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Cryothermy has potential advantages over RF energy for catheter ablation, including reversibility of lesion formation, catheter stability, and less procedural discomfort. Cryoablation procedures were performed in 14 patients with atrioventricular reentrant tachycardias (AVNRTs), 13 patients with accessory pathway (AP)-mediated tachycardias, and 5 patients with atrial fibrillation. The numbers of energy applications, pain scores, procedural times, and outcomes were recorded and compared with age- and sex-matched patients undergoing similar RF procedures. Cryoablation was successful in 26 of 32 patients (11/14 AVNRT, 10/13 AP, 5/5 AF) compared with 30 of 32 undergoing RF procedures, with similar numbers of energy applications and procedural times. Cryothermy was painless in all patients, and the overall procedural discomfort was significantly less than in patients treated with RF (1.3 +/- 2.2 vs 6.1 +/- 3.5). In patients with anteroseptal pathways, cryomapping successfully identified safe sites to target the delivery of energy. Cryothermy is a painless and safe alternative to RF. It may be particularly useful for catheter ablation of patients with pathways close to the atrioventricular node. [ABSTRACT FROM AUTHOR]

The thalamus has long been recognized for its role in relaying sensory information from the periphery, a function accomplished by its "first-order" nuclei. However, a second category of thalamic nuclei, termed "higher-order" nuclei, have been shown instead to mediate communication between cortical areas. The nucleus reuniens of the midline thalamus (RE) is a higher-order nucleus known to act as a conduit of reciprocal communication between the medial prefrontal cortex (mPFC) and hippocampus. While anatomical and behavioural studies of RE are numerous, circuit-based electrophysiological studies, particularly those examining the impact of cortical input and the thalamic reticular nucleus (TRN) on RE neuron firing, are sparse. To characterize RE neuron firing properties and dissect the circuit dynamics of the infralimbic subdivision of the mPFC (ilPFC), the TRN and RE, we used in vivo, extracellular, single-unit recordings in male Sprague Dawley rats and manipulated neural activity using targeted pharmacological manipulations, electrical stimulation and a projection-specific implementation of designer receptors exclusively activated by designer drugs (DREADDs). We show that ilPFC inhibition reduces multiple burst firing parameters in RE, whereas ilPFC stimulation drives burst firing and dampens tonic firing. In addition, TRN inhibition reduces the number of spontaneously active neurons in RE. Finally, inhibition of ilPFC terminals in RE selectively enhances a subset of burst firing parameters. These findings demonstrate that ilPFC input, both via direct projections and via the TRN, can modulate RE neuron firing pattern in nuanced and complex ways. They also highlight the ilPFC-TRN-RE circuit as a likely critical component of prefrontal-hippocampal interactions., (© 2018 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Acute RyR2 activation by exchange protein directly activated by cAMP (Epac) reversibly perturbs myocyte Ca 2+ homeostasis, slows myocardial action potential conduction, and exerts pro-arrhythmic effects. Loose patch-clamp studies, preserving in vivo extracellular and intracellular conditions, investigated Na + current in intact cardiomyocytes in murine atrial and ventricular preparations following Epac activation. Depolarising steps to varying test voltages activated typical voltage-dependent Na + currents. Plots of peak current against depolarisation from resting potential gave pretreatment maximum atrial and ventricular currents of -20.23 ± 1.48 (17) and -29.8 ± 2.4 (10) pA/μm 2 (mean ± SEM [n]). Challenge by 8-CPT (1 μmol/L) reduced these currents to -11.21 ± 0.91 (12) (P < .004) and -19.3 ± 1.6 (11) pA/μm 2 (P < .04) respectively. Currents following further addition of the RyR2 inhibitor dantrolene (10 μmol/L) (-19.91 ± 2.84 (13) and -26.6 ± 1.7 (17)), and dantrolene whether alone (-19.53 ± 1.97 (8) and -27.6 ± 1.9 (14)) or combined with 8-CPT (-19.93 ± 2.59 (12) and -29.9 ± 2.5(11)), were indistinguishable from pretreatment values (all P >> .05). Assessment of the inactivation that followed by applying subsequent steps to a fixed voltage 100 mV positive to resting potential gave concordant results. Half-maximal inactivation voltages and steepness factors, and time constants for Na + current recovery from inactivation in double-pulse experiments, were similar through all the pharmacological conditions. Intracellular sharp microelectrode membrane potential recordings in intact Langendorff-perfused preparations demonstrated concordant variations in maximum rates of atrial and ventricular action potential upstroke, (dV/dt) max . We thus demonstrate an acute, reversible, Na + channel inhibition offering a possible mechanism for previously reported pro-arrhythmic slowing of AP propagation following modifications of Ca 2+ homeostasis, complementing earlier findings from chronic alterations in Ca 2+ homeostasis in genetically-modified RyR2-P2328S hearts., (© 2017 The Authors. Clinical and Experimental Pharmacology and Physiology Published by John Wiley & Sons Australia, Ltd.)

Increasing evidence implicates chronic energetic dysfunction in human cardiac arrhythmias. Mitochondrial impairment through Pgc-1β knockout is known to produce a murine arrhythmic phenotype. However, the cumulative effect of this with advancing age and its electrocardiographic basis have not been previously studied. Young (12-16 weeks) and aged (>52 weeks), wild type (WT) (n = 5 and 8) and Pgc-1β -/- (n = 9 and 6), mice were anaesthetised and used for electrocardiographic (ECG) recordings. Time intervals separating successive ECG deflections were analysed for differences between groups before and after β1-adrenergic (intraperitoneal dobutamine 3 mg/kg) challenge. Heart rates before dobutamine challenge were indistinguishable between groups. The Pgc-1β -/- genotype however displayed compromised nodal function in response to adrenergic challenge. This manifested as an impaired heart rate response suggesting a functional defect at the level of the sino-atrial node, and a negative dromotropic response suggesting an atrioventricular conduction defect. Incidences of the latter were most pronounced in the aged Pgc-1β -/- mice. Moreover, Pgc-1β -/- mice displayed electrocardiographic features consistent with the existence of a pro-arrhythmic substrate. Firstly, ventricular activation was prolonged in these mice consistent with slowed action potential conduction and is reported here for the first time. Additionally, Pgc-1β -/- mice had shorter repolarisation intervals. These were likely attributable to altered K + conductance properties, ultimately resulting in a shortened QT c interval, which is also known to be associated with increased arrhythmic risk. ECG analysis thus yielded electrophysiological findings bearing on potential arrhythmogenicity in intact Pgc-1β -/- systems in widespread cardiac regions., (© 2017 The Authors. Clinical and Experimental Pharmacology and Physiology Published by John Wiley & Sons Australia, Ltd.)

