The cardiac electrical impulse depends on an orchestrated interplay of transmembrane ionic currents in myocardial cells. Two critical ionic current mechanisms are the inwardly rectifying potassium current (I K1 ), which is important for maintenance of the cell resting membrane potential, and the sodium current (I Na ), which provides a rapid depolarizing current during the upstroke of the action potential. By controlling the resting membrane potential, I K1 modifies sodium channel availability and therefore, cell excitability, action potential duration, and velocity of impulse propagation. Additionally, I K1 -I Na interactions are key determinants of electrical rotor frequency responsible for abnormal, often lethal, cardiac reentrant activity. Here, we have used a multidisciplinary approach based on molecular and biochemical techniques, acute gene transfer or silencing, and electrophysiology to show that I K1 -I Na interactions involve a reciprocal modulation of expression of their respective channel proteins (Kir2.1 and Na V 1.5) within a macromolecular complex. Thus, an increase in functional expression of one channel reciprocally modulates the other to enhance cardiac excitability. The modulation is model-independent; it is demonstrable in myocytes isolated from mouse and rat hearts and with transgenic and adenoviral-mediated overexpression/silencing. We also show that the post synaptic density, discs large, and zonula occludens-1 (PDZ) domain protein SAP97 is a component of this macromolecular complex. We show that the interplay between Na v 1.5 and Kir2.1 has electrophysiological consequences on the myocardium and that SAP97 may affect the integrity of this complex or the nature of Na v 1.5-Kir2.1 interactions. The reciprocal modulation between Na v 1.5 and Kir2.1 and the respective ionic currents should be important in the ability of the heart to undergo self-sustaining cardiac rhythm disturbances.reentry | scaffolding proteins | conduction velocity | protein trafficking I n the heart, the inward rectifying potassium current (I K1 ) is the major current responsible for the maintenance of the resting membrane potential (RMP), whereas the sodium current (I Na ) provides the largest fraction of the inward depolarizing current that flows during an action potential (1). It is well-known that a relationship exists between these two ionic currents that is crucial for proper cardiac electrical function; disruption of this balance results in changes in sodium channel availability, cell excitability, action potential duration, and conduction velocity (2). Accordingly, I K1 -I Na interactions are important in stabilizing and controlling the frequency of the electrical rotors that are responsible for the most dangerous cardiac arrhythmias, including ventricular tachycardia and fibrillation (3).Post synaptic density, discs large, and zonula occludens-1 (PDZ) domain proteins link different and in many cases, multiple proteins to macromolecular complexes through interactions with their various domains. More than 70 PDZ d...

Histone H3 lysine 4 (H3K4me) methyltransferases and their cofactors are essential for embryonic development and the establishment of gene expression patterns in a cell-specific and heritable manner. However, the importance of such epigenetic marks in maintaining gene expression in adults and in initiating human disease is unclear. Here, we addressed this question using a mouse model in which we could inducibly ablate PAX interacting (with transcription-activation domain) protein 1 (PTIP), a key component of the H3K4me complex, in cardiac cells. Reducing H3K4me3 marks in differentiated cardiomyocytes was sufficient to alter gene expression profiles. One gene regulated by H3K4me3 was Kv channel-interacting protein 2 (Kcnip2), which regulates a cardiac repolarization current that is downregulated in heart failure and functions in arrhythmogenesis. This regulation led to a decreased sodium current and action potential upstroke velocity and significantly prolonged action potential duration (APD). The prolonged APD augmented intracellular calcium and in vivo systolic heart function. Treatment with isoproterenol and caffeine in this mouse model resulted in the generation of premature ventricular beats, a harbinger of lethal ventricular arrhythmias. These results suggest that the maintenance of H3K4me3 marks is necessary for the stability of a transcriptional program in differentiated cells and point to an essential function for H3K4me3 epigenetic marks in cellular homeostasis.

