-mo-old mice) were used under the same experimental conditions as the human atrial preparations. Animals were handled and maintained according to approved protocols of the animal welfare committee of the University of Halle-Wittenberg (Halle, Germany). Details on these animal experiments have been described (38).


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  		Table 1. Clinical data of patients involved in the present study

 
The bathing solution contained (in mM) 119.8 NaCl, 5.4 KCl, 1.8 CaCl2 1.05 MgCl2, 0.42 NaH2PO4, 22.6 NaHCO3, 0.05 Na2EDTA, 0.28 ascorbic acid, and 5.05 glucose continuously gassed with 95% O2-5% CO2 and maintained at 37°C and pH 7.4 as described (8). Preparations were attached to a bipolar stimulating electrode and suspended individually in 10-ml glass tissue chambers for recording isometric contractions. Force of contraction was measured with inductive force transducers connected to a chart recorder. Time parameters of single contractions were evaluated at high chart speed. Each muscle was stretched to the length of maximal force of contraction. Trabeculae carneae were electrically stimulated at 1 Hz with rectangular pulses of 5-ms duration, and the voltage was ~10–20% greater than threshold. All preparations equilibrated in bathing solution until complete stabilization (≥30 min). During this period, the bathing solution was changed every 15 min. In single concentration experiments, the tested drugs were applied for 10 min. In experiments with antagonists/inhibitor, preparations were preincubated with the antagonist/inhibitor for 30 min, and then ATP was added in the continuous presence of the antagonist/inhibitor. In some experiments, mouse left atria were incubated with 100 µM UTP in the continuous presence of inhibitors.

Chemicals. ATP [no agonist at P2X6, antagonist at P2Y1,4 (1)], 2-methyl-thio-ATP [agonist at P2X1–7 (36); agonist at P2Y1,11,12, poor or no agonist at P2Y6 (1)], adenosine 5'-O-(2-thiotriphosphate) (ATP{gamma}S; good agonist at P2X1–6, P2Y1,2,11 poor or no agonist at P2X7, P2Y4,6), DPCPX [A1 adenosine receptor antagonist (42)], pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid [PPADS, potent inhibitor at P2Y1, poor inhibitor at P2Y4,6,13 no inhibition at P2Y2,11,12 (63), potent inhibitor at P2X1,2,3,5 poor or no inhibition at P2X4,6,7 (36)], RB2 [potent inhibitor at P2Y1,5,11,12,13, poor at P2Y2 (63)], suramin [potent inhibitor at P2Y1,2,11,12,13, poor inhibitor at P2Y6, no inhibition of P2Y4 (63), potent inhibitor at P2X1,2,3,5, poor or no inhibition at P2X4,6,7 (36), inhibits many additional proteins; see Ref. 40], SQ-22563 [adenylate cyclase inhibitor (66)], U-73122 [phospholipase C (PLC) inhibitor (66)], and isoproterenol hydrochloride were obtained from Sigma (München, Germany). All other chemicals were of analytic grade or best grade commercially available.

Data analysis. The experimental data given in text and Figs. 1–4 are means ± SE. Force of contraction is given in percentage of the predrug values where indicated. When both negative and PIEs were present, the latter was expressed as the difference between the minimal value of contractility (during the negative effect) and the maximum positive effect divided by the basal force of contraction (value before negative effect). In this way, both the negative and the positive effects were expressed as percentages of the same control value [see for previous use of this calculation of the effects of ATP (25)]. Data of multiple groups were compared using one-way ANOVA for paired observations followed by Bonferroni's t-test. A P value <0.05 was regarded as significant.


Figure 1
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  		Fig. 1. A: concentration-dependent inotropic effects of ATP. Typical original recording. Force of contraction in isolated electrically driven (1 Hz) atrial preparations from human hearts. At the indicated time, ATP was applied. Note the initial negative inotropic effect (NIE) followed by a positive inotropic effect (PIE). Time scale is indicated on the abscissa. Force of contraction is indicated at the ordinate. B: schematic drawing. It is indicated how the NIE and the PIE of ATP were quantified. C: quantification of the NIE and the PIE of ATP. Control is before drug applications (100%). NIE and PIE are given as mean values in %control ± SE of n = 8 experiments. *P < 0.05 vs. control. D: time parameters in isolated electrically driven (1 Hz) atrial preparations from human hearts. ATP (100 µM) was applied. Time to peak tension (Tr) and time of relaxation (Tf) are given on the ordinate as mean values ± SE. Nos. indicate no. of experiments.

 

Figure 4
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  		Fig. 4. Quantification of the NIE and the PIE of ATP (100 µM) in several experiments like those in Fig. 3. Control is before drug applications. NIE and PIE are given as mean values in %control ± SE. Nos. indicate no. of experiments. Before additional application of ATP, preparations were preincubated for 30 min with the antagonists DPCPX (1 µM), suramin (500 µM), PPADS (50 µM), reactive blue 2 (RB2, 500 µM), SQ-22536 (10 µM), or U-73122 (10 µM). *P < 0.05 vs. control.