Ageing is associated with increased prevalences of both atrial and ventricular arrhythmias, reflecting disruption of the normal sequence of ion channel activation and inactivation generating the propagated cardiac action potential. Experimental models with specific ion channel genetic modifications have helped clarify the interacting functional roles of ion channels and how their dysregulation contributes to arrhythmogenic processes at the cellular and systems level. They have also investigated interactions between these ion channel abnormalities and age-related processes in producing arrhythmic tendency. Previous reviews have explored the relationships between age and loss-of-function Na v 1.5 mutations in producing arrhythmogenicity. The present review now explores complementary relationships arising from gain-of-function Na v 1.5 mutations associated with long QT3 (LQTS3). LQTS3 patients show increased risks of life-threatening ventricular arrhythmias, particularly after 40 years of age, consistent with such interactions between the ion channel abnormailities and ageing. In turn clinical evidence suggests that ageing is accompanied by structural, particularly fibrotic, as well as electrophysiological change. These abnormalities may result from biochemical changes producing low-grade inflammation resulting from increased production of reactive oxygen species and superoxide. Experimental studies offer further insights into the underlying mechanisms underlying these phenotypes. Thus, studies in genetically modified murine models for LQTS implicated action potential recovery processes in arrhythmogenesis resulting from functional ion channel abnormalities. In addition, ageing wild type (WT) murine models demonstrated both ion channel alterations and fibrotic changes with ageing. Murine models then suggested evidence for interactions between ageing and ion channel mutations and provided insights into potential arrhythmic mechanisms inviting future exploration., (© 2017 The Authors. Clinical and Experimental Pharmacology and Physiology Published by John Wiley & Sons Australia, Ltd.© 2017 The Authors. Clinical and Experimental Pharmacology and Physiology Published by John Wiley & Sons Australia, Ltd.)

Deficits in neuronal network synchrony in hippocampus and prefrontal cortex have been widely demonstrated in disorders of cognitive dysfunction, including schizophrenia and Alzheimer's disease. The atypical dopamine agonist SKF 83959 has been shown to increase brain-derived neurotrophic factor signalling and suppress activity of glycogen synthase kinase-3 in PFC, two processes important to learning and memory. The purpose of this study was to therefore evaluate the impact of SKF 83959 on oscillatory deficits in methylazoxymethanol acetate (MAM) rat model of schizophrenia. To achieve this, local field potentials were recorded simultaneously from the hippocampus and prefrontal cortex of anesthetized rats at 15 and 90 min following both acute and repeated administration of SKF 83959 (0.4 mg/kg). In MAM rats, but not controls, repeated SKF 83959 treatment increased signal amplitude in hippocampus and enhanced the spectral power of low frequency delta and theta oscillations in this region. In PFC, SKF 83959 increased delta, theta and gamma spectral power. Increased HIP-PFC theta coherence was also evident following acute and repeated SKF 83959. In apparent contradiction to these oscillatory effects, in MAM rats, SKF 83959 inhibited spatial learning and induced a significant increase in thigmotactic behaviour. These findings have uncovered a previously unknown role for SKF 83959 in the positive regulation of hippocampal-prefrontal cortical oscillatory network activity. As SKF 83959 is known to have affinity for a number of receptors, delineating the receptor mechanisms that mediate the positive drug effects on neuronal oscillations could have significant future implications in disorders associated with cognitive dysfunction., (© 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Recent papers have attributed arrhythmic substrate in murine RyR2-P2328S hearts to reduced action potential (AP) conduction velocities (CV), reflecting acute functional inhibition and/or reduced expression of sodium channels. We explored for acute effects of direct exchange protein directly activated by cAMP (Epac)-mediated ryanodine receptor-2 (RyR2) activation on arrhythmic substrate and CV. Monophasic action potential (MAP) recordings demonstrated that initial steady (8 Hz) extrinsic pacing elicited ventricular tachycardia (VT) in 0 of 18 Langendorff-perfused wild-type mouse ventricles before pharmacological intervention. The Epac activator 8-CPT (8-(4-chlorophenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate) (VT in 1 of 7 hearts), and the RyR2 blocker dantrolene, either alone (0 of 11) or with 8-CPT (0 of 9) did not then increase VT incidence (P>.05). Both progressively increased pacing rates and programmed extrasystolic (S2) stimuli similarly produced no VT in untreated hearts (n=20 and n=9 respectively). 8-CPT challenge then increased VT incidences (5 of 7 and 4 of 8 hearts respectively; P<.05). However, dantrolene, whether alone (0 of 10 and 1 of 13) or combined with 8-CPT (0 of 10 and 0 of 13) did not increase VT incidence relative to those observed in untreated hearts (P>.05). 8-CPT but not dantrolene, whether alone or combined with 8-CPT, correspondingly increased AP latencies (1.14±0.04 (n=7), 1.04±0.03 (n=10), 1.09±0.05 (n=8) relative to respective control values). In contrast, AP durations, conditions for 2:1 conduction block and ventricular effective refractory periods remained unchanged throughout. We thus demonstrate for the first time that acute RyR2 activation reversibly induces VT in specific association with reduced CV., (© 2017 The Authors. Clinical and Experimental Pharmacology and Physiology Published by John Wiley & Sons Australia, Ltd.)

Normal cardiac excitation involves orderly conduction of electrical activation and recovery dependent upon surface membrane, voltage-gated, sodium (Na(+) ) channel α-subunits (Nav 1.5). We summarize experimental studies of physiological and clinical consequences of loss-of-function Na(+) channel mutations. Of these conditions, Brugada syndrome (BrS) and progressive cardiac conduction defect (PCCD) are associated with sudden, often fatal, ventricular tachycardia (VT) or fibrillation. Mouse Scn5a(+/-) hearts replicate important clinical phenotypes modelling these human conditions. The arrhythmic phenotype is associated not only with the primary biophysical change but also with additional, anatomical abnormalities, in turn dependent upon age and sex, each themselves exerting arrhythmic effects. Available evidence suggests a unified binary scheme for the development of arrhythmia in both BrS and PCCD. Previous biophysical studies suggested that Nav 1.5 deficiency produces a background electrophysiological defect compromising conduction, thereby producing an arrhythmic substrate unmasked by flecainide or ajmaline challenge. More recent reports further suggest a progressive decline in conduction velocity and increase in its dispersion particularly in ageing male Nav 1.5 haploinsufficient compared to WT hearts. This appears to involve a selective appearance of slow conduction at the expense of rapidly conducting pathways with changes in their frequency distributions. These changes were related to increased cardiac fibrosis. It is thus the combination of the structural and biophysical changes both accentuating arrhythmic substrate that may produce arrhythmic tendency. This binary scheme explains the combined requirement for separate, biophysical and structural changes, particularly occurring in ageing Nav 1.5 haploinsufficient males in producing clinical arrhythmia., (© 2015 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.)

