Phosphodiesterase 3 is present in rabbit and human erythrocytes and its inhibition potentiates iloprost-induced increases in cAMP

American Journal of Physiology-Heart and Circulatory Physiology

et al. 2010

Self Cite

164

179

is the conduit for low oxygen tension-induced ATP release from human erythrocytes. Am J Physiol Heart Circ Physiol 299: H1146 -H1152, 2010. First published July 9, 2010; doi:10.1152/ajpheart.00301.2010.-Erythrocytes release ATP in response to exposure to the physiological stimulus of lowered oxygen (O 2) tension as well as pharmacological activation of the prostacyclin receptor (IPR). ATP release in response to these stimuli requires activation of adenylyl cyclase, accumulation of cAMP, and activation of protein kinase A. The mechanism by which ATP, a highly charged anion, exits the erythrocyte in response to lowered O 2 tension or receptor-mediated IPR activation by iloprost is unknown. It was demonstrated previously that inhibiting pannexin 1 with carbenoxolone inhibits hypotonically induced ATP release from human erythrocytes. Here we demonstrate that three structurally dissimilar compounds known to inhibit pannexin 1 prevent ATP release in response to lowered O 2 tension but not to iloprost-induced ATP release. These results suggest that pannexin 1 is the conduit for ATP release from erythrocytes in response to lowered O 2 tension. However, the identity of the conduit for iloprost-induced ATP release remains unknown.iloprost; carbenoxolone; probenecid; red blood cell; cystic fibrosis transmembrane conductance regulator ERYTHROCYTES CONTRIBUTE to the regulation of vascular caliber by virtue of their ability to release adenosine 5=-triphosphate (ATP) (14 -17, 48, 50). ATP released from erythrocytes binds to purinergic receptors on the vascular endothelium, which leads to the local formation of endothelium-derived vasodilators such as nitric oxide, prostaglandins, and endotheliumderived hyperpolarizing factor (20,46,50).Erythrocytes release ATP in response to mechanical deformation and exposure to lowered oxygen (O 2 ) tension and in response to incubation with pharmacological agents such as the prostacyclin analog iloprost (Ilo) (5,44,45). ATP release in response to these various stimuli requires activation of adenylyl cyclase, accumulation of cAMP, and activation of PKA (3,43,47). Although several components of the signaling pathway for ATP release from erythrocytes have been investigated, the identity of the conduit by which ATP exits these cells is not fully characterized. In other cell types, several membrane channels, including connexin hemichannels, voltage-dependent anion channels, volume-regulated anion channels, and ATP-binding cassette proteins, have been implicated as conduits for ATP release (2,27,29,37,40,41). In addition, the cystic fibrosis transmembrane conductance regulator (CFTR), an ATP-binding cassette protein required for ATP release from erythrocytes in response to mechanical deformation (25, 45), was once considered to be a possible ATP conduit in cells. However, more recent studies demonstrate that CFTR is not likely to serve as an ATP conduit itself but rather regulates other channels that serve that role (1,7,19,24,51).Recently, the protein family of pannexins, orthologs of the invert...

Section: Isolation Of Healthy Human Erythrocytesmentioning

confidence: 99%

“…Isolated erythrocytes, exposed to room air and temperature, were diluted to a hematocrit of 20% in a buffer containing (in mM) 21 (28,34,42).…”

Section: Isolation Of Healthy Human Erythrocytesmentioning

confidence: 99%

Pannexin 1 is the conduit for low oxygen tension-induced ATP release from human erythrocytes

Sridharan

Adderley

American Journal of Physiology-Heart and Circulatory Physiology

et al. 2010

Self Cite

164

179

“…The activity of the heterotrimeric G protein, Gi, is required for ATP release from erythrocytes in response to reduced O 2 (32,33), wheras a second G protein, Gs, is associated with a prostacyclin receptor (IP) (32). It has been shown that G␣i2 expression is decreased in erythrocytes of humans with DM2 and this defect is associated with impairment of both cAMP accumulation and ATP release when these cells are incubated with a direct activator of Gi (21,39), suggesting that this defect in erythrocyte physiology could contribute to the associated vascular disease. Interestingly, expression of Gs is unaltered in DM2 erythrocytes, suggesting that prostacyclin-induced ATP release might remain intact.…”

mentioning

confidence: 99%

Divergent effects of low-O₂tension and iloprost on ATP release from erythrocytes of humans with type 2 diabetes: implications for O₂supply to skeletal muscle