 

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As depicted in the original recording in Fig. 1, ATP concentration-dependently exerted an initial NIE that rapidly (within <1 min) terminated and was followed by a pronounced sustained PIE. This sequence of events was very reproducible and occurred in tissue samples from all patients investigated. The effect was discernable at 10 µM ATP, the lowest concentration used. The effect could be rapidly and completely washed out and thereafter could be elicited again. At 100 µM ATP (the highest concentration investigated), the effect was most pronounced and was stable up to 30 min (the longest time studied); this argues against a rapid desensitization of the receptor involved. ATP (100 µM) was used in all subsequent experiments. The scheme in Fig. 1B depicts how we defined PIEs and NIEs of ATP (for quantification in Fig. 1C). With the use of the scheme in Fig. 1B, the effects of 100 µM ATP were quantified (Fig. 1C). Next, we quantified the time to peak tension and time of relaxation. Using β-adrenergic (by 1 µM isoproterenol) stimulation under the same experimental conditions, we noted a shortening of time of relaxation in accordance with the literature (from 123 ± 15 to 87 ± 4 ms, n = 6, P < 0.05). In contrast, ATP did not affect time parameters of contraction (Fig. 1D). It could be argued that the inotropic effects are mediated not by ATP per se but via its degradation products. Thus we studied a derivative of ATP, ATP{gamma}S, that is more slowly hydrolyzed than the parent compound ATP. This compound was indeed still active and induced both a negative (to 68%) and a positive inotropic (to 190%) effect like ATP itself (n = 4, P < 0.05; Fig. 2A). This is tentatively regarded as evidence that ATP itself acts on the preparations to exert its inotropic effect and not a degradation product of ATP. We further noted that 100 µM 2-methyl-thio-ATP exerted solely a positive inotropic under these experimental conditions (Fig. 2B). The effects were already detectable at 10 µM 2-methyl-thio-ATP. 2-Methyl-thio-ATP at 100 µM (the highest concentration tested) increased force of contraction from 5.42 ± 0.94 to 8.52 ± 1.24 mN or by ~57% (n = 6, P < 0.05), without the remotest NIE (Fig. 2B). Next, we wanted to find out which subtype of receptor mediates the PIE and/or the NIE of ATP. To this end, a number of commercially available antagonists for P1, P2X, and/or P2Y receptors were investigated. However, the PIE and NIE of ATP could neither be blocked by DPCPX [a P1 receptor antagonist more specifically, an A1 adenosine receptor antagonist; Fig. 3A (4, 60)] nor by suramin (a P2 antagonist; Fig. 3B), PPADS (a P2 antagonist; Fig. 3C), or RB2 (here 500 µM, a P2 antagonist; Fig. 3D). Data for these antagonists are quantified in Fig. 4 using the calculations exemplified in Fig. 1B. Apparently, the NIE of ATP is smaller in the presence of 1 µM DPCPX than in the presence of suramin, PPADS, and RB2. We do not think this implies an effect of DPCPX because the NIE was still present when we used 10 µM DPCPX (data not shown). This is in contrast to data by Froldi et al. (25) or Mantelli and coworkers (43) that 10 or 100 nM DPCPX completely blocked the NIE of ATP in rat or guinea pig atrium. DPCPX, suramin, and PPADS alone during 30 min incubation did not alter basal force of contraction (data not shown). In human cardiac tissue, the KD value for DPCPX at A1 adenosine receptors was reported as 2 nM (9). Hence, the concentrations of DPCPX we used should have blocked this receptor. Indeed we have used similar concentrations of DPCPX before in human atrial tissue (60, 67).


Figure 2
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  		Fig. 2. Typical original recording. Force of contraction in isolated electrically driven (1 Hz) atrial preparations from human hearts. At the indicated time, adenosine 5'-O-(2-thiotriphosphate) (ATP{gamma}S; 100 µM, A) or 2-methyl-thio-ATP (100 µM, B) was applied. Time scale is indicated on the abscissa. Force of contraction is indicated at the ordinate.

 

Figure 3
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  		Fig. 3. Typical original recordings. Force of contraction in isolated electrically driven (1 Hz) atrial preparations from human hearts. At the indicated time, ATP (100 µM) was applied. Before addition of ATP, preparations were preincubated with the antagonists 1,3-dipropyl-cyclopentvl-xanthine (DPCPX, 1 µM, A), suramin (500 µM, B), pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS, 50 µM, C), reactive blue 2 (500 µM, D), or the inhibitors SQ-22536 (10 µM, E) or U-73122 (10 µM, F). Preparations were incubated for 30 min with antagonist/inhibitors. Next, ATP was additionally applied. Time scale is indicated on the abscissa. Force of contraction is indicated at the ordinate.

 
The PIE of ATP in the presence of 500 µM RB2 (Fig. 3D) is weak or even absent. We do not interpret this as evidence that the PIE of ATP is mediated via RB2-sensitive receptors. In contrast, it is obvious that this is probably due to the strong innate PIE of RB2 (beginning of the recording; Fig. 3D) on top of which a further positive inotropic drug is hardly effective.

