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The cardiac late sodium current (INa) has been in the focus of research in the recent decade. The first reports on the sustained component of voltage activated sodium current date back to the seventies, but early studies interpreted this tiny current as a product of a few channels that fail to inactivate, having neither physiologic nor pathologic implications. Recently, the cardiac INa has emerged as a potentially major arrhythmogenic mechanism in various heart diseases, attracting the attention of clinicians and researchers. Research activity on INa has exponentially increased since Ranolazine, an FDA-approved antianginal drug was shown to successfully suppress cardiac arrhythmias by inhibiting INa. This review aims to summarize and discuss a series of papers focusing on the cardiac late sodium current and its regulation under physiological and pathological conditions. We will discuss critical evidences implicating INa as a potential target for treating myocardial dysfunction and cardiac arrhythmias.

Receptor-mediated changes in intracellular cyclic AMP concentration play critical role in the autonomic control of the heart, including regulation of a variety of ion channels via mechanisms involving protein kinase A, EPAC, or direct actions on cyclic nucleotide gated ion channels. In case of any ion channel, the actual signal transduction cascade can be identified by using properly modified cAMP derivatives with altered binding and activating properties. In this study we focus to structural modifications of cAMP resulting in specific activator and blocking effects on PKA or EPAC. Involvement of the cAMP-dependent signal transduction pathway in controlling rapid delayed rectifier K current was studied in canine ventricular myocytes using these specific cAMP analogues. Adrenergic stimulation increased the density of IKr in canine ventricular cells, which effect was mediated by a PKA-dependent but EPAC-independent pathway. It was also shown that intracellular application of large concentrations of cAMP failed to fully activate PKA comparing to the effect of isoproterenol, forskolin, or PDE-resistant cAMP derivatives. This difference was fully abolished following inhibition of phosphodiesterase by IBMX. These results are in line with the concept of compartmentalized release, action, and degradation of cAMP within signalosomes.

Action potential voltage-clamp (APVC) is a technique to visualize the profile of various currents during the cardiac action potential. This review summarizes potential applications and limitations of APVC, the properties of the most important ion currents in nodal, atrial, and ventricular cardiomyocytes. Accordingly, the profiles (“fingerprints”) of the major ion currents in canine ventricular myocytes, i.e. in cells of a species having action potential morphology and set of underlying ion currents very similar to those found in the human heart, are discussed in details. The degree of selectivity of various compounds, which is known to be a critical property of drugs used in APVC experiments, is overviewed. Thus the specificity of agents known to block sodium (tetrodotoxin, saxitoxin), potassium (chromanol 293B, HMR 1556, E-4031, dofetilide, sotalol, 4-aminopyridine, BaCl2), calcium (nifedipine, nisolpidine, nicardipine, diltiazem, verapamil, gallopamil), and chloride (anthracene-9-carboxylic acid, DIDS) channels, the inhibitor of the sodium-calcium exchanger (SEA0400), and the activator of sodium current (veratridine) are accordingly discussed. Based on a theory explaining how calcium current inhibitors block calcium channels, the structural comparison of the studied substances usually confirmed the results of the literature. Using these predictions, a hypothetical super-selective calcium channel inhibitor structure was designed. APVC is a valuable tool not only for studying the selectivity of the known ion channel blockers, but is also suitable for safety studies to exclude cardiac ion channel actions of any agent under development.

Calcium ions are crucial elements of excitation-contraction coupling in cardiac myocytes. The intracellular Ca2 concentration changes continously during the cardiac cycle, but the Ca2 entering to the cell serves as an intracellular second messenger, as well. The Ca2 as a second messenger influences the activity of many intracellular signalling pathways and regulates gene expression. In cardiac myocytes the major pathway for Ca2 entry into cells is L-type calcium channel (LTCC). The precise control of LTCC function is essential for maintaining the calcium homeostasis of cardiac myocytes. Dysregulation of LTCC may result in different diseases like cardiac hypertrophy, arrhytmias, heart failure. The physiological and pathological structural changes in the heart are induced in part by small G proteins. These proteins are involved in wide spectrum of cell biological functions including protein transport, regulation of cell proliferation, migration, apoptosis, and cytoskeletal rearrangement. Understanding the crosstalk between small G proteins and LTCC may help to understand the pathomechanism of different cardiac diseases and to develop a new generation of genetically-encoded Ca2 channel inhibitors.

