Nitric oxide (NO) produced by the endothelium diffuses both into the lumen and to the smooth muscle cells according to the concentration gradient in each direction. The extremely high reaction rate between NO and hemoglobin (Hb), k _Hb = 3–5 × 10 ⁷ M ⁻¹ ⋅s ⁻¹ , suggests that most of the NO produced would be consumed by Hb in the red blood cells (RBCs), which then would block the biological effect of NO. Therefore, specific mechanisms must exist under physiological conditions to reduce the NO consumption by RBCs, in which the Hb concentration is very high (24 mM heme). By using isolated microvessels as a bioassay, here we show that physiological concentrations of RBCs in the presence of intravascular flow does not inhibit NO-mediated vessel dilation, suggesting that RBCs under this condition are not an NO scavenger. On the other hand, RBCs (50% hematocrit) without intravascular flow reduce NO-mediated dilation to serotonin by 30%. In contrast, free Hb (10 μM) completely inhibits NO-mediated dilation with or without intravascular flow. The effect of flow on NO consumption by RBCs may be attributed to the formation of an RBC-free zone near the vessel wall, which is caused by hydrodynamic forces on particles. Intravascular flow does not affect the reaction rate between NO and free Hb in the lumen, because the latter forms a homogeneous solution and is not subject to the hydrodynamic separation. However, intravascular flow only partially contributes to the reduced consumption of NO by RBCs, because without the flow, the NO consumption by RBCs is already about 3 orders of magnitude slower than free Hb.

Nitric oxide (NO) produced by the vascular endothelium is an important biologic messenger that regulates vessel tone and permeability and inhibits platelet adhesion and aggregation. NO exerts its control of vessel tone by interacting with guanylyl cyclase in the vascular smooth muscle to initiate a series of reactions that lead to vessel dilation. Previous efforts to investigate this interaction by mathematical modeling of NO diffusion and reaction have been hampered by the lack of information on the production and degradation rate of NO. We use a mathematical model and previously published experimental data to estimate the rate of NO production, 6.8 × 10⁻¹⁴μmol ⋅ μm⁻²⋅ s⁻¹; the NO diffusion coefficient, 3,300 μm²s⁻¹; and the NO consumption rate coefficient in the vascular smooth muscle, 0.01 s⁻¹(1st-order rate expression) or 0.05 μM⁻¹⋅ s⁻¹(2nd-order rate expression). The modeling approach is discussed in detail. It provides a general framework for modeling the NO produced from the endothelium and for estimating relevant physical parameters.

Flow-mediated dilation has been documented in large conduit coronary arteries but not in coronary arterioles. The goal of this study was to determine whether this response occurs in coronary arterioles and whether it competes with myogenic constriction. Subepicardial arterioles (40-80 microns) were isolated and cannulated with two glass micropipettes connected to independent reservoir systems. During zero flow, myogenic responses were studied over the range of intraluminal pressure (IP) between 20 and 140 cmH2O. Myogenic constrictions and dilations was observed when IP was increased (greater than 60 cmH2O) and decreased (less than 60 cmH2O), respectively. Flow was initiated by simultaneously moving the reservoirs in equal and opposite directions, thus generating a pressure gradient (delta P) without changing the mean luminal pressure (range delta P = 4-60 cmH2O). Flow-induced responses were studied at low, intermediate, and high myogenic tones by setting IP at 20, 60, and 100 cmH2O, respectively. The threshold for flow-induced dilation was delta P = 4 cmH2O, and maximum dilation was observed at delta P = 20 cmH2O. Red cell velocities in isolated arterioles at delta P of 4 and 60 cmH2O were 1.2 +/- 0.2 and 15.9 +/- 1.3 mm/s, respectively, which are within the range of those reported for coronary microvessels in vivo. The magnitude of the flow-induced dilation was greatest at the intermediate tone (60 cmH2O IP) but was attenuated at lower and higher IP. After mechanical removal of the endothelium, spontaneous tone and myogenic responses were preserved, but flow-induced dilation and bradykinin-induced dilation were abolished.(ABSTRACT TRUNCATED AT 250 WORDS)

Background Coronary microvessels (<300 μm in diameter) have been demonstrated to be important in the regulation of local resistance and flow. Recent studies also suggest that these microvessels are more responsive to physiological and pharmacological stimuli than conduit vessels. However, little is known regarding the relative sensitivity of different microvascular segments in response to flow (shear stress) and agonists. The goal of this study was to test the hypothesis that a longitudinal gradient for shear stress– and agonist-induced dilation exists in the coronary microcirculation. Methods and Results Experiments were performed in four different sizes of porcine subepicardial coronary arterial microvessels: small arterioles (40±1-μm ID with resting tone); intermediate arterioles (60±1 μm); large arterioles (106±4 μm); and small arteries (179±9 μm). Vessels were isolated and cannulated to allow luminal pressure and flow to be independently controlled. All vessels developed active tone (to ≈65% to 75% of maximum diameter) at their control luminal pressures and showed graded dilations to stepwise increases in shear stress (0 to 10 dynes/cm ² ). For arterioles, the magnitude of the dilations increased as vessel size increased. The highest shear stress produced 21±3%, 32±2%, and 52±5% increases in diameter in small, intermediate, and large arterioles, respectively. Small arteries dilated only 22±6%. The endothelium-dependent vasodilator substance P (SP) produced dose-dependent dilation of all vessels with a threshold at 10 ⁻¹⁶ mol/L. Arterioles were maximally dilated at 10 ⁻⁹ mol/L SP. However, this dose produced only 80% dilation in small arteries. The ED ₅₀ for SP was shifted to the right by two orders of magnitude in small arteries compared with the arterioles. Adenosine preferentially dilated small arterioles, and the dose-response curves shifted to the right for larger vessels. The thresholds for adenosine-induced dilation were 10 ⁻¹² , 10 ⁻¹¹ , and 10 ⁻⁹ mol/L for small, intermediate, and large arterioles, respectively. The endothelium-independent vasodilator nitroprusside produced identical dose-dependent dilations in all vessel segments. Conclusions The results indicate that the pig coronary circulation exhibits a heterogeneity in physiological and pharmacological responses along the microvascular network. Small arterioles are more sensitive to adenosine, but large arterioles are more responsive to shear-stress stimulation. We speculate that site-specific preferential responses may play a crucial role in coordinating overall vascular function in the coronary microvascular network.

It has been reported that free hemoglobin (Hb) reacts with NO at an extremely high rate (K _Hb∼10⁷ m ⁻¹ s⁻¹) and that the red blood cell (RBC) membrane is highly permeable to NO. RBCs, however, react with NO 500–1000 times slower. This reduction of NO reaction rate by RBCs has been attributed to the extracellular diffusion limitation. To test whether additional limitations are also important, we designed a competition test, which allows the extracellular diffusion limitation to be distinguished from transmembrane or intracellular resistance. This test exploited the competition between free Hb and RBCs for NO generated in a homogenous phase by an NO donor. If the extracellular diffusion resistance is negligible, then the results would follow a kinetic model that assumes homogenous reaction without extracellular diffusion limitation. In this case, the measured effective reaction rate constant,K _RBC, would remain invariant of the hematocrit, extracellular-free Hb concentration, and NO donor concentration. Results show that the K _RBC approaches a constant only when the hematocrit is greater than 10%, suggesting that at higher hematocrit, the extracellular diffusion resistance is negligible. Under such a condition, the NO consumption by RBCs is still 500–1000 times slower than that by free Hb. This result suggests that intrinsic RBC factors, such as transmembrane diffusion limitation or intracellular mechanisms, exist to reduce the NO consumption by RBCs.