1 The possible role of sarcoplasmic reticulum (SR) Ca 2+ stores in hypoxic pulmonary vasoconstriction (HPV) is not well understood. In order to assess the possible role of intracellular Ca 2+ release from SR Ca 2+ stores in HPV, we examined the e ects of: (1) ryanodine (10 mM) depletion of intracellular Ca 2+ stores, and (2) thapsigargin (THAPS, 2 mM) or cyclopiazonic acid (CPA, 10 mM) depletion of intracellular Ca 2+ stores on HPV in canine pulmonary artery. 2 Isometric tension was measured from arterial ring suspended in Krebs-Henseliet solution (K-H) bubbled with 95%O 2 /5%CO 2 . Hypoxia was induced by bubbling phenylephrine (PE, 1 mM) precontracted rings with 95%N 2 /5%CO 2 . HPV was observed in both intact and endothelial-denuded arteries and expressed as % of maximal KCl contraction (%T kmax )=21.3+3.2%; n=13 and 21.7+4%; n=4, respectively. 3 When SR ca eine sensitive Ca 2+ stores were depleted by pretreatment with ryanodine and brief ca eine (15 mM) exposure, the hypoxic response was signifcantly reduced to 19.1+9.2% of the control hypoxic contraction (n=7; P50.001) with little or no e ect on PE or KCl contractions. On the other hand, in normoxic rings pretreated with THAPS or CPA, the PE responses were signi®cantly reduced (%T kmax =18.2+3.1% compared to 39.0+3.9% in control; n=16; P50.001; %T kmax =3.4+1.6% compared to 49.9+7.9% in control; n=6; P50.001; respectively) with no signi®cant e ect on ca eineinduced contractions, suggesting that both THAPS and CPA preferentially deplete InsP 3 -sensitive Ca 2+ stores, without a ecting the ca eine-sensitive Ca 2+ store; consistent with the existence of separate and independent InsP 3 and ca eine-sensitive Ca 2+ stores in this preparation. 4 When hypoxia was induced in the presence of THAPS or CPA, developed tension was signi®cantly larger than control (%T kmax =64.5+6.0%; n=16; P50.05%; %T kmax =78.2+15%; n=6; P50.05; respectively), was partially blocked by nisoldipine (10 mM) and ryanodine (%T kmax =20.3+3.7%; n=6), and nearly completely blocked by SK&F 96365 (50 mM). However, the actions of SK&F 96365 appeared to be nonselective since this compound also signi®cantly reduced contractions elicited by KCl, PE and ca eine. 5 Finally, evidence was obtained suggesting: (a) that at least some of the Ca 2+ released from the ca eine-and ryanodine-sensitive Ca 2+ stores by hypoxia may be taken up and bu ered by the InsP 3 -sensitive Ca 2+ stores, and (b) the apparent dependence of HPV on extracellular Ca 2+ entry pathways may be partially due to the dependence of the Ca 2+ content of intracellular SR Ca 2+ stores on sarcolemmal Ca 2+ entry pathways. 6 These data suggest that ca eine-and ryanodine-sensitive SR Ca 2+ stores contribute signi®cantly to HPV under normal conditions and, in the presence of THAPS or CPA, an additional nisoldipine-and ryanodine-insensitive Ca 2+ entry pathway is evoked by hypoxia.

Circadian rhythms organize many aspects of cell biology and physiology to a daily temporal program that depends on clock gene expression cycles in most mammalian cell types. However, circadian rhythms are also observed in isolated mammalian red blood cells (RBCs), which lack nuclei, suggesting the existence of post-translational cellular clock mechanisms in these cells. Here we show using electrophysiological and pharmacological approaches that human RBCs display circadian regulation of membrane conductance and cytoplasmic conductivity that depends on the cycling of cytoplasmic K+ levels. Using pharmacological intervention and ion replacement, we show that inhibition of K+ transport abolishes RBC electrophysiological rhythms. Our results suggest that in the absence of conventional transcription cycles, RBCs maintain a circadian rhythm in membrane electrophysiology through dynamic regulation of K+ transport.