Therapeutic strategy for cardiac arrhythmias has undergone a remarkable change during the last decades. Currently implantable cardioverter defibrillator therapy is considered to be the most effective therapeutic method to treat malignant arrhythmias. Some even argue that there is no room for antiarrhythmic drug therapy in the age of implantable cardioverter defibrillators. However, in clinical practice, antiarrhythmic drug therapies are frequently needed, because implantable cardioverter defibrillators are not effective in certain types of arrhythmias (i.e. premature ventricular beats or atrial fibrillation). Furthermore, given the staggering cost of device therapy, it is economically imperative to develop alternative effective treatments. Cardiac ion channels are the target of a number of current treatment strategies, but therapies based on ion channel blockers only resulted in moderate success. Furthermore, these drugs are associated with an increased risk of proarrhythmia, systemic toxicity, and increased defibrillation threshold. In many cases, certain ion channel blockers were found to increase mortality. Other drug classes such as blockers, angiotensin-converting enzyme inhibitors, aldosterone antagonists, and statins appear to have proven efficacy for reducing cardiac mortality. These facts forced researchers to shift the focus of their research to molecular targets that act upstream of ion channels. One of these potential targets is calcium/calmodulin-dependent kinase II (CaMKII). Several lines of evidence converge to suggest that CaMKII inhibition may provide an effective treatment strategy for heart diseases. (1) Recent studies have elucidated that CaMKII plays a key role in modulating cardiac function and regulating hypertrophy development. (2) CaMKII activity has been found elevated in the failing hearts from human patients and animal models. (3) Inhibition of CaMKII activity has been shown to mitigate hypertrophy, prevent functional remodeling and reduce arrhythmogenic activity. In this review, we will discuss the structural and functional properties of CaMKII, the modes of its activation and the functional consequences of CaMKII activity on ion channels.

Currents through single cardiac sodium channels have been measured in inside-out patches from guinea pig ventricular cells. To abolish the fast inactivation, Na channels were modified by DPI 201–106. In symmetrical Na solutions, a diminution of outward sodium currents can be observed that depends on the intracellular magnesium concentration and the membrane potential. Inward currents were not altered by the concentrations of magnesium used (between 0 and 22.5 mmol/1). In Mg free solutions a linear current-voltage relation can also be measured in the range of outward Na currents. At +60 mV (symmetrical Na solutions, single channel conductance 24 pS) a half maximal block of cardiac Na channels by intracellular magnesium was found at 2.1 mmol/l. From the analysis of single channel current-voltage relationships the concentration and voltage-dependent block by intracellular magnesium of cardiac sodium channels could be described as binding of Mg at one site with a Kd value of 5.1 mmol/1 at 0 mV. The site is located at an electrical distance of 0.18 from the inside.

Recent reports indicate that oxidized cobalamin, Cbl(III), can interfere with the biological effects of nitric oxide (NO) on vascular and visceral smooth muscle and in other systems. In attempting to elucidate the mechanism of these effects of Cbl(III), we reported that a Cbl(III)NO complex could be detected by electron paramagnetic resonance (EPR) spectroscopy, but not by ultraviolet/visible spectroscopy. Subsequently, others concluded that the alleged Cbl(III)NO complex is detectable by ultraviolet/visible, but not by EPR spectroscopy and provided ultraviolet/visible evidence for an alleged Cbl(III)NO complex. We report further investigation of the interaction of NO with Cbl, using both techniques, Fourier transform infrared (FTIR) spectroscopy and mass spectrometry. Our EPR results and the UV/VIS results of others appear to be experimental artifacts that can now, at least in part, be explained. Under conditions where FTIR measurements readily detect a N-O stretching frequency of NO bound to Fe(II), we do not detect a similar signal that can be ascribed to either Cbl(III)NO or Cbl(II)NO, indicating that neither Cbl(III) nor Cbl(II) form a stable complex with NO. Loss of the Cbl(II) EPR signal and mass spectral detection of N2O upon addition of NO to Cbl(II) solutions, demonstrates that Cbl(II), which is present in aerobic Cbl(III) solutions, reduces NO; however, this reaction does not appear to be fast enough to account for the observed biological effects in aerated media. Nitric oxide also reacts rapidly and irreversibly with the superoxo complex of Cbl(III), Cbl(III)O2-, which is always present in aerated solutions of Cbl(III). We believe that this latter reaction accounts for the observed inactivation of NO by Cbl(III) in biological systems. Because Cbl(III)O2- is spontaneously regenerated from Cbl(II) and O2 in aerated solutions, this may constitute a cyclic mechanism for the rapid elimination (oxidation) of NO. Thus, several physicochemical techniques fail to provide convincing evidence for the existence of stable Cbl(III)NO or Cbl(II)NO complexes but do provide evidence that Cbl species participate in redox reactions with NO under aerobic conditions, thereby inhibiting its physiological roles.