Drugs of abuse elicit dopamine release in the ventral striatum, possibly biasing dopamine-driven reinforcement learning towards drug-related reward at the expense of non-drug-related reward. Indeed, in alcohol-dependent patients, reactivity in dopaminergic target areas is shifted from non-drug-related stimuli towards drug-related stimuli. Such 'hijacked' dopamine signals may impair flexible learning from non-drug-related rewards, and thus promote craving for the drug of abuse. Here, we used functional magnetic resonance imaging to measure ventral striatal activation by reward prediction errors (RPEs) during a probabilistic reversal learning task in recently detoxified alcohol-dependent patients and healthy controls (N = 27). All participants also underwent 6-[(18) F]fluoro-DOPA positron emission tomography to assess ventral striatal dopamine synthesis capacity. Neither ventral striatal activation by RPEs nor striatal dopamine synthesis capacity differed between groups. However, ventral striatal coding of RPEs correlated inversely with craving in patients. Furthermore, we found a negative correlation between ventral striatal coding of RPEs and dopamine synthesis capacity in healthy controls, but not in alcohol-dependent patients. Moderator analyses showed that the magnitude of the association between dopamine synthesis capacity and RPE coding depended on the amount of chronic, habitual alcohol intake. Despite the relatively small sample size, a power analysis supports the reported results. Using a multimodal imaging approach, this study suggests that dopaminergic modulation of neural learning signals is disrupted in alcohol dependence in proportion to long-term alcohol intake of patients. Alcohol intake may perpetuate itself by interfering with dopaminergic modulation of neural learning signals in the ventral striatum, thus increasing craving for habitual drug intake., (© 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Aim: RyR2 mutations are associated with catecholaminergic polymorphic tachycardia, a condition characterized by ventricular and atrial arrhythmias. The present experiments investigate the atrial electrophysiology of homozygotic murine RyR2-P2328S (RyR2(S/S)) hearts for ectopic triggering events and for conduction abnormalities that might provide a re-entrant substrate., Methods: Electrocardiograph recordings were made from regularly stimulated RyR2(S/S) and wild type (WT) hearts, perfused using a novel modified Langendorff preparation. This permitted the simultaneous use of either floating intracellular microelectrodes to measure action potential (AP) parameters, or a multielectrode array to measure epicardial conduction velocity (CV)., Results: RyR2(S/S) showed frequent sustained tachyarrhythmias, delayed afterdepolarizations and ectopic APs, increased interatrial conduction delays, reduced epicardial CVs and reduced maximum rates of AP depolarization ((dV/dt)(max)), despite similar effective refractory periods, AP durations and AP amplitudes. Effective interatrial CVs and (dV/dt)(max) values of APs following ectopic (S2) stimulation were lower than those of APs following regular stimulation and decreased with shortening S1S2 intervals. However, although RyR2(S/S) atria showed arrhythmias over a wider range of S1S2 intervals, the interatrial CV and (dV/dt)(max) of S2 APs provoking such arrhythmias were similar in RyR2(S/S) and WT., Conclusions: These results suggest that abnormal intracellular Ca(2+) homoeostasis produces both arrhythmic triggers and a slow-conducting arrhythmic substrate in RyR2(S/S) atria. A similar mechanism might also contribute to arrhythmogenesis in other conditions, associated with diastolic Ca(2+) release, such as atrial fibrillation., (© 2012 The Authors Acta Physiologica © 2012 Scandinavian Physiological Society.)

Aim: QT interval prolongation reflecting delayed action potential (AP) repolarization is associated with polymorphic ventricular tachycardia and early after depolarizations potentially initiating extrasystolic APs if of sufficient amplitude. The current experiments explored contributions of altered re-excitation thresholds for, and conduction of, such extrasystolic APs to arrhythmogenesis in Langendorff-perfused, normokalaemic, control wild-type hearts and two experimental groups modelling long QT (LQT). The two LQT groups consisted of genetically modified, Scn5a(+/ΔKPQ) and hypokalaemic wild-type murine hearts., Methods: Hearts were paced from their right ventricles and monophasic AP electrode recordings obtained from their left ventricular epicardia, with recording and pacing electrodes separated by 1 cm. An adaptive programmed electrical stimulation protocol applied pacing (S1) stimulus trains followed by premature (S2) extrastimuli whose amplitudes were progressively increased with progressive decrements in S1S2 interval to maintain stimulus capture. Such protocols culminated in either arrhythmic or refractory endpoints., Results: Arrhythmic outcomes were associated with (1) lower conduction velocities in their initiating extrasystolic APs than refractory outcomes and (2) higher conduction velocities in the LQT groups than in controls. Furthermore, (3) the endpoints were reached at longer S1S2 coupling intervals and with smaller stimulus amplitudes in the LQT groups compared with controls. This was despite (4) similar relationships between conduction velocity and S1S2 coupling interval and between re-excitation thresholds and S1S2 coupling interval in all three experimental groups., Conclusions: Arrhythmias induced by extrasystolic APs in the LQT groups thus occur under conditions of higher conduction velocity and greater sensitivity to extrastimuli than in controls., (© 2012 The Authors Acta Physiologica © 2012 Scandinavian Physiological Society.)

1. In the present study, we investigated the effect of age on atrial electrophysiological properties in Scn5a(+/-) hearts used to model corresponding increases in atrial arrhythmic tendency in human Brugada syndrome. 2. Atrial action potential initiation, propagation and recovery were compared in young (3 month old) and aged (12 month old), wild-type (WT) and Scn5a(+/-) hearts. Multielectrode array recordings assessed the spatial propagation of intrinsic electrical activity in superfused atrial preparations, whereas bipolar electrogram recordings measured basic cycle lengths (BCL) in Langendorff preparations. The duration of electrogram activity (EGD) during regular and extrasystolic stimulation with programmed electrical stimulation provided EGD ratios and atrial effective refractory periods (AERP). Monophasic recordings measured action potential durations (APD). 3. Systematic statistical explorations for independent and interacting effects of age and the Scn5a(+/-) condition demonstrated that both young and aged Scn5a(+/-) mice exhibited slowed propagation of atrial excitation relative to corresponding WT mice, with the greatest effects in aged Scn5a(+/-) mice, which additionally exhibited increased intrinsic BCL. 4. Young Scn5a(+/-) mice exhibited greater EGD and EGD ratios, as well as APD/AERP ratios, suggesting increased arrhythmic tendency compared with WT mice. 5. Aged Scn5a(+/-) mice exhibited normal EGD, EGD ratios and APD compared to aged WT and young Scn5a(+/-), and increased AERP and smaller APD/AERP ratios compared with young Scn5a(+/-). 6. These electrophysiological findings indicate increased atrial arrhythmogenicity with maximal effects on both conduction and repolarization characteristics in young compared with aged Scn5a(+/-) mice., (© 2012 The Authors Clinical and Experimental Pharmacology and Physiology © 2012 Blackwell Publishing Asia Pty Ltd.)

