β-blockade abolishes the augmented cardiac tPA release induced by transactivation of heterodimerised bradykinin receptor-2 and β₂ -adrenergic receptor in vivo

Bradykinin (BK) receptor-2 (B₂R) and β₂-adrenergic receptor (β₂AR) have been shown to form heterodimers in vitro. However, in vivo proofs of the functional effects of B₂R-β₂AR heterodimerisation are missing. Both BK and adrenergic stimulation are known inducers of tPA release. Our goal was to demonstrate the existence of B₂R-β₂AR heterodimerisation in myocardium and to define its functional effect on cardiac release of tPA in vivo. We further investigated the effects of a non-selective β-blocker on this receptor interplay. To investigate functional effects of B₂R-β₂AR heterodimerisation (i. e. BK transactivation of β₂AR) in vivo, we induced serial electrical stimulation of cardiac sympathetic nerves (SS) in normal pigs that underwent concomitant BK infusion. Both SS and BK alone induced increases in cardiac tPA release. Importantly, despite B₂R desensitisation, simultaneous BK infusion and SS (BK+SS) was characterised by 2.3 ± 0.3-fold enhanced tPA release compared to SS alone. When β-blockade (propranolol) was introduced prior to BK+SS, tPA release was inhibited. A persistent B₂R-β₂AR heterodimer was confirmed in BK-stimulated and nonstimulated left ventricular myocardium by immunoprecipitation studies and under non-reducing gel conditions. All together, these results strongly suggest BK transactivation of β₂AR leading to enhanced β₂AR-mediated release of tPA. Importantly, non-selective β-blockade inhibits both SS-induced release of tPA and the functional effects of B₂R-β₂AR heterodimerisation in vivo, which may have important clinical implications.