Pannexin 1 channels control the hemodynamic response to hypoxia by regulating O2-sensitive extracellular ATP in blood

Abstract

Pannexin 1 (Panx1) channels export ATP and may contribute to increased concentration of the vasodilator ATP in plasma during hypoxia in vivo. We hypothesized that Panx1 channels and associated ATP export contribute to hypoxic vasodilation, a mechanism that facilitates the matching of oxygen delivery to metabolic demand of tissue. Male and female mice devoid of Panx1 (Panx1^-/-) and wild-type controls (WT) were anesthetized, mechanically ventilated, and instrumented with a carotid artery catheter or femoral artery flow transducer for hemodynamic and plasma ATP monitoring during inhalation of 21% (normoxia) or 10% oxygen (hypoxia). ATP export from WT vs. Panx1^-/-erythrocytes (RBC) was determined ex vivo via tonometer experimentation across progressive deoxygenation. Mean arterial pressure (MAP) was similar in Panx1^-/- (n = 6) and WT (n = 6) mice in normoxia, but the decrease in MAP in hypoxia seen in WT was attenuated in Panx1^-/- mice (-16 ± 9% vs. -2 ± 8%; P < 0.05). Hindlimb blood flow (HBF) was significantly lower in Panx1^-/- (n = 6) vs. WT (n = 6) basally, and increased in WT but not Panx1^-/- mice during hypoxia (8 ± 6% vs. -10 ± 13%; P < 0.05). Estimation of hindlimb vascular conductance using data from the MAP and HBF experiments showed an average response of 28% for WT vs. -9% for Panx1^-/- mice. Mean venous plasma ATP during hypoxia was 57% lower in Panx1^-/- (n = 6) vs. WT mice (n = 6; P < 0.05). Mean hypoxia-induced ATP export from RBCs from Panx1^-/- mice (n = 8) was 82% lower than that from WT (n = 8; P < 0.05). Panx1 channels participate in hemodynamic responses consistent with hypoxic vasodilation by regulating hypoxia-sensitive extracellular ATP levels in blood.NEW & NOTEWORTHY Export of vasodilator ATP from red blood cells requires pannexin 1. Blood plasma ATP elevations in response to hypoxia in mice require pannexin 1. Hemodynamic responses to hypoxia are accompanied by increased plasma ATP in mice in vivo and require pannexin 1.

Keywords: blood flow; erythrocyte; hypoxia; pannexin; vasodilation.

Figure 1.

Mean arterial blood pressure (MAP) for WT or Panx1^−/− mice during a 30-min hypoxic challenge. Individual values and means ± SD are shown. A and B absolute values and percent change from baseline. *P < 0.05 vs. baseline for WT but not for Panx1^−/− mice. †P < 0.05 compared to WT as determined by two-way repeated-measures ANOVA with Student–Newman–Keuls post hoc analysis. n = 6 for WT mice (4 males, 2 females) and n = 6 for Panx1^−/− mice (6 males). Red shading denotes normoxia (21% FiO₂) and blue shading denotes hypoxia (10% FiO₂). Panx1, pannexin 1; WT, wild-type.

Figure 2.

Heart rate (HR) for WT or Panx1^−/− mice during a 30-min hypoxic challenge. Individual values and means ± SD are shown. A and B: absolute values and percent change from baseline. n = 6 for WT mice (4 males, 2 females) and n = 6 for Panx1^−/− mice (6 males). Red shading denotes normoxia (21% FiO₂) and blue shading denotes hypoxia (10% FiO₂). Panx1, pannexin 1; WT, wild-type.

Figure 3.

Hindlimb blood flow (HBF) for WT or Panx1^−/− mice during a 30-min hypoxic challenge. Individual values and means ± SD are shown. A and B: absolute values and percent change from baseline. *P < 0.05 vs. respective baseline. †P < 0.05 compared to WT as determined by two-way repeated-measures ANOVA with Student–Newman–Keuls post hoc analysis. n = 6 for WT mice (5 males, 1 females) and n = 6 for Panx1^−/− mice (5 males, 1 females). Red shading denotes normoxia (21% FiO₂) and blue shading denotes hypoxia (10% FiO₂). Panx1, pannexin 1; WT, wild-type.

Figure 4.

Calculated, estimated hindlimb vascular conductance during a 30-min hypoxic challenge using MAP from mouse group 1 and HBF from mouse group 2. Hindlimb vascular conductance = (group average HBF/group average MAP) × 100. Values are % change from baseline. n = 12 for WT mice (9 males, 3 females) and n = 12 for Panx1^−/− mice (11 males, 1 females). Red shading denotes normoxia (21% FiO₂) and blue shading denotes hypoxia (10% FiO₂). HBF, hindlimb blood flow; MAP, mean arterial pressure; Panx1, pannexin 1; WT, wild-type.

Figure 5.

Plasma ATP concentration (nM) for WT or Panx1^−/− mice after 30 min of breathing either 21% or 10% O₂. Individual values and means ± SD are shown. †P < 0.05 compared to WT as determined by unpaired t test. For normoxia time control, n = 4 for WT mice (4 males) and n = 4 for Panx1^−/− mice (4 males). For hypoxia, n = 6 for WT mice (4 males, 2 females) and n = 6 for Panx1^−/− mice (6 males). Panx1, pannexin 1; WT, wild-type.

Figure 6.

Extracellular ATP increased progressively with the deoxygenation of WT and, to a lesser extent, Panx1^−/− RBCs. Individual values and means ± SD are shown. Absolute values are summarized in (A) and the percent change from baseline in (B). *P < 0.05 vs. respective baseline and †P < 0.05 compared to WT as determined by two-way repeated-measures ANOVA with Student–Newman–Keuls post hoc analysis. n = 8 for WT mice (6 males, 2 females) and n = 8 for Panx1^−/− mice (6 males, 2 females). Panx1, pannexin 1; WT, wild-type.

Figure 7.

Percent change in extracellular ATP following RBC incubation of RBCs from WT or Panx1^−/− mice with mastoparan 7 (10 µM). Individual values and means ± SD are shown. †P < 0.05 compared to WT as determined by unpaired t test. n = 4 for WT mice (4 males) and n = 4 for Panx1^−/− mice (4 males). Panx1, pannexin 1; WT, wild-type.

Figure 8.

Oxygen binding curves of RBCs from WT (n = 6 males) or Panx1^−/− mice (n = 7 males). The insert shows mean and standard error of P₅₀ values. Mean P₅₀ values (inset) did not differ significantly as determined by unpaired t test. Panx1, pannexin 1; WT, wild-type.