␤-Adrenergic receptors (␤-ARs) are members of the superfamily of G-protein-coupled receptors that mediate the effects of catecholamines in the sympathetic nervous system. Three distinct ␤-AR subtypes have been identified (␤1-AR, ␤2-AR, and ␤3-AR). In order to define further the role of the different ␤-AR subtypes, we have used gene targeting to inactivate selectively the ␤2-AR gene in mice. Based on intercrosses of heterozygous knockout (␤2-AR ؉/؊) mice, there is no prenatal lethality associated with this mutation. Adult knockout mice (␤2-AR ؊/؊) appear grossly normal and are fertile. Their resting heart rate and blood pressure are normal, and they have a normal chronotropic response to the ␤-AR agonist isoproterenol. The hypotensive response to isoproterenol, however, is significantly blunted compared with wild type mice. Despite this defect in vasodilation, ␤2-AR ؊/؊ mice can still exercise normally and actually have a greater total exercise capacity than wild type mice. At comparable workloads, ␤2-AR ؊/؊ mice had a lower respiratory exchange ratio than wild type mice suggesting a difference in energy metabolism. ␤2-AR ؊/؊ mice become hypertensive during exercise and exhibit a greater hypertensive response to epinephrine compared with wild type mice. In summary, the primary physiologic consequences of the ␤2-AR gene disruption are observed only during the stress of exercise and are the result of alterations in both vascular tone and energy metabolism.

Endothelial surface expression of P-selectin and subsequent leukocyte rolling in venules can be induced by mast cell-derived histamine and binding of thrombin to protease-activated receptor-1 (PAR1). We hypothesized that activation of endothelial PAR2 by mast cell tryptase or other proteases also contributes to inflammatory responses. Leukocyte rolling flux and rolling velocity were assessed by intravital microscopy of the cremaster muscles of wild-type mice following perivenular micropipette injections of a control (LSIGRL) or PAR2-activating (SLIGRL) oligopeptide. Injection of SLIGRL increased mean rolling leukocyte flux fraction from 34 ± 11 to 71 ± 24% (p < 0.05) and decreased mean rolling velocity from 63 ± 29 to 32 ± 2 μm/s (p < 0.05). No significant changes occurred with control peptide injection. To further evaluate the role of PAR2 in inflammatory responses, PAR2-deficient mice were generated by gene targeting and homologous recombination. Perivenular injections of SLIGRL resulted in only a small increase in rolling leukocyte flux fraction (from 21 ± 8 to 30 ± 2%) and no change in rolling velocity. Leukocyte rolling after surgical trauma was assessed in 9 PAR2-deficient and 12 wild-type mice. Early (0–15 min) after surgical trauma, the mean leukocyte rolling flux fraction was lower (10 ± 3 vs 30 ± 6%, p < 0.05) and mean rolling velocity was higher (67 ± 46 vs 52 ± 36 μm/s, p < 0.01) in PAR2-deficient compared with control mice. The defect in leukocyte rolling in PAR2-deficient mice did not persist past 30 min following surgical trauma. These results indicate that activation of PAR2 produces microvascular inflammation by rapid induction of P-selectin-mediated leukocyte rolling. In the absence of PAR2, the onset of inflammation is delayed.

The activation state of ␤-adrenergic receptors (␤-ARs) in vivo is an important determinant of hemodynamic status, cardiac performance, and metabolic rate. In order to achieve homeostasis in vivo, the cellular signals generated by ␤-AR activation are integrated with signals from a number of other distinct receptors and signaling pathways. We have utilized genetic knockout models to test directly the role of ␤1-and/or ␤2-AR expression on these homeostatic control mechanisms. Despite total absence of ␤1-and ␤2-ARs, the predominant cardiovascular ␤-adrenergic subtypes, basal heart rate, blood pressure, and metabolic rate do not differ from wild type controls. However, stimulation of ␤-AR function by ␤-AR agonists or exercise reveals significant impairments in chronotropic range, vascular reactivity, and metabolic rate. Surprisingly, the blunted chronotropic and metabolic response to exercise seen in ␤1/ ␤2-AR double knockouts fails to impact maximal exercise capacity. Integrating the results from single ␤1-and ␤2-AR knockouts as well as the ␤1-/␤2-AR double knockout suggest that in the mouse, ␤-AR stimulation of cardiac inotropy and chronotropy is mediated almost exclusively by the ␤1-AR, whereas vascular relaxation and metabolic rate are controlled by all three ␤-ARs (␤1-, ␤2-, and ␤3-AR). Compensatory alterations in cardiac muscarinic receptor density and vascular ␤3-AR responsiveness are also observed in ␤1-/␤2-AR double knockouts. In addition to its ability to define ␤-AR subtype-specific functions, this genetic approach is also useful in identifying adaptive alterations that serve to maintain critical physiological setpoints such as heart rate, blood pressure, and metabolic rate when cellular signaling mechanisms are perturbed.

Manipulations of the murine genome that alter cardiovascular function have created the need for methods to study cardiovascular physiology in genetically altered animals in vivo. We adapted chronic physiological measurement techniques to the nonanesthetized, nonrestrained murine model, established strain-specific cardiovascular and metabolic norms, and evaluated responses to anesthesia, exercise, and adrenergic stimulation. Anesthesia resulted in alterations in heart rate (HR), blood pressure (BP), and O2 consumption (V(O2)) and CO2 production (V(CO2)) for up to 6 h postoperatively. There were significant interstrain differences in resting values of HR and BP Graded treadmill exercise resulted in linear increases in HR, V(O2), V(CO2), and respiratory exchange ratio (RER) similar to those seen in larger species. Response to beta-adrenergic stimulation showed a classic sigmoidal dose-response curve; however, there was very little tachycardiac response to vagal blockade, indicating low resting vagal tone. This study demonstrates the feasibility of performing chronic cardiovascular measurements in nonanesthetized mice and stresses the importance of allowing for anesthetic recovery and strain variability. Murine cardiovascular responses to exercise can be reliably measured and are qualitatively similar to those in humans.