B2 bradykinin receptors in cultured neonatal rat cardiomyocytes mediate a negative chronotropic and negative inotropic response.

Abstract

Receptors for bradykinin (BK) were characterized in primary cultures of beating neonatal rat cardiomyocytes using [3H]BK was radioligand. Degradation studies demonstrated that [3H]BK was stable for at least 2 h when incubated with cardiomyocytes at 2 and 37 degrees C in the presence of bacitracin in combination with captopril or ramiprilat. Without these inhibitors, > 80% of the [3H]BK was degraded within 2 h at 37 degrees C. This indicates that angiotensin-converting enzyme (ACE) is responsible for the main BK-degrading activity in cardiomyocytes. Scatchard plots were linear and gave a Kd of 1.5 +/- 0.8 nmol/l (mean +/- SD, n = 4) and a maximum binding capacity of 55-125 fmol/mg protein. Association and dissociation studies showed that binding of [3H]BK was saturable and reversible. Binding of [3H]BK at 37 degrees C led to internalization of the ligand. Competition studies with B1 and B2 agonists and antagonists were consistent with a B2 subtype of receptor. Addition of BK to beating cardiomyocytes (> 1 nmol/l) at 37 degrees C gave a strong but transient negative chronotropic effect. This response was paralleled by changes in the pulsation amplitude, which indicated a simultaneous negative inotropic effect of BK. These results provide a basis for the hypothesis that ACE inhibition exerts its cardioprotective effect at the level of a population of cardiomyocytes by virtue of kinin receptor-mediated mechanisms.