Aims Collecting electrophysiological and molecular data from the murine conduction system presents technical challenges. We have developed an approach for the isolation of murine Purkinje cells (PCs), characterized the major ionic currents and use the ionic data to simulate action potentials (APs) recorded from the isolated PCs. Methods and Results Light microscopy was used to isolate and identify PCs from apical and septal cells. Current and voltage clamp techniques were used to record APs and whole cell currents. We simulated a PC action potential, based on our experimental data. APs recorded from PCs were significantly longer than those recorded from ventricular cells. The prominent plateau phase of the PC AP was very negative (~−40mV). Spontaneous activity was observed only in PCs. The inward rectifier current, IK1, demonstrated no significant differences compared to ventricular myocytes (VMs). However, sodium current density was larger, and the voltage-gated potassium current (Ito) density was significantly less in PCs compared to myocytes. T-Type Ca2+ currents (ICa-T) were present in PCs but not VMs. Computer simulations suggest that ICa-T and cytosolic calcium diffusion significantly modulate AP profile recorded in PCs, as compared to VMs. Conclusions Our study provides the first comprehensive ionic profile of murine PCs. The data show unique features of PC ionic mechanisms that govern its excitation process. Experimental data and numerical modeling results suggest that a smaller Ito and the presence of the ICa-T are important determinants of the longer and relatively negative plateau phase of the APs.

IntroductionStudies were performed to assess whether ATP-sensitive K+ (KATP) channels on rabbit preglomerular vessels can influence afferent arteriolar (AA) tone. K+ channels with a slope conductance of 258±13 (n = 7) pS and pronounced voltage dependence were demonstrated in excised patches from vascular smooth muscle cells of microdissected preglomerular segments. Channel activity was markedly reduced by 1 mM ATP and in a dose-dependent fashion by glibenclamide (10-9 M to 10-6 M), a specific antagonist of KAT channels. 10 -M diazoxide, a K+ channel opener, activated these channels in the presence of ATP, and this effect was also blocked by glibenclamide. To determine the role of these KAT channels in the con-

Rationale:The rapid delayed rectifier potassium current, I Kr , which flows through the human ether-a-go-gorelated (hERG) channel, is a major determinant of the shape and duration of the human cardiac action potential (APD). However, it is unknown whether the time dependency of I Kr enables it to control APD, conduction velocity (CV), and wavelength (WL) at the exceedingly high activation frequencies that are relevant to cardiac reentry and fibrillation.Objective: To test the hypothesis that upregulation of hERG increases functional reentry frequency and contributes to its stability. Methods and Results:Using optical mapping, we investigated the effects of I Kr upregulation on reentry frequency, APD, CV, and WL in neonatal rat ventricular myocyte (NRVM) monolayers infected with GFP (control), hERG (I Kr ), or dominant negative mutant hERG G628S. Reentry frequency was higher in the I Kr -infected monolayers (21.12؎0.8 Hz; n‫34؍‬ versus 9.21؎0.58 Hz; n‫;61؍‬ P<0.001) but slightly reduced in G628S-infected monolayers. APD 80 in the I Kr -infected monolayers was shorter (>50%) than control during pacing at 1 to 5 Hz. CV was similar in both groups at low frequency pacing. In contrast, during high-frequency reentry, the CV measured at varying distances from the center of rotation was significantly faster in I Kr -infected monolayers than controls. Simulations using a modified NRVM model predicted that rotor acceleration was attributable, in part, to a transient hyperpolarization immediately following the AP. The transient hyperpolarization was confirmed experimentally. Key Words: hERG Ⅲ delayed rectifier potassium channel Ⅲ reentry Ⅲ ventricular fibrillation Ⅲ optical mapping V entricular fibrillation (VF) is a major cause of sudden cardiac death. Evidence suggests that, in mammalian species ranging in size from the mouse to human, VF may be maintained by highly periodic reentrant waves called rotors. 1,2 The spiral waves that emanate from high-frequency rotors propagate through the ventricles and undergo intermittent, spatially distributed wavebreaks, resulting in complex patterns of conduction also known as fibrillatory conduction. 1,3 Although the existence of rotors has been known for years, the ionic mechanisms responsible for their behavior remain incompletely understood. Recently, it was shown that the frequency and the stability of rotors are determined in great measure by the resting membrane potential (RMP) and the conduction velocity (CV), which are controlled by the dynamic interplay between the outward component of the inward rectifier potassium current (I K1 ) and the rapid inward sodium current (I Na ). 4 Other experiments have demonstrated that overexpression of the slow delayed rectifier current (I Ks ) does not affect reentry frequency but significantly increases the incidence of fibrillatory conduction in cardiomyocyte monolayers. 5 However, the consequences of upregulation or gain of function in the rapid delayed rectifier current (I Kr ) on reentry frequency and dynamics has never been directly i...