We found that the maximum PIE of isoproterenol (1 nM to 1 µM were cumulatively applied) was unchanged by RB2; the maximum PIE of isoproterenol (at 1 µM) amounted to 12.9 ± 1.01 and 10.7 ± 3.40 mN in the absence and presence of RB2, respectively (n = 4, P > 0.05).

Furthermore, lower concentrations of RB2 [30 and 100 µM, which can at least partially block P2 receptors, compare Mantelli et al. (43), and did not by themselves increase force of contraction] did not affect either the NIE or PIE of ATP (data not shown). RB2 at concentrations >100 µM completely blocked human ATP-degrading enzymes (46). Hence it is conceivable that RB2 amplifies the effect of ATP by inhibiting its degradation. Whatever the underlying mechanism(s) are, others also descriptively reported on a PIE of RB2 at a similar concentration [isolated guinea pig heart preparations (64)].

To better understand the signal transduction system that mediates the inotropic effects of ATP, we tested whether its effects could be blocked by inhibitors previously used to inhibit PLC or cAMP-dependent protein kinase (PKA). We tested the involvement of these pathways because P2Y receptors (as well as some P1 receptors) have been noted to act via these well-studied pathways (for review, see Ref. 52). However, U-73122 (10 µM), a PLC inhibitor, and SQ-22536 (10 µM), a PKA inhibitor, did not affect the inotropic effects of ATP (Fig. 3, E and F). These compounds at these concentrations have been used before to study signal transduction via P2Y receptors in the heart [mouse (66) and chicken (50)]. It could be argued that, under our experimental conditions, these inhibitors are not effective. Therefore, we studied the effect of UTP on force of contraction in isolated mouse left atria, which were electrically paced like our human atrial preparations in the very same set up using the same buffers. In these mouse heart preparations, 100 µM UTP increased force of contraction. The effect was maximum after 10 min and amounted to 140 ± 18.6% of the predrug value (= UTP alone, P < 0.05). After preincubation (for 30 min) of the cardiac preparations with 10 µM SQ-22536 or 10 µM U-73122, the PIE of 100 µM UTP in the presence of these inhibitors was blunted and amounted to 113 ± 12 and 111 ± 17% of the predrug value, respectively (n = 7–11, P < 0.05 vs. UTP alone).


   DISCUSSION
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 ABSTRACT
 MATERIALS AND METHODS
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The main new finding of the present work is that ATP can exert PIEs in the human cardiac atrium. It remains to be elucidated whether the same holds true for human cardiac ventricle.

The concentration of ATP within cells, including cardiomyocytes (~10 mM), is much higher than in the extracellular space (~1–40 nM). Thus there is a huge gradient for ATP across the membrane. ATP (usually as MgATP2–) can pass through the intact sarcolemma from the inside to the outside, at a very small rate under normal conditions. The exact mechanism how ATP is transported through the intact cell membrane or released from muscle cells is not yet fully understood. It is possible that the cystic fibrosis transmembrane conductance regulator may act as an ATP channel or alternatively may regulate an ATP channel (56, 62). For instance, the release pathway from epithelial cells has been assumed to be either by a conductive pore (maxi anion channel: 20, volume regulated anion channels: 30), vesicular release, ATP-binding cassette transporters like the P-glycoprotein that mediated multidrug resistance (2), by connexins (17), an ectoapyrase (7), or through exocytosis (57). There might be cell-type-specific processes involved. When necrosis occurs during a myocardial infarction, a huge amount of ATP can be released from the dying myocyte because the intracellular concentration of ATP is in the millimolar range (58).

ATP can be degraded by soluble- or cell surface-located enzymes (ectonucleotidases and others) resident on endothelial cells but also on cardiomyocytes and other cell types (11). Apparently, it is the (low abundant) ATP4-moiety that binds to receptors, which have been divided into P1 and P2 receptors. P2 receptors are more sensitive to ATP than to adenosine; the opposite holds true for P1 receptors.

P1 receptors are further subclassified into A1, A2a, A2b, and A3 adenosine receptors (for review, see Ref. 52). In the heart, A1 adenosine receptors usually mediate NIEs via inhibition of the activity of adenylate cyclase and/or by activation of phosphatases via pertussis-sensitive G proteins (8, 27, 28). ATP is an agonist at P1 receptors. A1 adenosine receptor agonists like N6-phenylisopropyladenonsine and adenosine itself decreased (without any initial PIE) the force of contraction in the isolated electrically driven preparations from the human atrium (9). Their occupation could therefore only explain the NIE of ATP. From these data with the agonist adenosine it is unlikely that ATP acting on P1 receptors exerts its PIE. The conclusion is supported by the following data with a selective A1 antagonist and a selective A2 adenosine receptor agonist. Thus the NIE of ATP in the present work was not blocked by DPCPX, an A1 adenosine receptor antagonist (e.g., see Refs. 9 and 48), even at high concentrations as 10 µM DPCPX. This finding is unlike the case in rat and guinea pig atrium where the NIE of ATP is sensitive to 0.01 or 0.1 µM DPCPX (25, 43). Thus the A1 adenosine receptor is probably not involved in the NIE of ATP. Stimulation of A2 adenosine receptor by the selective agonist CGS-21680 can increase force of contraction in animal studies [isolated perfused rat heart (45)]. In contrast, we observed no PIE but a weak and slowly developing NIE with CGS-21680 (up to 10 µM) in human atrium (unpublished observation). Hence, based on the present findings and the literature, the PIE of ATP in the human atrium certainly does not involve P1 receptors, and the NIE of ATP in the human atrium probably involves neither A1 adenosine receptors nor A2 adenosine receptors.