Sprague

Goldman

American Journal of Physiology-Heart and Circulatory Physiology

et al. 2010

Self Cite

ML. Divergent effects of low-O 2 tension and iloprost on ATP release from erythrocytes of humans with type 2 diabetes: implications for O 2 supply to skeletal muscle. Am J Physiol Heart Circ Physiol 299: H566 -H573, 2010. First published May 28, 2010 doi:10.1152/ajpheart.00430.2010.-Erythrocytes release both O 2 and a vasodilator, ATP, when exposed to reduced O2 tension. We investigated the hypothesis that ATP release is impaired in erythrocytes of humans with type 2 diabetes (DM2) and that this defect compromises the ability of these cells to stimulate dilation of resistance vessels. We also determined whether a general vasodilator, the prostacyclin analog iloprost (ILO), stimulates ATP release from healthy human (HH) and DM2 erythrocytes. Finally, we used a computational model to compare the effect on tissue O 2 levels of increases in blood flow directed to areas of increased O2 demand (erythrocyte ATP release) with nondirected increases in flow (ILO). HH erythrocytes, but not DM2 cells, released increased amounts of ATP when exposed to reduced O2 tension (PO2 Ͻ 30 mmHg). In addition, isolated hamster skeletal muscle arterioles dilated in response to similar decreases in extraluminal O 2 when perfused with HH erythrocytes, but not when perfused with DM2 erythrocytes. In contrast, both HH and DM2 erythrocytes released ATP in response to ILO. In the case of DM2 erythrocytes, amounts of ATP released correlated inversely with glycemic control. Modeling revealed that a functional regulatory system that directs blood flow to areas of need (low O2-induced ATP release) provides appropriate levels of tissue oxygenation and that this level of the matching of O2 delivery with demand in skeletal muscle cannot be achieved with a general vasodilator. These results suggest that the inability of erythrocytes to release ATP in response to exposure to low-O2 tension could contribute to the peripheral vascular disease of DM2. muscle blood flow; red blood cells; oxygen delivery; prostacyclin CARDIOVASCULAR DISEASE ACCOUNTS for nearly half of deaths in humans with type 2 diabetes (DM2) (31, 41). It is estimated that in 2010 there will be in excess of 300 million cases of DM2 worldwide making this disease a major public health challenge (4). A significant complication of DM2 is impaired vascular function that contributes to a fourfold increased risk for claudication (4) and as much as a 16-fold increased risk for lower limb amputation (12,30,43). Although individuals with DM2 have an increased incidence of atherosclerosis in largeconduit vessels (4, 30), there is extensive evidence that microvascular circulatory control is also abnormal in humans with DM2 (23,24,27,35). Although direct studies of the skeletal muscle microcirculation are not possible in humans, such studies have been performed in several animal models of diabetes and reveal reduced convective O 2 delivery and diffusive O 2 transport that could contribute to a failure of the skeletal muscle vasculature to deliver appropriate amounts of O 2 to meet metabolic need bot...

“…12,13 Since human erythrocytes possess PDE5, a PDE that hydrolyzes cGMP, we hypothesized that inhibition of PDE5 would increase cAMP levels in these cells in response to activation of the IPR by the prostacyclin analog, UT-15C. As depicted in Figure 2, in the presence of the PDE5 inhibitor tadalafil (10 mmol/L, n ¼ 7), UT-15C-stimulated increases in cAMP were potentiated (P < 0.01).…”

Section: Characteristics Of Subjectsmentioning

confidence: 99%

“…10,11 ATP release requires increases in intracellular cAMP . [10][11][12] The level of this cyclic nucleotide is modulated by phosphodiesterase 3 (PDE3). 12 The critical role of PDE3 is demonstrated by the observation that, in healthy human erythrocytes, pharmacological inhibitors of PDE3 increase both intracellular cAMP and ATP release in response to IPR activation.…”

Section: Introductionmentioning

confidence: 99%

Phosphodiesterase 5 inhibitors augment UT-15C-stimulated ATP release from erythrocytes of humans with pulmonary arterial hypertension

Moody

Yeragunta

et al. 2014

Exp Biol Med (Maywood)

Both prostacyclin analogs and phosphodiesterase 5 (PDE5) inhibitors are effective treatments for pulmonary arterial hypertension (PAH). In addition to direct effects on vascular smooth muscle, prostacyclin analogs increase cAMP levels and ATP release from healthy human erythrocytes. We hypothesized that UT-15C, an orally available form of the prostacyclin analog, treprostinil, would stimulate ATP release from erythrocytes of humans with PAH and that this release would be augmented by PDE5 inhibitors. Erythrocytes were isolated and the effect of UT-15C on cAMP levels and ATP release were measured in the presence and absence of the PDE5 inhibitors, zaprinast or tadalafil. In addition, the ability of a soluble guanylyl cyclase inhibitor to prevent the effects of tadalafil was determined. Erythrocytes of healthy humans and humans with PAH respond to UT-15C with increases in cAMP levels and ATP release. In both groups, UT-15C-induced ATP release was potentiated by zaprinast and tadalafil. The effect of tadalafil was prevented by pre-treatment with an inhibitor of soluble guanylyl cyclase in healthy human erythrocytes. Importantly, UT-15C-induced ATP release was greater in PAH erythrocytes than in healthy human erythrocytes in both the presence and the absence of PDE5 inhibitors. The finding that prostacyclin analogs and PDE5 inhibitors work synergistically to enhance release of the potent vasodilator ATP from PAH erythrocytes provides a new rationale for the co-administration of these drugs in this disease. Moreover, these results suggest that the erythrocyte is a novel target for future drug development for the treatment of PAH.