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.