The present renal hemodynamic study tested the hypothesis that CD38 and superoxide anion (O2(·-)) participate in the vasoconstriction produced by activation of thromboxane prostanoid (TP) receptors in the mouse kidney. CD38 is the major mammalian ADP-ribosyl cyclase contributing to vasomotor tone through the generation of cADP-ribose, a second messenger that activates ryanodine receptors to release Ca(2+) from the sarcoplasmic reticulum in vascular smooth muscle cells. We evaluated whether the stable thromboxane mimetic U-46619 causes less pronounced renal vasoconstriction in CD38-deficient mice and the involvement of O2(·-) in U-46619-induced renal vasoconstriction. Our results indicate that U-46619 activation of TP receptors causes renal vasoconstriction in part by activating cADP-ribose signaling in renal resistance arterioles. Based on maximal renal blood flow and renal vascular resistance responses to bolus injections of U-46619, CD38 contributes 30-40% of the TP receptor-induced vasoconstriction. We also found that the antioxidant SOD mimetic tempol attenuated the magnitude of vasoconstriction by U-46619 in both groups of mice, suggesting mediation by O2(·-). The degree of tempol blockage of U-46619-induced renal vasoconstriction was greater in wild-type mice, attenuating renal vasoconstriction by 40% compared with 30% in CD38-null mice. In other experiments, U-46619 rapidly stimulated O2(·-) production (dihydroethidium fluorescence) in isolated mouse afferent arterioles, an effect abolished by tempol. These observations provide the first in vivo demonstration of CD38 and O2(·-) involvement in the vasoconstrictor effects of TP receptor activation in the kidney and in vitro evidence for TP receptor stimulation of O2(·-) production by the afferent arteriole.

Renal blood flow (RBF) responses to arginine vasopressin (AVP) were tested in anesthetized wild-type (WT) and CD38(-/-) mice that lack the major calcium-mobilizing second messenger cyclic ADP ribose. AVP (3-25 ng) injected intravenously produced dose-dependent decreases in RBF, reaching a maximum of 25 ± 2% below basal RBF in WT and 27 ± 2% in CD38(-/-) mice with 25 ng of AVP. Renal vascular resistance (RVR) increased 75 ± 6% and 78 ± 6% in WT and CD38(-/-) mice. Inhibition of nitric oxide (NO) synthase with nitro-L-arginine methyl ester (L-NAME) increased the maximum RVR response to AVP to 308 ± 76% in WT and 388 ± 81% in CD38(-/-) (P < 0.001 for both). Cyclooxygenase inhibition with indomethacin increased the maximum RVR response to 125 ± 15% in WT and 120 ± 14% in CD38(-/-) mice (P < 0.001, <0.05). Superoxide suppression with tempol inhibited the maximum RVR response to AVP by 38% in both strains (P < 0.005) but was ineffective when administered after L-NAME. The rate of RBF recovery (relaxation) after AVP was slowed by L-NAME and indomethacin (P < 0.001, <0.005) but was unchanged by tempol. All vascular responses to AVP were abolished by an AVP V1a receptor antagonist. A V2 receptor agonist or antagonist had no effect on AVP-induced renal vasoconstriction. Taken together, the results indicate that renal vasoconstriction by AVP in the mouse is strongly buffered by vasodilatory actions of NO and prostanoids. The vasoconstriction depends on V1a receptor activation without involvement of CD38 or concomitant vasodilatation by V2 receptors. The role of superoxide is to enhance the contractile response to AVP, most likely by reducing the availability of NO rather than directly stimulating intracellular contraction signaling pathways.

The calcium mobilizing second messenger, cyclic ADP ribose (cADPR, the product of ADP‐ribosyl cyclase) is an important component in the renal vasoconstrictor response to G‐protein coupled receptor stimulation. The role of cADPR in TpR‐mediated renal vasoconstriction was tested with the stable thromboxane mimetic U‐46619 in wild type (WT) and CD38 −/− mice. Arterial blood pressure (ABP) and renal blood flow (RBF) were not different between WT and CD38−/− mice during basal conditions. Bolus injections of U‐46619 (10–50ng in 10μl) caused a dose‐dependent decrease in RBF and increase in ABP in both strains but to a lesser extent for both in CD38−/− mice. Compared to control, 50 ng of U‐46619 reduced RBF by 37±4% and 22±3% and increased ABP by 36±6% and 22±6% in WT and CD38−/− mice, respectively (p<0.05 n=7). Intravenous infusion of Tempol to scavenge superoxide reduced renal vasoconstrictor response to U‐46619 by 40–45% in both strains of mice. The data show that the renal vasoconstriction produced by TpR stimulation is mediated by ADP‐ribosyl cyclase activity and depends upon superoxide for full expression. The comparable inhibition of the RBF response by Tempol in both strains of mice indicates that the vasoconstrictor action of superoxide is independent of CD38 and cannot explain the attenuated response to TpR activation in CD38−/− mice. Funded by NIH grant RO1 HL‐02334

Renal vascular responses to AVP were tested in anesthetized wild‐type (WT) and CD38‐/‐ mice that lack the calcium mobilizing second messenger cyclic ADP ribose. Iv injections of AVP (3‐25ng) produced a dose‐dependent decrease in renal blood flow with a minimum of 25±2% below control in WT and 27±2% in CD38‐/‐ mice at the 25ng dose. Renal vascular resistance (RVR) increased by 75±6% in WT and 78±6% in CD38‐/‐ mice. Inhibition of NO synthase with L‐NAME increased the maximum RVR response to AVP from 71±7 to 308±76% in WT and from 86±7 to 388±81% in CD38‐/‐. Cyclooxygenase inhibition with indomethacin increased the maximum RVR response to 125±15% in WT and 120±14% in CD38‐/‐ mice (P<0.001, <0.05). Superoxide suppression with tempol inhibited the maximum RVR response to AVP by 38% in both strains (p<0.005). All vascular responses to AVP were abolished by an AVP V1a receptor antagonist. Thus, vasoconstriction by AVP in the mouse is caused by V1a receptor activation with no concomitant vasodilatation by V2 receptors. However, renal vasoconstrictor responses to AVP are strongly buffered by vasodilatory actions of NO and prostanoids and augmented by the vasoconstrictor actions of superoxide. Unlike AVP, other vasoconstrictor agents show attenuated vasoconstrictor activity in CD38‐/‐ mice which points to a distinct activation mechanism for AVP that appears to be independent of CD38 and its downstream calcium signaling pathway. Grant Funding Source: Supported by HL‐02334