Inhibition of Thromboxane A₂-Induced Arrhythmias and Intracellular Calcium Changes in Cardiac Myocytes by Blockade of the Inositol Trisphosphate Pathway

Abstract

We have recently reported that left atrial injections of the thromboxane A 2 (TXA 2 ) mimetic, (5Z)-7-[(1R,4S,5S,6R)-6-[(1E,3S)-3-hydroxy-1-octenyl]-2 -oxabicyclo[2.2.1]hept-5-yl]-5-heptenoic acid (U46619), induced ventricular arrhythmias in the anesthetized rabbit. Data from this study led us to hypothesize that TXA 2 may be inducing direct actions on the myocardium to induce these arrhythmias. The aim of this study was to further elucidate the mechanism responsible for these arrhythmias. We report that TXA 2 R is expressed at both the gene and protein levels in atrial and ventricular samples of adult rabbits. In addition, TXA 2 R mRNA was identified in single, isolated ventricular cardiac myocytes. Furthermore, treatment of isolated cardiac myocytes with U46619 increased intracellular calcium in a dose-dependent manner and these increases were blocked by the specific TXA 2 R antagonist, 7-(3-((2-((phenylamino)carbonyl)-hydrazino)methyl)-7-oxabicyclo(2.2.1)hept-2-yl)-5-heptenoic acid (SQ29548). Pretreatment of myocytes with an inhibitor of inositol trisphosphate (IP 3 ) formation, gentamicin, or with an inhibitor of IP 3 receptors, 2-aminoethoxydiphenylborate (2-APB), blocked the increase in intracellular calcium. In vivo pretreatment of anesthetized rabbits with either gentamicin or 2-APB subsequently inhibited the formation of ventricular arrhythmias elicited by U46619. These data support the hypothesis that TXA 2 can induce arrhythmias via a direct action on cardiac myocytes. Furthermore, these arrhythmogenic actions were blocked by inhibitors of the IP 3 pathway. In summary, this study provides novel evidence for direct TXA 2 -induced cardiac arrhythmias and provides a rationale for IP 3 as a potential target for the treatment of TXA 2 -mediated arrhythmias.

Store-operated calcium entry is present in HL-1 cardiomyocytes and contributes to resting calcium

“…Protein extraction, quantification, and subsequent Western blotting using STIM1 (BD Laboratories) and Orai1 (Millipore) antibodies were performed as previously described [18]. …”

FGF23 is a novel regulator of intracellular calcium and cardiac contractility in addition to cardiac hypertrophy

“…Clamp-frozen ventricular tissues were weighed and homogenized in a 12:1 (vol/wt) ratio of ice-cold cell extraction buffer (Invitrogen), as described previously (56). Total protein concentration of the samples was determined by use of the microbicinchoninic acid protein assay (Pierce Chemical) and then normalized to tissue weight (g/mg).…”

“…Ca 2ϩ imaging. Imaging during FGF23 perfusion was conducted as reported previously (50,51,56 , and verapamil (10 M ϩ FGF23)] were tested five times, which totaled six to 10 cardiomyocytes from a given animal, which was repeated in three to five different animals. The fluorescent changes from each cell were averaged, and the data were grouped by animal and then used for data analysis.…”

International Union of Basic and Clinical Pharmacology. LXXXIII: Classification of Prostanoid Receptors, Updating 15 Years of Progress

“…Cardiovascular TP receptor expression even extends to cardiomyocytes, and the TxA 2 mimetic U-46619 (Fig. 10) causes arrhythmia (Wacker et al, 2009). TP receptors are also located on afferent sympathetic nerve endings in the heart and may participate in the sympathoexcitatory reflex that occurs during myocardial ischemia (Fu et al, 2008).…”