New in vitro model for proarrhythmia safety screening: IKs inhibition potentiates the QTc prolonging effect of IKr inhibitors in isolated guinea pig hearts.

INTRODUCTION: Preclinical in vivo QT measurement as a proarrhythmia essay is expensive and not reliable enough. The aim of the present study was to develop a sensitive, cost-effective, Langendorff perfused guinea pig heart model for proarrhythmia safety screening. METHODS: Low concentrations of dofetilide and cisapride (inhibitors of the rapid delayed rectifier potassium current, IKr) were tested alone and co-perfused with HMR-1556 (inhibitor of the slow delayed rectifier potassium current, IKs) in Langendorff perfused guinea pig hearts. The electrocardiographic rate corrected QT (QTc) interval, the Tpeak-Tend interval and the beat-to-beat variability and instability (BVI) of the QT interval were determined in sinus rhythm. RESULTS: Dofetilide and HMR-1556 alone or co-perfused, prolonged the QTc interval by 20±2%, 10±1% and 55±10%, respectively. Similarly, cisapride and HMR-1556 alone or co-perfused, prolonged the QTc interval by 11±3%, 11±4% and 38±6%, respectively. Catecholamine-induced fast heart rate abolished the QTc prolonging effects of the IKr inhibitors, but augmented the QTc prolongation during IKs inhibition. None of the drug perfusions increased significantly the Tpeak-Tend interval and the sinus BVI of the QT interval. DISCUSSION: IKs inhibition increased the QTc prolonging effect of IKr inhibitors in a super-additive (synergistic) manner, and the QTc interval was superior to other proarrhythmia biomarkers measured in sinus rhythm in isolated guinea pig hearts. The effect of catecholamines on the QTc facilitated differentiation between IKr and IKs inhibitors. Thus, QTc measurement in Langendorff perfused guinea pig hearts with pharmacologically attenuated repolarization reserve and periodic catecholamine perfusion seems to be suitable for preclinical proarrhythmia screening.

Predictive validity of endpoints used in electrophysiological modelling of migraine in the trigeminovascular system.

The trigeminovascular system has a pivotal role in the pathomechanism of migraine. The aim of the present study was to further develop existing models of migraine making them more suitable for testing the effects of compounds with presumed antimigraine activity in anaesthetised rats. Simultaneous recording of ongoing activity of spontaneously active neurons in the trigeminocervical complex as well as their discharges evoked by electrical stimulation of the dura mater via activation of A- and C-sensory fibres were carried out. Effects of sumatriptan, propranolol and topiramate were evaluated prior to and after application of a mixture containing inflammatory mediators on the dura. Propranolol (10 mg/kg s.c) and topiramate (30 mg/kg s.c.) resulted in a tendency to decrease the level of both spontaneous and evoked activity, while sumatriptan (1 mg/kg s.c.) did not exhibit any effect on recorded parameters. Application of an inflammatory soup to the dura mater boosted up spontaneous activity, which could be significantly attenuated by propranolol and topiramate but not by sumatriptan. In addition, all compounds prevented the delayed increase of spontaneous firing. In contrast to the ongoing activity, evoked responses were not augmented by inflammatory mediators. Nevertheless, inhibitory effect of propranolol and topiramate was evident when considering A- or C-fibre responses. Findings do not support the view that electrically evoked responses are useful for the measurement of trigeminal sensitization. It is proposed however, that inhibition of enhanced firing (immediate and/or delayed) evoked by inflammatory mediators as an endpoint have higher predictive validity regarding the clinical effectiveness of compounds.

Assessment of efficacy of proarrhythmia biomarkers in isolated rabbit hearts with attenuated repolarization reserve.

Isolated hearts with reduced repolarization reserve would be suitable for assessing the proarrhythmic liability of drugs. However, it is not known which proarrhythmia biomarkers indicate the increased susceptibility to torsades de pointes arrhythmia (TdP) in such experimental setting. Thus, we estimated the efficacy of proarrhythmia biomarkers in isolated hearts with attenuated repolarization reserve. Langendorff-perfused rabbit hearts were used. Repolarization reserve was reduced by concomitant inhibition of the rapid (IKr) and slow (IKs) delayed rectifier potassium currents by dofetilide and HMR-1556, respectively. Rate corrected QT (QTc) interval and beat-to-beat variability of the QT interval measured in sinus rhythm or irrespective of rhythm even during arrhythmias (sinus and absolute QT variability, respectively) were tested. QTc failed to predict increased proarrhythmic risk. Sinus QT variability indicated proarrhythmic liability when low concentration of dofetilide was used. However, when arrhythmias compromised sinus variability measurement during coperfusion of catecholamines and elevated concentration of dofetilide, only absolute QT variability indicated increased proarrhythmic risk. Absolute QT variability parameters seem to be the most practical and sensitive biomarkers of proarrhythmic liability in rabbit hearts with reduced repolarization reserve. Absolute QT variability parameters could serve as surrogates for torsades de pointes in drug-safety investigations in isolated rabbit hearts with attenuated repolarization reserve.

Cariprazine (RGH-188), a dopamine D(3) receptor-preferring, D(3)/D(2) dopamine receptor antagonist-partial agonist antipsychotic candidate: in vitro and neurochemical profile.