The muscarinic-activated K+ channel K(ACh), a prototype of channels regulated by neuroendocrine agonists via G proteins, was used to investigate the mode of action of isoflurane and halothane on G protein-coupled signal transduction processes. The evolution of the muscarinic current I(K(ACh)) was characterized through rapid agonist application and washout. At physiologically relevant concentrations, halothane and isoflurane reduced the rate of I(K(ACh)) activation without comparable effects on deactivation. Furthermore, both anesthetics reduced or eliminated the spontaneous decay (rapid desensitization) typical of the muscarinic response. In contrast to these similarities of anesthetic action on the time course of the response, the magnitude of I(K(ACh)) was slowly reduced by isoflurane but rapidly augmented by halothane. Neither halothane nor isoflurane altered the conductance of single I(K(ACh)) channels, indicating that these volatile anesthetics act on channel open-close kinetics. The reduced I(K(ACh)) activation rates suggest that impaired receptor/G protein interactions are induced by both anesthetics. For halothane, the increased amplitude of the response, also seen for I(K(ACh)) activated in a receptor-independent manner by guanosine-5'-O-(3-thio)triphosphate, suggests a direct action on the channel. Alteration of signal transduction processes by halothane and isoflurane may underlie some anesthetic actions of these compounds as well as secondary effects on the cardiovascular system.

We have previously shown that the neural adhesion molecules L1 and NCAM interact with each other to form a complex which binds more avidly to L1 than L1 to L1 alone (Kadmon, G., A. Kowitz, P. Altevogt, and M. Schachner. 1990a. J. Cell Biol. 110:193-208). This cis-association between L1 and NCAM is carbohydrate-dependent (Kadmon, G., A. Kowitz, P. Altevogt, and M. Schachner. 1990b. J. Cell Biol. 110:209-218). In the present study, we report that L1 and NCAM bind to each other via oligomannosidic carbohydrates expressed by L1, but not by NCAM, as shown in several experiments: (a) complex formation between L1 and NCAM is inhibited by a mAb to oligomannosidic carbohydrates and by the oligosaccharides themselves; (b) NCAM binds to oligomannosidic carbohydrates; (c) within the L1/NCAM complex, the oligomannosidic carbohydrates are hidden from accessibility to a mAb against oligomannosidic carbohydrates; (d) the recombinant protein fragment of NCAM containing the immunoglobulin-like domains and not the fragment containing the fibronectin type III homologous repeats binds to oligomannosidic glycans. Furthermore, the fourth immunoglobulin-like domain of NCAM shows sequence homology with carbohydrate recognition domains of animal C-type lectins and, surprisingly, also with plant lectins. A peptide comprising part of the C-type lectin consensus sequence in the fourth immunoglobulin-like domain of NCAM interferes with the association between L1 and NCAM. The functional importance of oligomannosidic glycans at the cell surface was shown for neurite outgrowth in vitro. When neurons from early postnatal mouse cerebellum were maintained on laminin or poly-L-lysine, neurite outgrowth was inhibited by oligomannosidic glycans, by glycopeptides, glycoproteins, or neoglycolipids containing oligomannosidic glycans, but not by nonrelated oligosaccharides or oligosaccharide derivates. Neurite outgrowth was also inhibited by the peptide comprising part of the C-type lectin consensus sequence in the fourth immunoglobulin-like domain of NCAM. The combined results suggest that carbohydrate-mediated cis-associations between adhesion molecules at the cell surface modulate their functional properties.

Expression of the neural adhesion molecule L1 and its potential involvement in axonal sprouting were examined in the deafferented rat dentate gyrus. We focused on the dentate gyrus because of its well-defined cytoarchitecture and well-characterized neuronal degeneration and sprouting response following entorhinal cortex lesions. In the molecular layer of the dentate gyrus, a trilaminar staining pattern was observed, with the middle molecular layer exhibiting slightly denser immunolabeling compared to both inner and outer molecular layers. Two to 12 days after a unilateral entorhinal cortex lesion, a progressive loss of L1 immunolabeling was noted in the ipsilateral middle and outer molecular layers, followed by a substantial reappearance of immunostaining 65 days after lesion incidence. The width of the immunostained ipsilateral inner molecular layer revealed a progressive widening and by postlesion day 65 occupied about 50% of the total width of the molecular layer. Immunoelectron microscopy localized Ll to the surface of unmyelinated axons in both normal and deafferented dentate gyrus. In situhybridization revealed L1 messenger RNA confined to neurons throughout the hippocampul formation, but did not indicate changes in L1 messenger RNA levels in the hippocampus, dentate gyrus, entorhinal cortex or basal forebrain in response to unilateral entorhinal cortex lesions. Changes in Ll immunolabeling in the deafferented dentate gyrus corresponded in a spatial and temporal manner to changes of the synaptic marker synaptophysin and axonal marker phosphorylated tau.

