Vessel diameter was measured with a video caliper (Colorado Video, Boulder, CO, USA). Vessels without leaks were allowed to develop spontaneous tone (≥17% less initial diameter). Ca++-free PSS was superfused at the end of all experiments to determine passive arteriolar diameters. Compounds were introduced via a syringe pump and at concentrations that previously described elsewhere [24]. A23187, a calcium ionophore, was this website introduced into the lumen of the arterioles at a concentration of 1 μm, as previously described [36]. l-NMMA (Calbiochem,
Gibbstown, NJ, USA) was used at a final tissue bath concentration of 0.1 mm to competitively inhibit NOS activity. The superfusate concentration of phentolamine, an α-adrenergic receptor blocker, was 1 μm. ADO was superfused at the end of all experiments (0.1 mm) to determine passive arteriolar diameters. Compounds were added directly to the superfusate solution as previously described [26, 27]. ACh, Spermine NONOate,
and PE, were added at increasing concentrations of 0.001–100 μm or A23187 1–1000 nm. All chemicals were from Sigma (St. Louis, MO, USA), unless otherwise noted. Arteriolar diameter, D (μm), was recorded during a control, BMS-777607 purchase intraluminal infusion or PVNS period and immediately following AH. Resting vascular tone was calculated by: %tone = [(Dpass − Dc)/Dpass] × 100, where Dpass is passive diameter under ADO and Dc is the diameter measured during the control period. Arteriolar responses were normalized as follows: percent change from control = [(Dss/Dc) − 1] × 100, where Dss is the steady-state diameter following intraluminal infusion, AH, and PVNS. Dc immediately prior to the beginning of any experimental procedure was used to calculate %tone and reported as
0 PSI diameter measurements in the A23187 experimental series. All data are reported as mean ± SE. Spontaneous tone was calculated by: % tone = [(Dpass − DI)/Dpass] × 100, where Dpass is the maximal diameter recorded under Ca++ free PSS for coronary or mesenteric arterioles, respectively. DI is the initial diameter of the arteriole O-methylated flavonoid prior to the experimental period. Active responses to pressure changes were normalized to the maximal diameter according to the following formula: % Normalized diameter = [(Dss/Dpass)] × 100, Dss is the steady-state diameter during each pressure step. The experimental responses to ACh, A23187, and Spermine NONOate are expressed using the following equation: % relaxation = [(Dss − Dcon)/(Dpass − Dcon)] × 100, where Dss is the steady-state arteriolar diameter during the experimental period, Dcon is the control diameter recorded immediately prior to experimental period. Responses to PE were calculated by the following formula: % constriction = [(Dss − Dcon)/(Dcon)] × 100.
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