Reactivation of the fetal cardiac gene program is a characteristic feature of hypertrophied and failing hearts that correlates with impaired cardiac function and poor prognosis. However, the mechanism governing the reversible expression of fetal cardiac genes remains unresolved. Here we show that neuronrestrictive silencer factor (NRSF), a transcriptional repressor, selectively regulates expression of multiple fetal cardiac genes, including those for atrial natriuretic peptide, brain natriuretic peptide and a-skeletal actin, and plays a role in molecular pathways leading to the re-expression of those genes in ventricular myocytes. Moreover, transgenic mice expressing a dominant-negative mutant of NRSF in their hearts exhibit dilated cardiomyopathy, high susceptibility to arrhythmias and sudden death. We demonstrate that genes encoding two ion channels that carry the fetal cardiac currents I f and I Ca,T , which are induced in these mice and are potentially responsible for both the cardiac dysfunction and the arrhythmogenesis, are regulated by NRSF. Our results indicate NRSF to be a key transcriptional regulator of the fetal cardiac gene program and suggest an important role for NRSF in maintaining normal cardiac structure and function.

Individual types of ion channels play a unique role in generating membrane excitation based on their gating and conductance properties. The contribution of a given ion channel has been extensively discussed in original experimental papers. However, the complicated interactions of more than 10 ionic current systems through a common membrane potential make it difficult to clarify their roles in membrane excitability. ; I ext , current applied through the electrode (pA); I ha , hyperpolarization-activated cation current (pA); I Kl , inward rectifier K ϩ current (pA); I KACh , ACh-activated K ϩ current (pA); I KATP , ATP-sensitive K ϩ current (pA); I Kpl , non-specific, voltage-dependent outward current (plateau current) (pA); I Kr , delayed rectifier K ϩ current, rapid component (pA); I Ks , delayed rectifier K ϩ current, slow component (pA); I l , total of background current (time-independent) components (pA); I l(Ca) , Ca 2ϩ -activated background cation current (pA); I Na , Na ϩ current (pA); I NaCa , Na ϩ /Ca 2ϩ exchange current (pA); I NaK , Na ϩ /K ϩ pump current (pA); I net X, whole cell current carried by ion X (pA); I RyR , Ca 2ϩ release through the RyR channel in SR (pA); I SR L, Ca 2ϩ leak from the SR (pA); I SR U, Ca 2ϩ uptake in the SR (pA); I SR T, Ca 2ϩ transfer from the SR uptake site to the release site (pA); I st , sustained inward current (pA); I to , transient outward current (pA); I tot , total current of ion channels and ion exchangers (pA); K mX , Michaelis constant for ion X binding; N, total number of channels; P x , convert factor (pA mM Ϫ1 ); p(X), probability of state X in a multiple states gate; R, gas constant, 8.3143 C mV K Ϫ1 mmol Ϫ1; SA factor, scaling factor for SA node cell sarcoplasmic reticulum (0.03); T, absolute temperature K; T, T*, TCa, TCa*, the 4 states of NL model (1996) ).

The matrix metalloproteinase (MMP) family (B25 members in mammals) has been implicated in extracellular matrix remodeling associated with embryonic development, cancer formation and progression, and various other physiological and pathological events. Inactivating mutations in individual matrix metalloproteinase genes in mice described so far, however, are nonlethal at least up to the first few weeks after birth, suggesting functional redundancy among MMP family members. Here, we report that mice lacking two MMPs, MMP-2 (nonmembrane type) and MT1-MMP (membrane type), die immediately after birth with respiratory failure, abnormal blood vessels, and immature muscle fibers reminiscent of central core disease. In the absence of MMP-2 and MT1-MMP, myoblast fusion in vitro is also significantly retarded. These findings suggest functional overlap in mice between the two MMPs with distinct molecular natures.

The cardiac cell model (Kyoto Model) described in the accompanying paper [1] is developed to simulate membrane excitation and contraction in both ventricular and sinoatrial (SA) node cells using a set of equations common for both cell types. Using the Kyoto model, we aim to clarify the relationship between the role of individual current systems in membrane excitation and their unique gating and conductance properties in the SA node cell. So far, the contribution of various time-and voltage-dependent current systems has been evaluated simply by comparing the magnitude of individual currents or by examining the effects of excluding the particular current system of interest. In the present study, we reconstruct the spontaneous action potential by varying not only the current size, but also the voltage dependency of the channel gating according to the experimental data. Furthermore, we evaluate the contribution of each current system by introducing a new hypothetical equilibrium potential during the course of pacemaker depolarization. The Kyoto model is compared with the models of Wilders et al. [2], Demir et al. [3], and the Oxsoft SA node model (Oxsoft Heart Program; Biologic, Claix, France). METHODSThe methods have been fully described in the accompanying paper [1]. The gating and conductance properties of ion channels are common for both the ventricular and SA node cell versions.The sequential numbers of equations and tables and abbreviations referred to in the present paper indicate those in the accompanying paper [1].

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These results suggest that the novel enhancer CNS13 and MEF2 may play a critical role in the transcription of Hcn4 in the heart.

Types and distributions of inwardly rectifying potassium (Kir) channels are one of the major determinants of the electrophysiological properties of cardiac myocytes. Kir2.1 (classical inward rectifier K(+) channel), Kir6.2/SUR2A (ATP-sensitive K(+) channel) and Kir3.1/3.4 (muscarinic K(+) channels) in cardiac myocytes are commonly upregulated by a membrane lipid, phosphatidylinositol 4,5-bisphosphates (PIP(2)). PIP(2) interaction sites appear to be conserved by positively charged amino acid residues and the putative alpha-helix in the C-terminals of Kir channels. PIP(2) level in the plasma membrane is regulated by the agonist stimulation. Kir channels in the cardiac myocytes seem to be actively regulated by means of the change in PIP(2) level rather than by downstream signal transduction pathways.

The mean sarcomere length (SL) of guinea-pig cardiac myocytes was recorded simultaneously with the whole-cell current under voltage-clamp conditions. After blocking both sarcoplasmic reticulum (SR) and L-type Ca(2+) channels with ryanodine, cyclopiazonic acid and nicardipine, strong depolarizing pulses induced only the tonic component of SL shortening through the reverse mode of Na(+)/Ca(2+) exchange (NCX). A positive staircase of SL shortening was observed on applying a train pulses to +60~+100 mV at 2 Hz and trans-membrane Ca(2+) flux was calculated from the time integral of the Na(+)/Ca(2+) exchange current ( I(NCX)). Changes in cytosolic [Ca(2+)] ([Ca(2+)](i)) were determined indirectly using the experimental [Ca(2+)](i)/SL relationship. Cellular Ca(2+) buffering was characterized by a lumped single-component system with a maximum binding capacity of 200 micro M and a dissociation constant of 613 nM. Despite the decrease in driving force, the amplitude of the outwards I(NCX) at +60 mV gradually increased along with the positive staircase. The model simulation suggested that this increase of outwards I(NCX) is caused by a dramatic increase in Ca(2+)-mediated activation of NCX.