In all likelihood, ATP acts via P2 receptors in the human atrium. P2 receptors are furthermore subdivided into P2X (ligand-gated ion channels) and P2Y receptors (G protein-coupled receptors). The mRNAs for all presently known P2X receptors (P2X1–7) and P2Y receptors (P2Y1,2,4,6,11,12,13,14) were detected in the human heart (3, 66). On protein level, P2Y1,4,6,12 and P2X1,2,3,4,6,7 have been detected in the human heart (3, 66). P2Y1,2,6,11,12,13 receptors are blocked by either suramin or PPADS or by RB2 (63). Because these antagonists do not block the contractile effects of ATP, this is one argument why P2Y receptors are not involved. In addition, P2Y receptors typically couple to PLC and/or adenylate cyclase. Because the inotropic effects of ATP are not blocked under conditions described in the literature (use of 10 µM U-73122 or SQ-22536), a further argument against the involvement of P2Y receptors can be raised. Moreover, ATP is not an agonist but an antagonist at human P2Y4 receptors (34); therefore, this receptor cannot play a role in the PIE of ATP. Moreover, 2-methyl-thio-ATP is inactive at P2Y2 and P2Y4 receptors (63), providing further evidence that these receptors are not involved because the PIE of ATP is mimicked by 2-methyl-thio-ATP (Fig. 2). As concerns the P2X receptors, only the human P2X6 and P2X4 receptors are poorly sensitive or insensitive to the tested P2 antagonists, namely PPADS, suramin, and RB2 (6, 16, 32). The fact that 2-methyl-thio-ATP induces a PIE suggests that this effect might be P2X4 receptor-mediated. Consistent with this assumption, the P2X4 receptors from rat mouse and humans are relatively insensitive to suramin. For instance, 100 µM suramin reduced P2X4-mediated currents by only 11% (32). Furthermore, 30 µM suramin inhibited the human P2X4-mediated current in cardiomyocytes from P2X4-overexpressing transgenic mice only marginally (55). In noncardiac cells, there is also evidence that P2X4 receptor-mediated effects of ATP cannot be blocked by suramin or PPADS (macrophages; see Ref. 12). In heterologous expression systems, the P2X4 receptor quickly (in 4 s by 30%) desensitizes to agonists (23), which is in contrast to the observed sustained PIE toward ATP, which we report here. However, our results concur with data on P2X4 receptor-infected chicken cardiac cells or cardiomyocytes from transgenic mice overexpressing P2X4 receptors (31). Overexpression of human P2X4 receptors in transgenic mice led to enhanced basal contractility (31). Thus P2X4 receptors may have effects in the heart, at least after genetic cardiac overexpression. Enhanced activity and/or expression of P2X4 receptors may be beneficial: crossbreeding of P2X4-overexpressing mice with calsequestrin-overexpressing mice (a genetic model of heart failure) had a beneficial effect on cardiac function and survival (67).

Some caveats are in order. There are heteromeric P receptors that are not yet fully studied with respect to agonists and antagonists (35) that could play a role in the human atrium, and a role of orphan receptors cannot be ruled out.

We can only speculate on the physiological meaning of the contractile effects of ATP. As mentioned above, the intracellular concentration of ATP in cells amounts to ~10 mM (58, 62). The concentration of ATP in the interstitium can profoundly increase during ischemia or stretch (5, 10, 39, 59). For instance, aortic endothelial cells released ATP at a rate of 2,900 fmol/min for one million cells (49). Another source of ATP is corelease from perivascular nerves (14). The concentration of ATP in erythrocytes amounts to ~2 mM and 0.2 µM in plasma (e.g., see Refs. 37 and 58). Hence leak of ATP from erythrocytes can rapidly lead to high ATP concentrations in the plasma.

The NIEs of ATP can only play a feedback role because they are transient over time. The PIE of ATP is sustained over time and may therefore be physiologically important to increase contractility, should this become necessary. Hence, a conceivable scenario may be as follows: during myocardial infarction, high amounts of ATP are released from dying myocytes to increase force in areas of the heart not yet damaged by cell death. Thus ATP may subserve to maintain contractile homeostasis in acute myocardial failure. It is tempting to speculate that, in chronic end-stage heart failure, inhibitors of ATP degradation can be useful to maintain contractility.

In summary, we describe a novel PIE of ATP in the human cardiac atrium. The receptors have been tentatively as P2X4-like receptors. The nature of the receptor mediating the initial NIE of ATP remains to be elucidated. Further work using more specific receptor blockers (presently not available) will be necessary to identify the receptors involved more rigorously.