Stressors can exert a wide variety of responses, ranging from adaptive responses to pathological changes; moreover, recent studies suggest that mild stressors can attenuate the response of a system to major stressful events. We have previously shown that 2-week exposure to cold, a comparatively mild inescapable stressor, induced a pronounced reduction in ventral tegmental area (VTA) dopamine (DA) neuron activity, whereas restraint stress increases DA neuron activity. However, it is not known if these stressors differentially impact the VTA in a region-specific manner, if they differentially impact behavioral responses, or whether the effects of such different stressors are additive or antagonistic with regard to their impact on DA neuron firing. To address these questions, single-unit extracellular recordings were performed in anesthetized control rats and rats exposed to chronic cold, and tested after delivery of a 2-h restraint session. Chronic cold stress strongly attenuated the number of DA neurons firing in the VTA, and this effect occurred primarily in the medial and central VTA regions that preferentially project to reward-related ventral striatal regions. Chronic cold exposure also prevented the pronounced increase in DA neuron population activity without affecting the behavioral sensitization to amphetamine produced by restraint stress. Taken together, these data show that a prolonged inescapable mild stressor can induce plastic changes that attenuate the DA system response to acute stress., (© 2012 The Authors. European Journal of Neuroscience © 2012 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.)

Loss- and gain-of-function cardiac Na(+) channel (SCN5A) mutations are associated with the Brugada and long QT type 3 syndromes (LQT3), respectively. Both result in ventricular arrhythmias, but have differing atrial effects. We have recently reported decreased incidences of atrial arrhythmias in a murine Scn5a+/Delta cardiac system used to model LQT3. We now explore for atrial arrhythmias in a murine Brugada syndrome model. Programmed electrical stimulation and burst pacing were applied to 10 wild-type (WT) and 10 Scn5a+/- Langendorff-perfused murine hearts. These induced arrhythmias in similar proportions of the Scn5a+/- and WT hearts, in contrast to their decreased incidences previously reported for Scn5a+/Delta. Almost half of the Scn5a+/- hearts displayed spontaneous atrial arrhythmias never observed in WT. Both these WT and Scn5a+/- hearts showed positive critical intervals, given by the difference between action potential durations at 90% recovery (APD(90)) and atrial effective refractory periods, before pharmacological intervention. Both flecainide and quinidine then prevented both induced and spontaneous atrial arrhythmias in all the arrhythmic WT and Scn5a+/- hearts, in contrast to their differing ventricular effects. However, flecainide prolonged APD(90) in WT but not Scn5a+/-, whereas quinidine prolonged APD(90) in both WT and Scn5a+/-. Nevertheless, addition of both agents markedly increased atrial effective refractory periods to values exceeding the corresponding values of APD(90). This resulted in negative critical intervals in both WT and Scn5a+/-. These findings demonstrate a clear-cut relationship between critical interval and atrial arrhythmic tendency for the first time and extend previous reports that had related critical interval to ventricular arrhythmia.

Aim: clinical observations suggest that alternans in action potential (AP) characteristics presages breakdown of normal ordered cardiac electrical activity culminating in ventricular arrhythmogenesis. We compared such temporal nonuniformities in monophasic action potential (MAP) waveforms in left (LV) and right ventricular (RV) epicardia and endocardia of Langendorff-perfused murine wild-type (WT), and Scn5a(+/-) hearts modelling Brugada syndrome (BrS) for the first time., Methods: a dynamic pacing protocol imposed successively incremented steady pacing rates between 5.5 and 33 Hz. A signal analysis algorithm detected sequences of >10 beats showing alternans. Results were compared before and following the introduction of flecainide (10 microm) and quinidine (5 microm) known to exert pro- and anti-arrhythmic effects in BrS., Results: sustained and transient amplitude and duration alternans were both frequently followed by ventricular ectopic beats and ventricular tachycardia or fibrillation. Diastolic intervals (DIs) that coincided with onsets of transient (tr) or sustained (ss) alternans in MAP duration (DI*) and amplitude (DI') were determined. Kruskal-Wallis tests followed by Bonferroni-corrected Mann-Whitney U-tests were applied to these DI results sorted by recording site, pharmacological conditions or experimental populations. WT hearts showed no significant heterogeneities in any DI. Untreated Scn5a (+/-) hearts showed earlier onsets of transient but not sustained duration alternans in LV endocardium compared with RV endocardium or LV epicardium. Flecainide administration caused earlier onsets of both transient and sustained duration alternans selectively in the RV epicardium in the Scn5a (+/-) hearts., Conclusion: these findings in a genetic model thus implicate RV epicardial changes in the arrhythmogenicity produced by flecainide challenge in previously asymptomatic clinical BrS.

Aim: To investigate the interacting effects of age and sex on electrocardiographic (ECG) features of Scn5a(+/-) mice modelling Brugada syndrome., Methods: Recordings were performed on anaesthetized wild-type (WT) and Scn5a(+/-) mice and differences attributable to these risk factors statistically stratified., Results: Scn5a(+/-) exerted sex-dependent effects upon sino-atrial function that only became apparent with age. RR intervals were greater in old male than in old female Scn5a(+/-). Atrio-ventricular (AV) conduction was slower in young female mice, whether WT and Scn5a(+/-), than the corresponding young male WT and Scn5a(+/-). However, PR intervals lengthened with age in male but not in female Scn5a(+/-) giving the greatest PR intervals in old male Scn5a(+/-) compared with either old male WT or young male Scn5a(+/-) mice. In contrast, PR intervals were similar in old female Scn5a(+/-) and in old female WT. QTc was prolonged in Scn5a(+/-) compared with WT, and female Scn5a(+/-) compared with female WT. Age-dependent alterations in durations of ventricular repolarization relative to WT affected male but not female Scn5a(+/-). Thus, T-wave durations were greater in old male Scn5a(+/-) compared with old male WT, but indistinguishable between old female Scn5a(+/-) and old female WT. Finally, analysis for combined interactions of genotype, age and sex demonstrated no effects on P wave and QRS durations and QTc intervals., Conclusion: We demonstrate for the first time that age, sex and genotype exert both independent and interacting ECG effects. The latter suggest alterations in cardiac pacemaker function, atrio-ventricular conduction and ventricular repolarization greatest in ageing male Scn5a(+/-).

