Effect of ivabradine and endotoxin on human If

Background: Endotoxins (lipopolysaccharides, LPSs) trigger the development of sepsis by gram negative bacteria and cause a variety of biological effects on host cells including alterations in ionic channels. Recently we published that LPS impairs the pacemaker current I f in single cardiomyocytes (after 6 h incubation) isolated from human right atrial appendages. The LPS impairment of I f in human heart was discussed to be relevant for understanding the reduction in heart rate variability observed in human sepsis and endotoxemia. Aims: Based on our findings of I f impairment by endotoxin we want to address two main questions in this study using human cardiomyocytes isolated from right atrial appendages. 1.) Interference of Ivabradine and LPS on I f Ivabradine is a novel I f blocker, prescribed to patients with stable angina pectoris with a contraindication of beta- blockers. Medication of ivabradine is also discussed in cardiac diseases, where a reduction of heart rate is thought to be beneficial. What happens when ivabradine is administered to patients with concomitant elevated endotoxin levels? Due to a cooperation we have the exceptional opportunity to obtain ivabradine (provided by Servier) which enables us to study the interference of this blocker and endotoxin on the human cellular level. 2.) Mechanism of LPS action on I f Further experiments will provide substantial insight if the effect of LPS on I f is mediated by an intracellular signal (e.g. decrease in intracellular cAMP level). Different endotoxin variants and an antagonist will allow us to detect the "minimal" endotoxin structure necessary to affect I f and to elucidate if the action of LPS on I f is in line with current hypothesis of biological LPS action. Membrane properties may be altered due to interaction of endotoxin with the cardiac plasmamembrane affecting thereby ionic channel properties. LPS induced differences in local motional restriction and order in lipid bilayer will be assessed using spin labelling technique. Experimental studies on I f will be accompanied by mathematical modelling to explain the role of alterations in pacemaker channel activity for single cell excitability.

Although sepsis is among the leading causes of death (third highest in Austria and Germany) it is a rather unknown disease in general population. It can be defined as a systemic inflammatory response that occurs during infections induced by microorganisms and may lead to death due to a massive drop of blood pressure and multiple organ failure. Recently, we showed that endotoxin (lipopolysaccharide, LPS), the trigger of gram-negative sepsis impairs the human cardiac pacemaker current (If). This ionic current plays an important role for generation and autonomic modulation of sinoatrial pacemaker activity and hence heart rate control. Ivabradine, a new selective (If)-blocker, is currently investigated whether it improves survival in septic patients by lowering heart rate. In the scope of the project we further investigated the effect of LPS on the pacemaker current and showed that the impairment of (If) by LPS is caused by a direct interaction of a specific entity of the molecule, the so-called O-chain, with the pacemaker channel proteins. Thus, we discovered a new biological mode of LPS-action which is not receptor-mediated as described so far. A central aim of this study was to clarify whether ivabradine is still capable to block the pacemaker current under elevated endotoxin levels (septic conditions). Using a special electrophysiological method (patch-clamp technique) we investigated the human pacemaker current in cardiomyocytes isolated from the right atrial appendage. The efficacy of ivabradine to block (If) was reduced under septic conditions, an observation that correlated well with lowered intracellular ivabradine concentrations in LPS-treated myocytes as detected by high pressure liquid chromatography. Computational analysis using a sinoatrial pacemaker cell model revealed that despite the reduction of (If) under septic conditions, ivabradine was able to further decelerate pacemaking activity. This novel finding, i.e. (If)-inhibition by ivabradine under elevated endotoxin levels on the cellular level, may provide a molecular understanding for the efficacy of this drug on heart rate reduction in the septic patient. Thus, the medical application of ivabradine as heart rate reducing agent in critically ill patients (severe sepsis and septic shock) might favour a better therapeutic outcome.