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This work was supported by the Deutsche Forschungsgemeinschaft and by the Wilhelm-Roux-Program of the Medical Faculty of the University of Halle-Wittenberg.


   ACKNOWLEDGMENTS
 
The technical assistance of S. Reber is greatly appreciated.


   FOOTNOTES
 

Address for reprint requests and other correspondence: J. Neumann, Institut für Pharmakologie und Toxikologie, Medizinische Fakultät, Martin-Luther-Universität Halle-Wittenberg, Magdeburger Str. 4, D-06112 Halle, Germany (e-mail: joachim.neumann{at}medizin.uni-halle.de)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


   REFERENCES
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 ABSTRACT
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  1. Abbracchio MP, Burnstock G, Boeynaems JM, Barnard EA, Boyer JL, Kennedy C, Knight GE, Fumagalli M, Gachet C, Jacobson KA, Weisman GA. International Union of Pharmacology LVIII: update on the P2Y G protein-coupled nucleotide receptors: from molecular mechanisms and pathophysiology to therapy. Pharmacol Rev 58: 281–341, 2006.[Abstract/Free Full Text]
  2. Abraham EH, Prat AG, Gerweck L, Seneveratne T, Arceci RJ, Kramer R, Guidotti G, Cantiello HF. The multidrug resistance (mdr1) gene product functions as an ATP channel. Proc Natl Acad Sci USA 90: 312–316, 1993.[Abstract/Free Full Text]
  3. Banfi C, Ferrario S, De Vincenti O, Ceruti S, Fumagalli M, Mazzola A, D' Ambrosi N, Volonte C, Fratto P, Vitali E, Burnstock G, Beltrami E, Parolari A, Polvani G, Biglioli P, Tremoli E, Abbracchio MP. P2 receptors in human heart: upregulation of P2X6 in patients undergoing heart transplantation, interaction with TNFalpha and potential role in myocardial cell death. J Mol Cell Cardiol 39: 929–939, 2005.[CrossRef][Web of Science][Medline]
  4. Behnke N, Müller W, Neumann J, Schmitz W, Scholz H, Stein B. Differential antagonism by 1,3-dipropylxanthine-8-cyclopentylxanthine and 9-chloro-2(2-furanyl)-5,6-dihydro-1,2,4-triazolo (1,5-c)quinazolin-5-imine of the effects of adenosine derivatives in the presence of isoprenaline on contractile response and cAMP content in cardiomyocytes. Evidence for the coexistence of A1- and A2-adenosine receptors on cardiomyocytes. J Pharmacol Exp Ther 254: 1017–1023, 1990.[Abstract/Free Full Text]
  5. Berne RM. Cardiac nucleotides in hypoxia: possible role in regulation of coronary blood flow. Am J Physiol 204: 317–322, 1963.[Abstract/Free Full Text]
  6. Bo X, Zhang Y, Nassar M, Burnstock G, Schoepfer R. A P2X purinoceptor cDNA conferring a novel pharmacological profile. FEBS Lett 375: 129–133, 1995.[CrossRef][Web of Science][Medline]
  7. Bodas E, Aleu J, Pujol G, Martin-Satué M, Marsal J, Solsona C. ATP crossing the cell plasma membrane generates an ionic current in xenopus oocytes. J Biol Chem 275: 20268–20273, 2000.[Abstract/Free Full Text]
  8. Böhm M, Brückner R, Neumann J, Schmitz W, Scholz H, Starbatty J. Role of guanine nucleotide-binding protein in the regulation by adenosine of cardiac potassium conductance and force of contraction. Evaluation with pertussis toxin. Naunyn-Schmiedeberg's Arch Pharmacol 332: 403–405, 1986.[CrossRef][Web of Science][Medline]
  9. Böhm M, Pieske B, Ungerer M, Erdmann E. Characterization of A1 adenosine receptors in atrial and ventricular myocardium from diseased human hearts. Circ Res 65: 1201–1211, 1989.[Abstract/Free Full Text]
  10. Borst MM, Schrader J. Adenine nucleotide release from isolated perfused guinea pig hearts and extracellular formation of adenosine. Circ Res 68: 797–806, 1991.[Abstract/Free Full Text]
  11. Bowditch J, Brown AK, Dow JW. Accumulation and salvage of adenosine and inosine by isolated mature cardiac myocytes. Biochim Biophys Acta 844: 119–128, 1985.[Medline]
  12. Bowler JW, Bailey RJ, North RA, Surprenant A. P2X4, P2Y1 and P2Y2 receptors on rat alveolar macrophages. Br J Pharmacol 140: 567–575, 2003.[CrossRef][Web of Science][Medline]
  13. Burnstock G, Campbell G, Satchell D, Smythe A. Evidence that adenosine triphosphate or a related nucleotide is the transmitter substance released by non-adrenergic inhibitory nerves in the gut. Br J Pharmacol 40: 668–688, 1970.[Web of Science][Medline]