1. Flecainide and quinidine exert contrasting pro-arrhythmic and anti-arrhythmic effects in mouse hearts containing the loss-of-function, Scn5a(+/-), and the gain-of-function, Scn5a(+/DeltaKPQ), mutations in their sodium channel alpha-subunits. 2. The following properties were accordingly compared in wild-type and Scn3b(-/-) hearts modelling modifications in the beta-subunit, before and after introduction of either agent: (i) ventricular arrhythmogenecity and effective refractory periods (VERP) in response to programmed electrical stimulation (PES); (ii) monophasic action potential waveforms recorded from the left ventricular epicardium and endocardium; (iii) action potential durations (APD) obtained from the monophasic action potentials; and (iv) critical intervals derived from the APD and VERP values. 3. Ventricular tachycardia was induced by PES in 11 out of 15 Scn3b(-/-) hearts and 0 out of 17 wild-type hearts. This incidence was reduced to three out of eight Scn3b(-/-) hearts but increased to three out of eight wild-type hearts with flecainide. 4. Arrhythmogenic incidence was reduced to two out of eight Scn3b(-/-) hearts and remained at 0 out of eight wild-type hearts in the presence of quinidine. 5. Ventricular effective refractory periods were prolonged and endocardial and epicardial APD shortened, resulting in negative critical intervals in both Scn3b(-/-) and wild-type hearts treated by either flecainide or quinidine. Nevertheless, endocardial APD remained consistently longer than epicardial APD, leaving similar, positive endocardial-epicardial, differences, DeltaAPD, in treated and untreated Scn3b(-/-) and wild-type hearts. 6. It is concluded that both flecainide and quinidine exert anti-arrhythmogenic effects in Scn3b(-/-) hearts, doing so through modifying VERP rather than DeltaAPD, in contrast to their differing effects in Scn5a(+/-) and Scn5a(+/DeltaKPQ) hearts.

Aim: To investigate the effect of increases in extracellular Ca(2+) entry produced by the L-type Ca(2+) channel agonist FPL-64176 (FPL) upon acute atrial arrhythmogenesis in intact Langendorff-perfused mouse hearts and its dependence upon diastolic Ca(2+) release from sarcoplasmic reticular Ca(2+) stores., Methods: Confocal microscope studies of Fluo-3 fluorescence in isolated atrial myocytes were performed in parallel with electrophysiological examination of Langendorff-perfused mouse hearts., Results: Atrial myocytes stimulated at 1 Hz and exposed to FPL (0.1 microm) initially showed (<10 min) frequent, often multiple, diastolic peaks following the evoked Ca(2+) transients whose amplitudes remained close to control values. With continued pacing (>10 min) this reverted to a regular pattern of evoked transients with increased amplitudes but in which diastolic peaks were absent. Higher FPL concentrations (1.0 microm) produced sustained and irregular patterns of cytosolic Ca(2+) activity, independent of pacing. Nifedipine (0.5 microm), and caffeine (1.0 mm) and cyclopiazonic acid (CPA) (0.15 microm) pre-treatments respectively produced immediate and gradual reductions in the F/F(0) peaks. Such nifedipine and caffeine, or CPA pre-treatments, abolished, or reduced, the effects of 0.1 and 1.0 microm FPL on cytosolic Ca(2+) signals. FPL (1.0 microm) increased the incidence of atrial tachycardia and fibrillation in intact Langendorff-perfused hearts without altering atrial effective refractory periods. These effects were inhibited by nifedipine and caffeine, and reduced by CPA., Conclusion: Enhanced extracellular Ca(2+) entry exerts acute atrial arrhythmogenic effects that is nevertheless dependent upon diastolic Ca(2+) release. These findings complement reports that associate established, chronic, atrial arrhythmogenesis with decreased overall inward Ca(2+) current.

Aim: In contrast to extensive reports on the roles of Na(v)1.5 alpha-subunits, there have been few studies associating the beta-subunits with cardiac arrhythmogenesis. We investigated the sino-atrial and conduction properties in the hearts of Scn3b(-/-) mice., Methods: The following properties were compared in the hearts of wild-type (WT) and Scn3b(-/-) mice: (1) mRNA expression levels of Scn3b, Scn1b and Scn5a in atrial tissue. (2) Expression of the beta(3) protein in isolated cardiac myocytes. (3) Electrocardiographic recordings in intact anaesthetized preparations. (4) Bipolar electrogram recordings from the atria of spontaneously beating and electrically stimulated Langendorff-perfused hearts., Results: Scn3b mRNA was expressed in the atria of WT but not Scn3b(-/-) hearts. This was in contrast to similar expression levels of Scn1b and Scn5a mRNA. Immunofluorescence experiments confirmed that the beta(3) protein was expressed in WT and absent in Scn3b(-/-) cardiac myocytes. Lead I electrocardiograms from Scn3b(-/-) mice showed slower heart rates, longer P wave durations and prolonged PR intervals than WT hearts. Spontaneously beating Langendorff-perfused Scn3b(-/-) hearts demonstrated both abnormal atrial electrophysiological properties and evidence of partial or complete dissociation of atrial and ventricular activity. Atrial burst pacing protocols induced atrial tachycardia and fibrillation in all Scn3b(-/-) but hardly any WT hearts. Scn3b(-/-) hearts also demonstrated significantly longer sinus node recovery times than WT hearts., Conclusion: These findings demonstrate, for the first time, that a deficiency in Scn3b results in significant atrial electrophysiological and intracardiac conduction abnormalities, complementing the changes in ventricular electrophysiology reported on an earlier occasion.

