Prolonged hyperglycaemia impairs vascular reactivity and inhibits voltage-activated K(+) (Kv) channels. We examined acute effects of altering glucose concentration on the activity and inhibition by endothelin-1 (ET-1) of Kv currents of freshly isolated rat arterial myocytes. Peak Kv currents recorded in glucose-free solution were reversibly reduced within 200 s by increasing extracellular glucose to 4 mm. This inhibitory effect of glucose was abolished by protein kinase C inhibitor peptide (PKC-IP), and Kv currents were further reduced in 10 mm glucose. In current-clamped cells, membrane potentials were more negative in 4 than in 10 mm glucose. In 4 mm d-glucose, 10 nm ET-1 decreased peak Kv current amplitude at +60 mV from 23.5 +/- 3.3 to 12.1 +/- 3.1 pA pF(-1) (n = 6, P < 0.001) and increased the rate of inactivation, decreasing the time constant around fourfold. Inhibition by ET-1 was prevented by PKC-IP. When d-glucose was increased to 10 mm, ET-1 no longer inhibited Kv current (n = 6). Glucose metabolism was required for prevention of ET-1 inhibition of Kv currents, since fructose mimicked the effects of d-glucose, while l-glucose, sucrose or mannitol were without effect. Endothelin receptors were still functional in 10 mm d-glucose, since pinacidil-activated ATP-dependent K(+) (K(ATP)) currents were reduced by 10 nm ET-1. This inhibition was nearly abolished by PKC-IP, indicating that endothelin receptors could still activate PKC in 10 mm d-glucose. These results indicate that changes in extracellular glucose concentration within the physiological range can reduce Kv current amplitude and can have major effects on Kv channel modulation by vasoconstrictors.

We demonstrate the synergistic benefits of using multiple technologies to investigate complex multi-scale biological responses. The combination of reductionist and integrative methodologies can reveal novel insights into mechanisms of action by tracking changes of in vivo phenomena to alterations in protein activity (or vice versa). We have applied this approach to electrical and mechanical remodelling in right ventricular failure caused by monocrotaline-induced pulmonary artery hypertension in rats.We show arrhythmogenic T-wave alternans in the ECG of conscious heart failure animals. Optical mapping of isolated hearts revealed discordant action potential duration (APD) alternans. Potential causes of the arrhythmic substrate; structural remodelling and/or steep APD restitution and dispersion were observed, with specific remodelling of the Right Ventricular Outflow Tract. At the myocyte level, [Ca2+]i transient alternans were observed together with decreased activity, gene and protein expression of the sarcoplasmic reticulum Ca2+-ATPase (SERCA). Computer simulations of the electrical and structural remodelling suggest both contribute to a less stable substrate.Echocardiography was used to estimate increased wall stress in failure, in vivo. Stretch of intact and skinned single myocytes revealed no effect on the Frank-Starling mechanism in failing myocytes. In isolated hearts acute stretch-induced arrhythmias occurred in all preparations. Significant shortening of the early APD was seen in control but not failing hearts. These observations may be linked to changes in the gene expression of candidate mechanosensitive ion channels (MSCs) TREK-1 and TRPC1/6. Computer simulations incorporating MSCs and changes in ion channels with failure, based on altered gene expression, largely reproduced experimental observations.

Inward rectifier potassium current (IK1) and Kir2 composition of the zebrafish (Danio rerio) heart drKir2.1a, drKir2.1b, drKir2.2a, drKir2.2b, drKir2.3, and drKir2.4) were expressed in the zebrafish heart. drKir2.4 and drKir2.2a were the dominant isoforms in both the ventricle (92.9 ± 1.5 and 6.3 ± 1.5 %) and the atrium (28.9 ± 2.9 and 64.7 ± 3.0 %). The remaining four channels comprised together less than 1 and 7 % of the total transcripts in ventricle and atrium, respectively. The four main gene products (drKir2.1a, drKir2.2a, drKir2.2b, drKir2.4) were cloned, sequenced, and expressed in HEK cells for electrophysiological characterization. drKir2.1a was the most weakly rectifying (passed more outward current) and drKir2.2b the most strongly rectifying (passed less outward current) channel, while drKir2.2a and drKir2.4were intermediate between the two. In regard to sensitivity to Ba 2+ block, drKir2.4 was the most sensitive (IC50 = 1.8 μM) and drKir2.1a the least sensitive channel (IC50 = 132 μM). These findings indicate that the Kir2 isoform composition of the zebrafish heart markedly differs from that of the mammalian hearts. Furthermore orthologous Kir2 channels (Kir2.1 and Kir2.4) of zebrafish and mammals show striking differences in Ba 2+ -sensitivity. Those structural and functional differences needs to be taken into account when zebrafish is used as a model for human cardiac electrophysiology, cardiac diseases, and in screening cardioactive substances. AUTHOR'S PROOF!49 Keywords separated by ' -'Zebrafish -Heart -Inward rectifier potassium current -Kir2 channel 50 Foot note information ION CHANNELS, RECEPTORS AND TRANSPORTERS

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