  14. Burnstock g. Purinergic nerves. Pharmacol Rev 24: 509–581, 1972.[Free Full Text]
  15. Burnstock G, Meghji P. Distribution of P1- and P2-purinoceptors in the guinea-pig and frog heart. Br J Pharmacol 73: 879–885, 1981.[Web of Science][Medline]
  16. Collo G, North RA, Kawashima E, Merlo-Pich E, Neidhart S, Surprenant A, Buell G. Cloning OF P2X5 and P2X6 receptors and the distribution and properties of an extended family of ATP-gated ion channels. J Neurosci 16: 2495–2507, 1996.[Abstract/Free Full Text]
  17. Cotrina ML, Lin JH, Alves-Rodrigues A, Liu S, Li J, Azmi-Ghadimi H, Kang J, Naus CC, Nedergaard M. Connexins regulate calcium signaling by controlling ATP release. Proc Natl Acad Sci USA 95: 15735–15740, 1998.[Abstract/Free Full Text]
  18. Day HJ, Holmsen H. Concepts of the blood platelet release reaction. Ser Haematol 4: 3–27, 1971.[Medline]
  19. Drury AN, Szent-Gyorgyi A. The physiological activity of adenine compounds with especial reference to their action upon the mammalian heart. J Physiol 68: 213–237, 1929.[Free Full Text]
  20. Dutta AK, Sabirov RZ, Uramoto H, Okada Y. Role of ATP-conductive anion channel in ATP release from neonatal rat cardiomyocytes in ischaemic or hypoxic conditions. J Physiol 559: 799–812, 2004.[Abstract/Free Full Text]
  21. Fabiato A, Fabiato F. Effects of pH on the myofilaments and the sarcoplasmic reticulum of skinned cells from cardiace and skeletal muscles. J Physiol 276: 233–255, 1978.[Abstract/Free Full Text]
  22. Fischer Y, Becker C, Loken C. Purinergic inhibition of glucose transport in cardiomyocytes. J Biol Chem 274: 755–761, 1999.[Abstract/Free Full Text]
  23. Fountain SJ, North RA. A C-terminal lysine that controls human P2X4 receptor desensitization. J Biol Chem 281: 15044–15049, 2006.[Abstract/Free Full Text]
  24. Fredholm BB, Abbracchio MP, Burnstock G, Daly JW, Harden TK, Jacobson KA, Leff P, Williams M. Nomenclature and classification of purinoceptors. Pharmacol Rev 46: 143–156, 1994.[Web of Science][Medline]
  25. Froldi G, Pandolfo L, Chinellato A, Ragazzi E, Caparrotta L, Fassina G. Dual effect of ATP and UTP on rat atria: which types of receptors are involved? Naunyn Schmiedeberg's Arch Pharmacol 349: 381–386, 1994.[Web of Science][Medline]
  26. Gergs U, Boknik P, Schmitz W, Simm A, Silber RE, Illes P, Neumann J. ATP modified contraction in the human heart (Abstract). Naunyn-Schmiedeberg's Arch Pharmacol 375, Suppl 1: 63, 2007.
  27. Gupta RC, Neumann J, Durant P, Watanabe AM. A1-adenosine-receptor mediated inhibition of isoproterenol-stimulated protein phosphorylation in ventricular myocytes. Evidence against a cyclic AMP-dependent effect. Circ Res 72: 65–74, 1993.[Abstract/Free Full Text]
  28. Gupta RC, Neumann J, Watanabe AM. Comparison of adenosine and muscarinic receptor mediated effects on phosphatase inhibitor-1 activity in the heart. J Pharmacol Exp Ther 266: 16–22, 1993.[Abstract/Free Full Text]
  29. Hirano Y, Abe S, Sawanobori T, Hiraoka M. Arachidonic acid induced increase in intracellular free calcium in guinea-pig hepatocytes. Jpn J Physiol 41: 327–332, 1991.[CrossRef][Web of Science][Medline]
  30. Hisadome K, Koyama T, Kimura C, Droogmans G, Ito Y, Oike M. Volume-regulated anion channels serve as an auto/paracrine nucleotide release pathway in aortic endothelial cells. J Gen Physiol 119: 511–520, 2002.[Abstract/Free Full Text]
  31. Hu B, Mei QB, Yao XJ, Smith E, Barry WH, Liang BT. A novel contractile phenotype with cardiac transgenic expression of the human P2X4 receptor. FASEB J 15: 2739–2741, 2001.[Free Full Text]
  32. Jones CA, Chessell IP, Simon J, Barnard EA, Miller KJ, Michel AD, Humphrey PP. Functional characterization of the P2X(4) receptor orthologues. Br J Pharmacol 129: 388–394, 2000.[CrossRef][Web of Science][Medline]
  33. Jorgensen S. Breakdown of adenine and hypoxanthine nucleotides and nucleosides in human plasma. Acta Pharmacol Toxicol (Copenh) 12: 294–302, 1956.[Medline]