1. Intracellular Ca(2+) overload has been associated with established atrial arrhythmogenesis. The present experiments went on to correlate acute initiation of atrial arrhythmogenesis in Langendorff-perfused mouse hearts with changes in Ca(2+) homeostasis in isolated atrial myocytes following pharmacological procedures that modified the storage or release of sarcoplasmic reticular (SR) Ca(2+) or inhibited entry of extracellular Ca(2+). 2. Caffeine (1 mmol/L) elicited diastolic Ca(2+) waves in regularly stimulated atrial myocytes immediately following addition. This was followed by a decline in the amplitude of the evoked transients and the disappearance of such diastolic events, suggesting partial SR Ca(2+) depletion. 3. Cyclopiazonic acid (CPA; 0.15 micromol/L) produced more gradual reductions in evoked Ca(2+) transients and abolished diastolic Ca(2+) events produced by the further addition of caffeine. 4. Nifedipine (0.5 micromol/L) produced immediate reductions in evoked Ca(2+) transients. Further addition of caffeine produced an immediate increase followed by a decline in the amplitude of the evoked Ca(2+) transients, without eliciting diastolic Ca(2+) events. 5. These findings correlated with changes in spontaneous and provoked atrial arrhythmogenecity in mouse isolated Langendorf-perfused hearts. Thus, caffeine was pro-arrhythmogenic immediately following but not > 5 min after application and both CPA and nifedipine pretreatment inhibited such arrhythmogenesis. 6. Together, these findings relate acute atrial arrhythmogenesis in intact hearts to diastolic Ca(2+) events in atrial myocytes that, in turn, depend upon a finite SR Ca(2+) store and diastolic Ca(2+) release following Ca(2+)-induced Ca(2+) release initiated by the entry of extracellular Ca(2+).

The experiments explored the extent to which alterations in L-type Ca(2+) channel-mediated Ca(2+) entry triggers Ca(2+)-mediated arrhythmogenesis in Langendorff-perfused murine hearts through use of the specific L-type Ca(2+) channel modulator FPL-64716 (FPL). Introduction of FPL (1 microm) resulted in a gradual development (>10 min) of diastolic electrical events and alternans in spontaneously beating hearts from which monophasic action potentials were recorded. In regularly paced hearts, they additionally led to non-sustained and sustained ventricular tachycardia (nsVT and sVT). Programmed electrical stimulation (PES) resulted in nsVT and sVT after 5-10 and >10 min perfusion, respectively. Pretreatments with nifedipine, diltiazem and cyclopiazonic acid abolished arrhythmogenic tendency induced by subsequent introduction of FPL, consistent with its dependence upon both extracellular Ca(2+) entry and the degree of filling of the sarcoplasmic reticular Ca(2+) store. Values for action potential duration at 90% repolarization when any of these agents were applied to FPL-treated hearts became indistinguishable from those shown by untreated control hearts, in contrast to earlier reports of their altering in long QT syndrome type 3 and hypokalaemic murine models for re-entrant arrhythmogenesis. These arrhythmic effects instead correlated with alterations in Ca(2+) homeostasis at the single-cell level found in investigations of the effects of both FPL and the same agents in regularly stimulated fluo-3 loaded myocytes. These findings are compatible with a prolonged extracellular Ca(2+) entry that potentially results in an intracellular Ca(2+) overload and produces the cardiac arrhythmogenecity following addition of FPL.

With no further intervention, relapse rates in detoxified alcoholics are high and usually exceed 80% of all detoxified patients. It has been suggested that stress and exposure to priming doses of alcohol and to alcohol-associated stimuli (cues) contribute to the relapse risk after detoxification. This article focuses on neuronal correlates of cue responses in detoxified alcoholics. Current brain imaging studies indicate that dysfunction of dopaminergic, glutamatergic and opioidergic neurotransmission in the brain reward system (ventral striatum including the nucleus accumbens) can be associated with alcohol craving and functional brain activation in neuronal systems that process attentional relevant stimuli, reward expectancy and experience. Increased functional brain activation elicited by such alcohol-associated cues predicted an increased relapse risk, whereas high brain activity elicited by affectively positive stimuli may represent a protective factor and was correlated with a decreased prospective relapse risk. These findings are discussed with respect to psychotherapeutic and pharmacological treatment options.

Aim: We compared the clinical and physiological consequences of the novel mutation R878C in a highly conserved pore residue in domain II (S5-S6) of human, hNa(v)1.5, cardiac Na(+) channels., Methods: Full clinical evaluation of pedigree members through three generations of a Chinese family combined with SCN5A sequencing from genomic DNA was compared with patch and voltage-clamp results from two independent expression systems., Results: The four mutation carriers showed bradycardia, and slowed sino-atrial, atrioventricular and intraventricular conduction. Two also showed sick sinus syndrome; two had ST elevation in leads V1 and V2. Unlike WT-hNa(v)1.5, whole-cell patch-clamped HEK293 cells expressing R878C-hNa(v)1.5 showed no detectable Na(+) currents (i(Na)), even with substitution of a similarly charged lysine residue. Voltage-clamped Xenopus oocytes injected with either 0.04 or 1.5 microg microL(-1) R878C-hNa(v)1.5 cRNA similarly showed no i(Na), yet WT-hNa(v)1.5 cRNA diluted to 0.0004-0.0008 ng microL(-1)resulted in expression of detectable i(Na). i(Na) was simply determined by the amount of injected WT-hNa(v)1.5: doubling the dose of WT-hNa(v)1.5 cRNA doubled i(Na). i(Na) amplitudes and activation and inactivation characteristics were similar irrespective of whether WT-hNa(v)1.5 cRNA was given alone or combined with equal doses of R878C-hNa(v)1.5 cRNA therefore excluding dominant negative phenotypic effects. Na(+) channel function in HEK293 cells transfected with R878C-hNa(v)1.5 was not restored by exposure to mexiletine (200 microM) and lidocaine (100 microM). Fluorescence confocal microscopy using E3-Nav1.5 antibody demonstrated persistent membrane expression of both WT and R878C-hNa(v)1.5. Modelling studies confirmed that such i(Na) reductions reproduced the SSS phenotype., Conclusion: Clinical consequences of the novel R878C mutation correlate with results of physiological studies.

Aim: To explore the physiological consequences of the ryanodine receptor (RyR2)-P2328S mutation associated with catecholaminergic polymorphic ventricular tachycardia (CPVT)., Methods: We generated heterozygotic (RyR2 p/s) and homozygotic (RyR2 s/s) transgenic mice and studied Ca2+ signals from regularly stimulated, Fluo-3-loaded, cardiac myocytes. Results were compared with monophasic action potentials (MAPs) in Langendorff-perfused hearts under both regular and programmed electrical stimulation (PES)., Results: Evoked Ca2+ transients from wild-type (WT), heterozygote (RyR2 p/s) and homozygote (RyR2 s/s) myocytes had indistinguishable peak amplitudes with RyR2 s/s showing subsidiary events. Adding 100 nm isoproterenol produced both ectopic peaks and subsidiary events in WT but not RyR2 p/s and ectopic peaks and reduced amplitudes of evoked peaks in RyR2 s/s. Regularly stimulated WT, RyR2 p/s and RyR2 s/s hearts showed indistinguishable MAP durations and refractory periods. RyR2 p/s hearts showed non-sustained ventricular tachycardias (nsVTs) only with PES. Both nsVTs and sustained VTs (sVTs) occurred with regular stimuli and PES with isoproterenol treatment. RyR2 s/s hearts showed higher incidences of nsVTs before but mainly sVTs after introduction of isoproterenol with both regular stimuli and PES, particularly at higher pacing frequencies. Additionally, intrinsically beating RyR2 s/s showed extrasystolic events often followed by spontaneous sVT., Conclusion: The RyR2-P2328S mutation results in marked alterations in cellular Ca2+ homeostasis and arrhythmogenic properties resembling CPVT with greater effects in the homozygote than the heterozygote demonstrating an important gene dosage effect.

