Predicting in vivo cardiovascular properties of β-blockers from cellular assays: a quantitative comparison of cellular and cardiovascular pharmacological responses

Abstract

β-Adrenoceptor antagonists differ in their degree of partial agonism. In vitro assays have provided information on ligand affinity, selectivity, and intrinsic efficacy. However, the extent to which these properties are manifest in vivo is less clear. Conscious freely moving rats, instrumented for measurement of heart rate (β1; HR) and hindquarters vascular conductance (β2; HVC) were used to measure receptor selectivity and ligand efficacy in vivo. CGP 20712A caused a dose-dependent decrease in basal HR (P<0.05, ANOVA) at 5 doses between 6.7 and 670 μg/kg (i.v.) and shifted the dose-response curve for isoprenaline to higher agonist concentrations without altering HVC responses. In contrast, at doses of 67 μg/kg (i.v.) and above, ICI 118551 substantially reduced the HVC response to isoprenaline without affecting HR responses. ZD 7114, xamoterol, and bucindolol significantly increased basal HR (ΔHR: +122 ± 12, + 129 ± 11, and + 59 ± 11 beats/min, respectively; n=6), whereas other β-blockers caused significant reductions (all at 2 mg/kg i.v.). The agonist effects of xamoterol and ZD 7114 were equivalent to that of the highest dose of isoprenaline. Bucindolol, however, significantly antagonized the response to the highest doses isoprenaline. An excellent correlation was obtained between in vivo and in vitro measures of β1-adrenoceptor efficacy (R(2)=0.93; P<0.0001).

Figure 1.

Inhibition of ³H-CGP 12177-specific binding in whole CHO cells expressing human β1-adrenoceptor (A) and human β2-adrenoceptor (B). Nonspecific binding was determined by 10 μM propranolol. Concentration of ³H-CGP 12177 present was 0.97 nM (A) and 1.11 nM (B). Data points are means ± se of triplicate determinations. Results of single experiments shown are representative of 8 separate experiments in each case.

Figure 2.

³H-cAMP accumulation in CHO cells expressing the human β1-adrenoceptor in response to xamoterol, ZD 7114, pronethalol, acebutolol and bisoprolol (A) and bucindolol (B). Bars represent basal ³H-cAMP accumulation and that in response to 10 μM isoprenaline. Data points are means ± se of triplicate determinations. Results are representative of 3 (A) and 6 (B) separate experiments.

Figure 3.

Absolute values for HR (top panel) and HVC (bottom panel) in conscious, atropine-treated, freely moving rats in response to isoprenaline (at 0, 4, 12, 40, and 120 ng/kg/min) before and after administration of CGP 20712A at doses of 0–670 μg/kg bolus, 0–333 μg/kg/h infusion. Values (mean±se; n=6) were measured at baseline (B; i.e., in the absence of isoprenaline) and at the end of 3-min infusions of isoprenaline.

Figure 4.

Absolute values for HR (top panel) and HVC (bottom panel) in conscious, atropine-treated, freely moving rats in response to isoprenaline (at 0, 4, 12, 40, and 120 ng/kg/min) before and after administration of ICI 118551 at doses of 0–670 μg/kg bolus and 0–333 μg/kg/h infusion. Values (means±se; n=6) were measured at baseline (B; i.e., in the absence of isoprenaline) and at the end of 3-min infusions of isoprenaline.

Figure 5.

Absolute values for HR (top panel) and HVC (bottom panel) in conscious, atropine-treated, freely moving rats in response to isoprenaline (at 0, 4, 12, 40, 120 ng/kg/min) following administration of bisoprolol at doses of 0–2000 μg/kg bolus, 0–1000 μg/kg/h infusion. Values (means±se; n=6) were measured at baseline (B; i.e., in the absence of isoprenaline) and at the end of 3-min infusions of isoprenaline.

Figure 6.

Absolute values for HR (top panel) and HVC (bottom panel) in conscious, atropine-treated, freely moving rats in response to isoprenaline (at 0, 4, 12, 40, and 120 ng/kg/min) following administration of saline and several different β-blockers (2 mg/kg bolus, 1 mg/kg/h infusion). Values (means±se; n=6) were measured at baseline (B; i.e., in the absence of isoprenaline) and at the end of 3-min infusions of isoprenaline.

Figure 7.

Absolute values for HR (top panel) and HVC (bottom panel) in conscious, atropine-treated, freely moving rats in response to isoprenaline (at 0, 4, 12, 40, and 120 ng/kg/min) following administration of vehicle (5% propylene glycol, 2% Tween 80 in saline), bucindolol, carvedilol, and nebivolol (2 mg/kg bolus, 1 mg/kg/h infusion). Values (means±se; n=6) were measured at baseline (B; i.e., in the absence of isoprenaline) and at the end of 3-min infusions of isoprenaline.

Figure 8.

Correlation plot of the efficacy of β-blockers for the in vitro vs. in vivo data. In vitro efficacy is the maximum ³H-cAMP accumulation response (given as percentage of the maximum response to isoprenaline) in cells expressing the human β1-adrenoceptor. The in vivo efficacy is the change in HR (beats/min) in response to the administration of the β-blocker alone in conscious freely moving rats (ISA). Ligands are at 2 mg/kg bolus, 1 mg/kg/h infusion except for CGP 20712A and ICI 118551, which were at 670 μg/kg bolus, 333 μg/kg/h infusion. For ligands with a 2-component ³H-cAMP accumulation response (bucindolol and carvedilol), efficacy is given as the maximum efficacy obtained for that ligand (i.e., includes response at site 1 plus that at site 2). Line is that of best fit through all the ligands.