The delayed component of intramembranous charge movement (hump, I gamma) was studied around the contraction threshold in cut skeletal muscle fibers of the frog (Rana esculenta) in a single Vaseline-gap voltage clamp. Charges (Q) were computed as 50-ms integrals of the ON (QON) and OFF (QOFF) of the asymmetric currents after subtracting a baseline. The hump appeared in parallel with an excess of QON over QOFF by approximately 2.5 nC/mu F. Caffeine (0.75 mM) not only shifted the contraction threshold but moved both the hump and the difference between the ON and OFF charges to more negative membrane potentials. When using 10-mV voltage steps on top of different prepulse levels, the delayed component, if present, was more readily observable. The voltage dependences of the ON and OFF charges measured with these pulses were clearly different: QON had a maximum at or slightly above the contraction threshold, while QOFF increased monotonically in the voltage range examined. Caffeine (0.75 mM) shifted this voltage dependence of QON toward more negative membrane potentials, while that of QOFF was hardly influenced. These results show that the delayed component of intramembranous charge movement either is much slower during the OFF than during the ON, or returns to the OFF position during the pulse. Tetracaine (25 microM) had similar effects on the charge movement currents, shifting the voltage dependence on the ON charge in parallel with the contraction threshold, but to more positive membrane potentials, and leaving QOFF essentially unchanged. The direct difference between the charge movement measured in the presence of caffeine and in control solution was either biphasic or resembled the component isolated by tetracaine, suggesting a common site of caffeine and tetracaine action. The results can be understood if the released Ca plays a direct role in the generation of the hump, as proposed in the first paper of this series (Csernoch et al. 1991. J. Gen. Physiol. 97:845-884).

In carbonate‐forming environments, authigenic minerals can cement surface sediments into centimeter‐sized intraclasts that are later reworked into “flat‐pebble” or “edgewise” conglomerates. Flat‐pebble conglomerates comprise only a small portion of facies in modern marine environments but are common in ancient strata, implying that seafloor cements were more widespread in the past. Flat‐pebble conglomerates nearly disappeared after the Ordovician radiation, yet it is unclear if this decline was due to changing seawater chemistry or if increased infaunalization and bioturbation simply worked to break down nascent clasts. We discovered a process analog that produces flat‐pebble conglomerates around the Great Salt Lake, Utah, USA, and studied these facies using field observations, wave models, satellite imagery, petrography, and microanalytic chemical data. Clasts were sourced from wave‐rippled grainstone that cemented in situ in offshore environments. Lake floor cements formed under aragonite saturation states that are lower than modern marine settings, suggesting that physical processes are at least as important as chemical ones. Results from our wave models showed that coarse sediments near the field site experience quiescent periods of up to 6 months between suspension events, allowing isopachous cements to form. Using a simple mathematical framework, we show that the main difference between Great Salt Lake and modern, low‐energy marine settings is that the latter has enough bioturbating organisms to break up clasts. Observations from Great Salt Lake demonstrate how geologic trends in flat‐pebble abundance could largely reflect changes in total infaunal biomass and ecology without requiring regional‐to‐global changes in seawater chemistry. Calcium carbonate is a common sedimentary mineral that precipitates quickly under Earth surface conditions. If carbonate sediments are at rest for long periods of time, they may solidify via cementation into flat, pebble‐sized clasts. Geologic trends show that flat pebbles are anticorrelated with burrows, suggesting they may tell us about animal evolution and ecology. However, it was unknown if animals disrupt clasts through chemical means, leading to slower rates of mineral growth, or if they physically push sediments apart, breaking up nascent cements. We studied flat pebbles forming in Great Salt Lake, Utah, where the water is too salty for burrowing animals. The composition of lake water should lead to slower growth rates for carbonate minerals, yet clasts still form. We conclude that pebbles form during quiescent intervals with little physical energy to move grains, either from waves or from burrowing organisms. As a result, ancient flat pebbles are probably not unique markers for seawater chemistry. Sediment cohesion within carbonate environments depends on both physical and chemical energy near the sediment‐water interfaceIf sediment transport is infrequent, burrowing animals provide a significant source of physical energySubstrate cohesion may record physical changes related to the number and behavior of animals rather than geochemical conditions