Aim: We explored the anti-arrhythmic efficacy of K(+) channel activation in the hypokalaemic murine heart using NS1643 and nicorandil, compounds which augment I(Kr) and I(KATP) respectively., Methods: Left ventricular epicardial and endocardial monophasic action potentials were compared in normokalaemic and hypokalaemic preparations in the absence and presence of NS1643 (30 microM) and nicorandil (20 microM)., Results: Spontaneously beating hypokalaemic hearts (3 mM K(+)) all elicited early afterdepolarizations (EADs) and episodes of ventricular tachycardia (VT). Perfusion with NS1643 and nicorandil suppressed EADs and VT in 7 of 13 and five of six hypokalaemic hearts. Provoked arrhythmia studies using programmed electrical stimulation induced VT in all hypokalaemic hearts, but failed to do so in 7 of 13 and five of six hearts perfused with NS1643 and nicorandil respectively. These anti-arrhythmic effects were accompanied by reductions in action potential duration at 90% repolarization (APD(90)) and changes in the transmural gradient of repolarization, reflected in DeltaAPD(90). NS1643 and nicorandil reduced epicardial APD(90) from 68.3 +/- 1.1 to 56.5 +/- 4.1 and 51.5 +/- 1.5 ms, respectively, but preserved endocardial APD(90) in hypokalaemic hearts. NS1643 and nicorandil thus restored DeltaAPD(90) from -9.6 +/- 4.3 ms under baseline hypokalaemic conditions to 3.9 +/- 4.1 and 9.9 +/- 2.1 ms, respectively, close to normokalaemic values., Conclusion: These findings demonstrate, for the first time, the anti-arrhythmic efficacy of K(+) channel activation in the setting of hypokalaemia. NS1643 and nicorandil are anti-arrhythmic through the suppression of EADs, reductions in APD(90) and restorations of DeltaAPD(90).

Chronic stress exposure alters the central noradrenergic neurons originating from the locus coeruleus (LC). Previously, we demonstrated that evoked increases in the firing rate of LC neurons and their release of norepinephrine are enhanced following chronic cold exposure. In the present studies, we tested the hypothesis that reduced feedback inhibition of LC neurons might underlie these alterations in LC activity by examining the effect of alpha(2)-autoreceptor stimulation on LC activity in chronically stressed rats using in vivo and in vitro single unit recordings. Given that regulators of G-protein signaling (RGS) proteins can impact the coupling of alpha(2)-autoreceptors to downstream signaling cascades, we also explored the expression of several RGS proteins following chronic stress exposure. We observed that the alpha(2)-autoreceptor-evoked inhibition of LC neurons was reduced and that the expression of RGS7 was increased following chronic stress exposure. Finally, we demonstrated that intracellular administration of RGS7 via patch clamp electrodes mimicked the stress-induced decrease in clonidine-evoked autoreceptor-mediated inhibition. These novel data provide a mechanism to explain how chronic stress-induced alterations in receptor coupling can result in changes in alpha(2)-autoreceptor control of noradrenergic function throughout the central nervous system, potentially leading to alterations in anxiety-related behaviors, and may suggest novel therapeutic targets for the treatment of mood and anxiety disorders.

Aim: To perform an empirical, pharmacological, separation of early afterdepolarizations (EADs) and transmural gradients of repolarization in arrhythmogenesis in a genetically modified mouse heart modelling human long QT syndrome (LQT) 3., Methods: Left ventricular endocardial and epicardial monophasic action potentials and arrhythmogenic tendency were compared in isolated wild type (WT) and Scn5a+/Delta hearts perfused with 0.1 and 1 microm propranolol and paced from the right ventricular epicardium., Results: All spontaneously beating bradycardic Scn5a+/Delta hearts displayed EADs, triggered beats and ventricular tachycardia (VT; n = 7), events never seen in WT hearts (n = 5). Perfusion with 0.1 and 1 microm propranolol suppressed all EADs, triggered beats and episodes of VT. In contrast, triggering of VT persisted following programmed electrical stimulation in 6 of 12 (50%), one of eight (12.5%), but six of eight (75%) Scn5a+/Delta hearts perfused with 0, 0.1 and 1 microm propranolol respectively in parallel with corresponding alterations in repolarization gradients, reflected in action potential duration (DeltaAPD(90)) values. Thus 0.1 microm propranolol reduced epicardial but not endocardial APD(90) from 54.7 +/- 1.6 to 44.0 +/- 2.0 ms, restoring DeltaAPD(90) from -3.8 +/- 1.6 to 3.5 +/- 2.5 ms (all n = 5), close to WT values. However, 1 microm propranolol increased epicardial APD(90) to 72.5 +/- 1.2 ms and decreased endocardial APD(90) from 50.9 +/- 1.0 to 24.5 +/- 0.3 ms, increasing DeltaAPD(90) to -48.0 +/- 1.2 ms., Conclusion: These findings empirically implicate EADs in potentially initiating spontaneous arrhythmogenic phenomena and transmural repolarization gradients in the re-entrant substrate that would sustain such activity when provoked by extrasystolic activity in murine hearts modelling human LQT3 syndrome.

Sudden cardiac death stemming from ventricular arrhythmogenesis is one of the major causes of mortality in the developed world. Congenital and acquired forms of long QT syndrome (LQTS) are in turn associated with life threatening arrhythmias. Over the past decade our understanding of arrhythmogenic mechanisms in the setting of these diseases has increased greatly due to the creation of a number of animal models. Of these, the genetically amenable mouse has proved to be a particularly powerful tool. This review summarizes the congenital and acquired LQTS and describes the various mouse models that have been created to further probe arrhythmogenic mechanisms.