Background-Gap junction resistivity, R j , has been proposed as a key determinant of conduction velocity (CV). However, studies in connexin-gene knockout mice demonstrated significant CV slowing only with near-complete connexin deletion, and these findings led to the concept of a significant redundancy of myocardial gap junctions. We challenged this prevailing concept and addressed the hypothesis that there is a continuous relationship between R j and CV, each independently measured in human and guinea-pig myocardium. Methods and Results-R j and CV were directly measured by oil-gap impedance and microelectrode techniques in human left ventricular myocardium from patients with hypertrophic cardiomyopathy and in guinea-pig atrial and ventricular myocardium before and during pharmacological uncoupling with 20-µmol/L carbenoxolone. There was a continuous relationship between R j and CV in human and guinea-pig myocardium, pre-and post-carbenoxolone (r 2 =0.946; P<0.01). In guinea-pig left ventricle, left atrium, and right atrium, carbenoxolone increased R j by 28±9%, 26±16%, and 25±14% and slowed CV by 17±3%, 23±8%, and 11±4% respectively (all P<0.05 versus control). As a clinically accessible measure of local microscopic myocardial conduction slowing in vivo in the intact human heart, carbenoxolone prolonged electrogram duration in the right atrium (39.7±4.2 to 42.3±4.3 ms; P=0.01) and right ventricle (48.1±2.5 to 53.3±5.3 ms; P<0.01). Conclusions-There is a continuous relationship between R j and CV that is consistent between cardiac chambers and across species, indicating that naturally occurring variations in cellular coupling can account for variations in CV, and that the concept that there is massive redundancy of coupling is not tenable. (Circ Arrhythm Electrophysiol. 2013;6:1208-1214.)

The effect of differential signalling by IL-6 and leukaemia inhibitory factor (LIF) which signal by gp130 homodimerisation or LIFRβ/gp130 heterodimerisation on survival and hypertrophy was studied in neonatal rat cardiomyocytes. Both LIF and IL-6 [in the absence of soluble IL-6 receptor (sIL-6Rα)] activated Erk1/2, JNK1/2, p38-MAPK and PI3K signalling peaking at 20 min and induced cytoprotection against simulated ischemia-reperfusion injury which was blocked by the MEK1/2 inhibitor PD98059 but not the p38-MAPK inhibitor SB203580. In the absence of sIL-6R, IL-6 did not induce STAT1/3 phosphorylation, whereas IL-6/sIL-6R and LIF induced STAT1 and STAT3 phosphorylation. Furthermore, IL-6/sIL-6R induced phosphorylation of STAT1 Tyr701 and STAT3 Tyr705 were enhanced by SB203580. IL-6 and pheneylephrine (PE), but not LIF, induced cardiomyocyte iNOS expression and nitric oxide (NO) production. IL-6, LIF and PE induced cardiomyocyte hypertrophy, but with phenotypic differences in ANF and SERCA2 expression and myofilament organisation with IL-6 more resembling PE than LIF. Transfection of cardiomyocytes with full length or truncated chimaeric gp130 cytoplasmic domain/Erythropoietin receptor (EpoR) extracellular domain fusion constructs showed that the membrane proximal Box 1 and Box 2 containing region of gp130 was necessary and sufficient for MAPK and PI3K activation; hypertrophy; SERCA2 expression and iNOS/NO induction in the absence of JAK/STAT activation. In conclusion, IL-6 can signal in cardiomyocytes independent of sIL-6R and STAT1/3 and furthermore, that Erk1/2 and PI3K activation by IL-6 are both necessary and sufficient for induced cardioprotection. In addition, p38-MAPK may act as a negative feedback regulator of JAK/STAT activation in cardiomyocytes.