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Venom-derived peptide modulators of ion channel gating are regarded as essential tools for understanding the molecular motions that occur during the opening and closing of ion channels. In this study, we present the characterization of five spider toxins on 12 human voltage-gated ion channels, following observations about the target promiscuity of some spider toxins and the ongoing revision of their "canonical" gating-modifying mode of action. The peptides were purified de novo from the venom of Grammostola rosea tarantulas, and their sequences were confirmed by Edman degradation and mass spectrometry analysis. Their effects on seven tetrodotoxin-sensitive Na+ channels, the three human ether-a-go-go (hERG)-related K+ channels, and two human Shaker-related K+ channels were extensively characterized by electrophysiological techniques. All the peptides inhibited ion conduction through all the Na+ channels tested, although with distinctive patterns. The peptides also affected the three pharmaceutically relevant hERG isoforms differently. At higher concentrations, all peptides also modified the gating of the Na+ channels by shifting the activation to more positive potentials, whereas more complex effects were recorded on hERG channels. No effects were evident on the two Shaker-related K+ channels at concentrations well above the IC50 value for the affected channels. Given the sequence diversity of the tested peptides, we propose that tarantula toxins should be considered both as multimode and target-promiscuous ion channel modulators; both features should not be ignored when extracting mechanistic interpretations about ion channel gating. Our observations could also aid in future structure-function studies and might help the development of novel ion channel-specific drugs.

A common feature of tumor cells is the aberrant expression of ion channels on their plasma membrane. The molecular mechanisms regulating ion channel expression in cancer cells are still poorly known. K+ channels that belong to the human ether-a-go-go-related gene 1 (herg1) family are frequently misexpressed in cancer cells compared to their healthy counterparts. We describe here a posttranslational mechanism for the regulation of hERG1 channel surface expression in cancer cells. This mechanism is based on the activity of hERG1 isoforms containing the USO exon. These isoforms (i) are frequently overexpressed in human cancers, (ii) are retained in the endoplasmic reticulum, and (iii) form heterotetramers with different proteins of the hERG family. (iv) The USO-containing heterotetramers are retained intracellularly and undergo ubiquitin-dependent degradation. This process results in decreased hERG1 current (IhERG1) density. We detailed such a mechanism in heterologous systems and confirmed its functioning in tumor cells that endogenously express hERG1 proteins. The silencing of USO-containing hERG1 isoforms induces a higher IhERG1 density in tumors, an effect that apparently regulates neurite outgrowth in neuroblastoma cells and apoptosis in leukemia cells. Copyright © 2008, American Society for Microbiology. All Rights Reserved.

OBJECTIVES: The present study was designed to determine the effects of long-term antialdosterone treatment on cardiac structural and functional alterations, portal and systemic hemodynamic as well as adrenergic dysfunction characterizing Child A cirrhotic patients with F1 esophageal varices. METHODS: Twenty-two Child A postviral preascitic cirrhotic patients were randomly allocated to 200 mg/day K-Canrenoate (13 patients, age 59.6 +/- 2.2 yr, mean + SEM) or no-drug treatment (9 patients, age 61.8 +/- 2.3) for a 6-month-period. Measurements, which included hepatic venous pressure gradient (HVPG), left ventricular wall thickness, left ventricular end-diastolic volume and diastolic function (LVWT, LVEDV, and E/A ratio, echocardiography), and muscle sympathetic nerve activity (MSNA, microneurography, peroneal nerve), were obtained at baseline and following 6 months of drug or no-drug treatment. Ten healthy age-matched subjects served as controls. RESULTS: Cirrhotic patients were characterized by increased HVPG, LVWT, and MSNA values and by a depressed E/A ratio. K-Canrenoate treatment significantly reduced HVPG (from 15.3 +/- 1.0 to 13.8 +/- 0.8 mmHg, p < 0.05), LVWT (from 21.8 +/- 0.5 to 20.7 +/- 0.6 mm, p < 0.02), and LVEDV (from 99.2 +/- 7 to 86.4 +/- 6 ml, p < 0.01), leaving E/A ratio and MSNA almost unaltered. No significant change was observed in the untreated group of cirrhotic patients followed for 6 months without intervention. CONCLUSIONS: These data provide evidence that aldosterone blockade by long-term K-Canrenoate administration improves hepatic hemodynamics by lowering HVPG and ameliorates cardiac structure and function by favoring a reduction in LVWT and LVEDV as well. They also show, however, that this therapeutic intervention neither improves left ventricular diastolic dysfunction nor exerts sympathoinhibitory effects.