Aims: We resolved roles for early afterdepolarizations (EADs) and transmural gradients of repolarization in arrhythmogenesis in Langendorff-perfused hypokalaemic murine hearts paced from the right ventricular epicardium., Methods: Left ventricular epicardial and endocardial monophasic action potentials (MAPs) and arrhythmogenic tendency were compared in the presence and absence of the L-type Ca(2+) channel blocker nifedipine (10 nm-1 microm) and the calmodulin kinase type II inhibitor KN-93 (2 microm)., Results: All the hypokalaemic hearts studied showed prolonged epicardial and endocardial MAPs, decreased epicardial-endocardial APD(90) difference, EADs, triggered beats and ventricular tachycardia (VT) (n = 6). In all spontaneously beating hearts, 100 (but not 10) nm nifedipine reduced both the incidence of EADs and triggered beats from 66.9 +/- 15.7% to 28.3 +/- 8.7% and episodes of VT from 10.8 +/- 6.3% to 1.2 +/- 0.7% of MAPs (n = 6 hearts, P < 0.05); 1 microm nifedipine abolished all these phenomena (n = 6). In contrast programmed electrical stimulation (PES) still triggered VT in six of six hearts with 0, 10 and 100 nm but not 1 microm nifedipine. 1 microm nifedipine selectively reduced epicardial (from 66.1 +/- 3.4 to 46.2 +/- 2.5 ms) but not endocardial APD(90), thereby restoring DeltaAPD(90) from -5.9 +/- 2.5 to 15.5 +/- 3.2 ms, close to normokalaemic values. KN-93 similarly reduced EADs, triggered beats and VT in spontaneously beating hearts to 29.6 +/- 8.9% and 1.7 +/- 1.1% respectively (n = 6) yet permitted PES-induced VT (n = 6), in the presence of a persistently negative DeltaAPD(90)., Conclusions: These findings empirically implicate both EADs and triggered beats alongside arrhythmogenic substrate of DeltaAPD(90) in VT pathogenesis at the whole heart level.

Aim: Hypokalaemia is associated with a lethal form of ventricular tachycardia (VT), torsade de pointes, through pathophysiological mechanisms requiring clarification., Methods: Left ventricular endocardial and epicardial monophasic action potentials were compared in isolated mouse hearts paced from the right ventricular epicardium perfused with hypokalaemic (3 and 4 mm [K(+)](o)) solutions. Corresponding K(+) currents were compared in whole-cell patch-clamped epicardial and endocardial myocytes., Results: Hypokalaemia prolonged epicardial action potential durations (APD) from mean APD(90)s of 37.2 +/- 1.7 ms (n = 7) to 58.4 +/- 4.1 ms (n =7) and 66.7 +/- 2.1 ms (n = 11) at 5.2, 4 and 3 mm [K(+)](o) respectively. Endocardial APD(90)s correspondingly increased from 51.6 +/- 1.9 ms (n = 7) to 62.8 +/- 2.8 ms (n = 7) and 62.9 +/- 5.9 ms (n = 11) giving reductions in endocardial-epicardial differences, DeltaAPD(90), from 14.4 +/- 2.6 to 4.4 +/- 5.0 and -3.4 +/- 6.0 ms respectively. Early afterdepolarizations (EADs) occurred in epicardia in three of seven spontaneously beating hearts at 4 mm [K(+)](o) with triggered beats followed by episodes of non-sustained VT in nine of 11 preparations at 3 mm. Programmed electrical stimulation never induced arrhythmic events in preparations perfused with normokalemic solutions yet induced VT in two of seven and nine of 11 preparations at 4 and 3 mm [K(+)](o) respectively. Early outward K(+) current correspondingly fell from 73.46 +/- 8.45 to 61.16+/-6.14 pA/pF in isolated epicardial but not endocardial myocytes (n = 9) (3 mm [K(+)](o))., Conclusions: Hypokalaemic mouse hearts recapitulate the clinical arrhythmogenic phenotype, demonstrating EADs and triggered beats that might initiate VT on the one hand and reduced transmural dispersion of repolarization reflected in DeltaAPD(90) suggesting arrhythmogenic substrate on the other.

The membrane protein KCNE1 has been implicated in cell volume regulation. Using a knockout mouse model, this study examined the role of KCNE1 in regulatory volume decrease (RVD) in freshly isolated renal proximal tubule cells. Cell diameter was measured using an optical technique in response to hypotonic shock and stimulation of Na(+)-alanine cotransport in cells isolated from wild-type and KCNE1 knockout mice. In HEPES buffered solutions 64% of wild-type and 56% of knockout cells demonstrated RVD. In HCO3- buffered solutions 100% of the wild-type cells showed RVD, while in the knockout cells the proportion of cells displaying RVD remained unchanged. RVD in the knockout cells was rescued by valinomycin, a K+ ionophore. In wild-type HCO3- dependent cells the K+ channel inhibitors barium and clofilium inhibited RVD. These data suggest that mouse renal proximal tubule is comprised of two cell populations. One cell population is capable of RVD in the absence of HCO3-, whereas RVD in the other cell population has an absolute requirement for HCO3-. The HCO3- dependent RVD requires the normal expression of KCNE1.

All drugs of abuse have been shown to act either directly or indirectly by increasing dopamine neurotransmission within the limbic system. Thus, alcohol has been shown to increase dopamine transmission primarily by activating dopamine cell spike activity, whereas psychostimulants increase dopamine transmission by inhibiting the removal of dopamine from the synaptic space after its release. The spike-dependent release of dopamine that is modulated by drugs of abuse to lead to their rewarding actions has been termed the phasic dopamine response. In contrast, with repeated drug administration, dopamine will also accumulate in the extracellular space of the nucleus accumbens in concentrations too low to stimulate postsynaptic receptors, but of sufficient magnitude to activate dopamine release-inhibiting autoreceptors. In addition, the level of extracellular dopamine is proposed to be under the regulatory influence of cortico-accumbens afferents. This steady-state level of extrasynaptic dopamine has been termed the tonic dopamine response. In this paper it is proposed that several of the aspects of drug addiction, withdrawal and craving associated with the continued use of these drugs can be explained on the basis of their effects on tonic versus phasic dopamine system function. Thus, the increase in tonic dopamine levels that occurs with repeated drug administration would serve to oppose phasic dopamine release via stimulation of dopamine terminal autoreceptors, causing the subject to increase drug administration to restore the phasic response. Moreover, after withdrawal from the drugs, exposure to priming doses of drug or to drug-related stimuli are proposed to increase tonic dopamine levels, again triggering drug-seeking behavior in order to restore balance between the tonic and phasic dopamine systems. Therefore, one consequence of continued drug use is that these parameters of dopamine system function that normally serve to keep the system stable will enter into a new steady-state homeostasis, from which the system is particularly susceptible to destabilizing influences that may precipitate relapse.