Cariprazine {RGH-188; trans-N-[4-[2-[4-(2,3-dichlorophenyl)piperazin-1-yl]ethyl]cyclohexyl]-N',N'-dimethylurea hydrochloride}, a novel candidate antipsychotic, demonstrated approximately 10-fold higher affinity for human D(3) versus human D(2L) and human D(2S) receptors (pKi 10.07, 9.16, and 9.31, respectively). It displayed high affinity at human serotonin (5-HT) type 2B receptors (pK(i) 9.24) with pure antagonism. Cariprazine had lower affinity at human and rat hippocampal 5-HT(1A) receptors (pK(i) 8.59 and 8.34, respectively) and demonstrated low intrinsic efficacy. Cariprazine displayed low affinity at human 5-HT(2A) receptors (pK(i) 7.73). Moderate or low affinity for histamine H(1) and 5-HT(2C) receptors (pK(i) 7.63 and 6.87, respectively) suggest cariprazine's reduced propensity for adverse events related to these receptors. Cariprazine demonstrated different functional profiles at dopamine receptors depending on the assay system. It displayed D(2) and D(3) antagonism in [(35)S]GTPgammaS binding assays, but stimulated inositol phosphate (IP) production (pEC(50) 8.50, E(max) 30%) and antagonized (+/-)-quinpirole-induced IP accumulation (pK(b) 9.22) in murine cells expressing human D(2L) receptors. It had partial agonist activity (pEC(50) 8.58, E(max) 71%) by inhibiting cAMP accumulation in Chinese hamster ovary cells expressing human D(3) receptors and potently antagonized R(+)-2-dipropylamino-7-hydroxy-1,2,3,4-tetrahydronaphtalene HBr (7-OH-DPAT)-induced suppression of cAMP formation (pK(b) 9.57). In these functional assays, cariprazine showed similar (D(2)) or higher (D(3)) antagonist-partial agonist affinity and greater (3- to 10-fold) D(3) versus D(2) selectivity compared with aripiprazole. In in vivo turnover and biosynthesis experiments, cariprazine demonstrated D(2)-related partial agonist and antagonist properties, depending on actual dopaminergic tone. The antagonist-partial agonist properties of cariprazine at D(3) and D(2) receptors, with very high and preferential affinity to D(3) receptors, make it a candidate antipsychotic with a unique pharmacological profile among known antipsychotics.

The role of the Na+/Ca2+ exchanger, I(Na) and I(CaL) in the genesis of dofetilide-induced torsades de pointes in isolated, AV-blocked rabbit hearts.

BACKGROUND AND PURPOSE: The Na+/Ca2+ exchanger (NCX) may contribute to triggered activity and transmural dispersion of repolarization, which are substrates of torsades de pointes (TdP) type arrhythmias. This study examined the effects of selective inhibition of the NCX by SEA0400 on the occurrence of dofetilide-induced TdP. EXPERIMENTAL APPROACH: Effects of SEA0400 (1 micromol x L(-1)) on dofetilide-induced TdP was studied in isolated, Langendorff-perfused, atrioventricular (AV)-blocked rabbit hearts. To verify the relevance of the model, lidocaine (30 micromol x L(-1)) and verapamil (750 nmol x L(-1)) were also tested against dofetilide-induced TdP. KEY RESULTS: Acute AV block caused a chaotic idioventricular rhythm and strikingly increased beat-to-beat variability of the RR and QT intervals. SEA0400 exaggerated the dofetilide-induced increase in the heart rate-corrected QT interval (QTc) and did not reduce the incidence of dofetilide-induced TdP [100% in the SEA0400 + dofetilide group vs. 75% in the dofetilide (100 nmol x L(-1)) control]. In the second set of experiments, verapamil further increased the dofetilide-induced QTc prolongation and neither verapamil nor lidocaine reduced the dofetilide-induced increase in the beat-to-beat variability of the QT interval. However, lidocaine decreased and verapamil prevented the development of dofetilide-induced TdP as compared with the dofetilide control (TdP incidence: 13%, 0% and 88% respectively). CONCLUSIONS AND IMPLICATIONS: Na+/Ca2+ exchanger does not contribute to dofetilide-induced TdP, whereas Na+ and Ca2+ channel activity is involved in TdP genesis in isolated, AV-blocked rabbit hearts. Neither QTc prolongation nor an increase in the beat-to-beat variability of the QT interval is a sufficient prerequisite of TdP genesis in rabbit hearts.

Importance of extracardiac alpha1-adrenoceptor stimulation in assisting dofetilide to induce torsade de pointes in rabbit hearts.

In anaesthetized rabbits, alpha(1)-adrenoceptor stimulation increases the propensity of repolarization-prolonging drugs to induce torsade de pointes ventricular tachycardia. However, it is not known whether the stimulation of intracardiac alpha(1)-adrenoceptors, or the increased ventricular stretch caused by extracardiac alpha(1)-adrenoceptor-mediated peripheral vasoconstriction and increased resistance, are the sensitizing factors. Accordingly, this study investigated whether a sustained load-induced left ventricular stretch or stimulation of the intracardiac alpha(1)-adrenoceptors with 100 nM methoxamine, or the co-application of these two, can assist dofetilide (100 nM) to elicit torsade de pointes in isolated Langendorff-perfused, rabbit hearts. In the stretched hearts, a constant high level of stretch was produced by a water-filled left ventricular balloon inflated to a volume of 1.4 ml, whereby the systolic and end-diastolic pressures virtually did not exceed the physiological range (