A series of stilbenoid analogues of resveratrol (trans-3,4',5-trihydroxystilbene)with a stilbenic or a bibenzylic skeleton have been prepared by partial synthesis from resveratrol and dihydroresveratrol. The synthesized compounds have been evaluated for their ability to modulate voltage-gated channels.

Sea anemones are an important source of various biologically active peptides, and it is known that ATX-II from Anemonia sulcata slows sodium current inactivation. Using six different sodium channel genes (from Nav1.1 to Nav1.6), we investigated the differential selectivity of the toxins AFT-II (purified from Anthopleura fuscoviridis) and Bc-III (purified from Bunodosoma caissarum) and compared their effects with those recorded in the presence of ATX-II. Interestingly, ATX-II and AFT-II differ by only one amino acid (L36A) and Bc-III has 70% similarity. The three toxins induced a low voltage-activated persistent component primarily in the Nav1.3 and Nav1.6 channels. An analysis showed that the 18 dose-response curves only partially fit the hypothesized binding of Lys-37 (sea anemone toxin Anthopleurin B) to the Asp (or Glu) residue of the extracellular IV/S3-S4 loop in cardiac (or nervous) Na+ channels, thus suggesting the substantial contribution of some nearby amino acids that are different in the various channels. As these channels are atypically expressed in mammalian tissues, the data not only suggest that the toxicity is highly dependent on the channel type but also that these toxins and their various physiological effects should be considered prototype models for the design of new and specific pharmacological tools

Native rat lactotrophs express thyrotrophin-releasing hormone-dependent K+ currents consisting of fast and slow deactivating components that are both sensitive to the class III anti-arrhythmic drugs that block the eag-related gene (ERG) K+ current (I(ERG)). Here we describe in MMQ prolactin-releasing pituitary cells the isolation of the slowly deactivating long-lasting component (I(ERGS)), which, unlike the fast component (I(ERGF)), is insensitive to verapamil 2 microm but sensitive to a novel scorpion toxin (ErgTx-2) that hardly affects I(ERGF). The time constants of I(ERGS) activation, deactivation, and recovery from inactivation are more than one order of magnitude greater than in I(ERGF), and the voltage-dependent inactivation is left-shifted by approximately 25 mV. The very slow MMQ firing frequency (approximately 0.2 Hz) investigated in perforated patch is increased approximately four times by anti-arrhythmic agents, by ErgTx-2, and by the abrupt I(ERGS) deactivation. Prolactin secretion in the presence of anti-arrhythmics is three- to fourfold higher in comparison with controls. We provide evidence from I(ERGS) and I(ERGF) simulations in a firing model cell to indicate that only I(ERGS) has an accommodating role during the experimentally observed very slow firing. Thus, we suggest that I(ERGS) potently modulates both firing and prolactin release in lactotroph cells

Overactivity of the sympathetic nervous system and portal hypertension are key factors in the development of ascites in cirrhosis. The sympathoexcitation that characterizes the more advanced stages of liver diseases is less clearly defined in preascitic cirrhosis. We measured sympathetic nerve traffic to skeletal muscle (peroneal nerve) and to skin districts by microneurography in (1) 12 Child class A cirrhotic patients with clinically significant portal hypertension (portal pressure gradient > 10 mm Hg, 14.8 +/- 1.2 mm Hg, mean +/- SEM) but without actual or previous ascites, (2) 16 Child class C cirrhotic patients with tense ascites, and (3) 10 patients with mild congestive heart failure, a condition paradigmatic of a marked sympathetic activation. Muscle sympathetic nerve traffic was markedly increased in Child class C subjects as compared with controls (23.9 +/- 1.6 bursts/min, P <.01) and superimposable to that recorded in heart failure patients (52.9 +/- 4.7 vs. 60.3 +/- 2 bursts/min, P = not significant). Muscle sympathetic nerve traffic was also increased in Child class A subjects (41.6 +/- 2 bursts/min, P <.01 vs. controls) although to a lesser extent (P <.05 vs. Child class C patients). Skin sympathetic nerve traffic was within the normal range in all patients. Neurohormones were all markedly increased in Child class C subjects. Only norepinephrine was increased in Child class A patients. Our data show that sympathetic nerve traffic activation (1) is already detectable in Child class A cirrhosis when clinically significant portal hypertension is present but ascites never developed and (2) is not generalized because although muscle traffic is increased, skin traffic is within normal range. The role of drugs modulating sympathoactivation should be investigated in preascitic cirrhosis.