Effects of bradykinin on intracellular calcium transients in cardiac myocytes

In spite many evidences has supported the cardioprotective effect of bradykinin, its direct effects at the cell level are still under question. We investigated the both effects of bradykinin (BK) on Ca2+-related ionic currents using whole cell voltage clamp technique in rabbit cardiomyocytes and on the intracellular Ca2+ transient using calcium sensitive fluorescence dye, indo-1AM. Simultaneously with recording intracellular Ca2+ transients, cell contractility was estimated from the changes in length of the electrical stimulated rat cardiac myocytes. L-type Ca2+ current decreased by bradykinin at the entire voltage range. Inward tail current increased initially up to its maximum about 4 min after exposing myocytes to BK, and then gradually decreased again by further exposure to BK. This tail current decreased remarkably at washing BK off but slowly recovered ca. 20 min later. The change in cell contractility was similar to that in tail current showing initial increase followed by gradual decrease. Removal of BK brought remarkable decrease in contractility, which was recovered 15~20 min after cessation of electrical stimulation. Bradykinin increased Ca2+ transient initially but after some time Ca2+ transient also decreased coincidentally with contractility. From these results, it is suggested that bradykinin exerts directly its cardioprotective effect on the single myocytes by decreasing the intracellular Ca2+ level followed by an initial increase in Ca2+ transient.

Modulation of ATP-induced activation of the muscarinic K+ channel activity by protein kinase C

The atrial acetylcholine-activated K+ (KACh) channel is gated by the pertussis toxin-sensitive inhibitory G (GK) protein. Earlier studies revealed that ATP alone can activate the KACh channel via transphosphorylation mediated by nucleoside-diphosphate kinase (NDPK) in atrial cells of rabbit and guinea pig. This channel can be activated by various agonists and also modulated its function by phosphorylation. ATP-induced KACh channel activation (AIKA) was maintained in the presence of the NDPK inhibitor, suggesting the existence of a mechanism other than NDPK-mediated process. Here we hypothesized the phosphorylation process as another mechanism underlying AIKA and was undertaken to examine what kinase is involved in atrial cells isolated from the rat heart. Single application of 1 mM ATP gradually increased the activity of KACh channels and reached its maximum 40 ~ 50 sec later following adding ATP. AIKA was not completely reduced but maintained by half even in the presence of NDPK inhibitor. Neither ADP nor a non-hydrolyzable ATP analogue, AMP-PNP can cause AIKA, while a non-specific phosphatase, alkaline phosphatase blocked completely AIKA. PKC antagonists such as sphingosine or tamoxifen, completely blocked AIKA, whereas PKC catalytic domain increased AIKA. Taken together, it is suggested that the PKC-mediated phosphorylation is partly involved in AIKA.