  34. Kennedy C, Qi AD, Herold CL, Harden TK, Nicholas RA. ATP, an agonist at the rat P2Y(4) receptor, is an antagonist at the human P2Y(4) receptor. Mol Pharmacol 57: 926–931, 2000.[Abstract/Free Full Text]
  35. Kennedy C. P2X receptors: targets for novel analgesics? Neuroscientist 11: 345–356, 2005.[Abstract/Free Full Text]
  36. Khakh BS, Burnstock G, Kennedy C, King BF, North RA, Séguéla P, Voigt M, Humphrey PP. International union of pharmacology. XXIV. Current status of the nomenclature and properties of P2X receptors and their subunits. Pharmacol Rev 53: 107–118, 2001.[Abstract/Free Full Text]
  37. Kichenin K, Decollogne S, Angignard J, Seman M. Cardiovascular and pulmonary response to oral administration of ATP in rabbits. J Appl Physiol 88: 1962–1968, 2000.[Abstract/Free Full Text]
  38. Kirchhefer U, Baba HA, Kobayashi YM, Jones LR, Schmitz W, Neumann J. Altered function in atrium of transgenic mice overexpressing triadin 1. Am J Physiol Heart Circ Physiol 283: H1334–H1343, 2002.[Abstract/Free Full Text]
  39. Kuzmin AI, Lakomkin VL, Kapelko VI, Vassort G. Interstitial ATP level and degradation in control and postmyocardial infracted rats. Am J Physiol Cell Physiol 275: C766–C771, 1998.[Abstract/Free Full Text]
  40. Lambrecht G. Agonists and antagonists acting at P2X receptors: selectivity profiles and functional implications. Naunyn Schmiedebergs Arch Pharmacol 362: 340–350, 2000.[CrossRef][Web of Science][Medline]
  41. Legssyer A, Poggioli J, Renard D, Vassort G. ATP and other adenine compounds increase mechanical activity and inositol trisphosphate production in rat heart. J Physiol 401: 185–199, 1988.[Abstract/Free Full Text]
  42. Lohse MJ, Boser S, Klotz KN, Schwabe U. Affinities of barbiturates for the GABA-receptor complex and A1 adenosine receptors: a possible explanation of their excitatory effects. Naunyn Schmiedebergs Arch Pharmacol 336: 211–217, 1987.[CrossRef][Web of Science][Medline]
  43. Mantelli L, Amerini S, Filippi S, Ledda F. Blockade of adenosine receptors unmasks a stimulatory effect of ATP on cardiac contractility. Br J Pharmacol 109: 1268–1271, 1993.[Web of Science][Medline]
  44. Mei Q, Liang BT. P2 purinergic receptor activation enhances cardiac contractility in isolated rat and mouse hearts. Am J Physiol Heart Circ Physiol 281: H334–H341, 2001.[Abstract/Free Full Text]
  45. Monahan TS, Sawmiller DR, Fenton RA, Dobson JG Jr. Adenosine A(2a)-receptor activation increases contractility in isolated perfused hearts. Am J Physiol Heart Circ Physiol 279: H1472–H1481, 2000.[Abstract/Free Full Text]
  46. Munkonda MN, Kauffenstein G, Kukulski F, Levesque SA, Legendre C, Pelletier J, Lavoie EG, Lecka J, Sevigny J. Inhibition of human and mouse plasma membrane bound NTPDases by P2 receptor antagonists. Biochem Pharmacol 74: 1524–1534, 2007.[CrossRef][Web of Science][Medline]
  47. Needleman P, Minkes MS, Douglas JR Jr. Stimulation of prostaglandin biosynthesis by adenine nucleotides. Profile of prostaglandin release by perfused organs. Circ Res 34: 455–460, 1974.[Abstract/Free Full Text]
  48. Neumann J, Schmitz W, Scholz H, Stein B. Effects of adenosine analogues on contractile response and cAMP content in guinea-pig isolated ventricular myocytes. Naunyn-Schmiedeberg's Arch Pharmacol 340: 689–695, 1989.[CrossRef][Web of Science][Medline]
  49. Oike M, Kimura C, Koyama T, Yoshikawa M, Ito Y. Hypotonic stress-induced dual Ca2+ responses in bovine aortic endothelial cells. Am J Physiol Heart Circ Physiol 279: H630–H638, 2000.[Abstract/Free Full Text]
  50. Podrasky Ernest David Xu, Bruce Liang T. A novel phospholipase C- and cAMP-independent positive inotropic mechanism via a P2 purinoceptor. Am J Physiol Heart Circ Physiol 273: H2380–H2387, 1997.[Abstract/Free Full Text]
  51. Puceat M, Clement-Chomienne O, Terzic A, Vassort G. Alpha 1-adrenoceptor and purinoceptor agonists modulate Na-H antiport in single cardiac cells. Am J Physiol Heart Circ Physiol 264: H310–H319, 1993.[Abstract/Free Full Text]
  52. Ralevic V, Burnstock G. Receptors for purines and pyrimidines. Pharmacol Rev 50: 413–492, 1998.[Abstract/Free Full Text]
  53. Saito D, Ueeda M, Abe Y, Tani H, Nakatsu T, Yoshida H, Haraoka S, Nagashima H. Treatment of paroxysmal supraventricular tachycardia with intravenous injection of adenosine triphosphate. Br Heart J 55: 291–294, 1986.[Abstract/Free Full Text]
  54. Scamps F, Legssyer A, Mayoux E, Vassort G. The mechanism of positive inotropy induced by adenosine triphosphate in rat heart. Circ Res 67: 1007–1016, 1990.[Abstract/Free Full Text]
  55. Shen JB, Pappano AJ, Liang BT. Extracellular ATP-stimulated current inwild-type and P2X4 receptor transgenic mouse ventricular myocytes: implications for a cardiac physiologic role of P2X4 receptors. FASEB J 20: 277–284, 2006.[Abstract/Free Full Text]
  56. Sugita M, Yue Y, Foskett JK. CFTR Cl- channel and CFTR-associated ATP channel: distinct pores regulated by common gates. EMBO J 17: 898–908, 1998.[CrossRef][Web of Science][Medline]
  57. Tatur S, Groulx N, Orlov SN, Grygorczyk R. Ca2+-dependent ATP release from A549 cells involves synergistic autocrine stimulation by coreleased uridine nucleotides. J Physiol 584: 419–435, 2007.[Abstract/Free Full Text]
  58. Traut TW. Physiological concentrations of purines and pyrimidines. Mol Cell Biochem 140: 1–22, 1994.[CrossRef][Web of Science][Medline]
  59. Uozumi H, Kudoh S, Zou Y, Harada K, Yamazaki T, Komuro I. Autocrine release of ATP mediates mechanical stress-induced cardiomyocytes hypertrophy (Abstract). Circulation 98: I624, 1998.
  60. Vahlensieck U, Boknik P, Knapp J, Linck B, Müller FU, Neumann J, Schmitz W, Herzig S, Gross I, Schlüter H, Zidek W, Deng MC, Scheld HH. Negative chronotropic and inotropic effects exerted by diadenosine hexaphosphate (AP6A) via A1-adenosine receptors. Br J Pharmacol 119: 835–844, 1996.[Web of Science][Medline]
  61. Vahlensieck U, Boknik P, Gombosova I, Huke S, Knapp J, Linck B, Lüss H, Müller FU, Neumann J, Deng MC, Scheld HH, Jankowski H, Schlüter H, Zidek W, Zimmermann N, Schmitz W. Inotropic effects of diadenosine tetraphosphate (AP4A) in human and animal cardiac preparations. J Pharmacol Exp Ther 288: 805–813, 1999.[Abstract/Free Full Text]
  62. Vassort G. Adenosine 5'-triphosphate: a P2-purinergic agonist in the myocardium. Physiol Rev 81: 767–806, 2001.[Abstract/Free Full Text]
  63. von Kügelgen I. Pharmacological profiles of cloned mammalian P2Y-receptor subtypes. Pharmacol Ther 110: 415–432, 2006.[CrossRef][Web of Science][Medline]
  64. Wee S, Peart JN, Headrick JP. P2 Purinoceptor-Mediated Cardioprotection in Ischemic-Reperfused Mouse Heart. J Pharmacol Exp Ther September 12, 2007;doi: 10.1124/jpet.107.125815.[Abstract/Free Full Text]
  65. Welford LA, Cusack NJ, Hourani SM. The structure-activity relationships of ectonucleotidases and of excitatory P2-purinoceptors: evidence that dephosphorylation of ATP analogues reduces pharmacological potency. Eur J Pharmacol 141: 123–130, 1987.[CrossRef][Web of Science][Medline]
  66. Wihlborg AK, Balogh J, Wang L, Borna C, Dou Y, Joshi BV, Lazarowski E, Jacobson KA, Arner A, Erlinge D. Positive inotropic effects by uridine triphosphate (UTP) and uridine diphosphate (UDP) via P2Y2 and P2Y6 receptors on cardiomyocytes and release of UTP in man during myocardial infarction. Circ Res 98: 970–976, 2006.[Abstract/Free Full Text]
  67. Yang A, Sonin D, Jones L, Barry WH, Liang BT. A beneficial role of cardiac P2X4 receptors in heart failure: rescue of the calsequestrin overexpression model of cardiomyopathy. Am J Physiol Heart Circ Physiol 287: H1096–H1103, 2004.[Abstract/Free Full Text]
  68. Zheng JS, Boluyt MO, Long X, O'Neill L, Lakatta EG, Crow MT. Extracellular ATP inhibits adrenergic agonist-induced hypertrophy of neonatal cardiac myocytes. Circ Res 78: 525–535, 1996.[Abstract/Free Full Text]
  69. Zimmermann N, Nacke P, Neumann J, Winter J, Gams E. Inotropic effects of diadenosine monophosphate (AP1A) in isolated human cardiac preparations. J Cardiovasc Pharmacol 35: 881–886, 2000.[CrossRef][Web of Science][Medline]



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D. Sonin, S.-Y. Zhou, C. Cronin, T. Sonina, J. Wu, K. A. Jacobson, A. Pappano, and B. T. Liang
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