It is now well established that stromal interaction molecule 1 (STIM1) is the calcium sensor of endoplasmic reticulum stores required to activate store-operated calcium entry (SOC) channels at the surface of non-excitable cells. However, little is known about STIM1 in excitable cells, such as striated muscle, where the complement of calcium regulatory molecules is rather disparate from that of non-excitable cells. Here, we show that STIM1 is expressed in both myotubes and adult skeletal muscle. Myotubes lacking functional STIM1 fail to show SOC and fatigue rapidly. Moreover, mice lacking functional STIM1 die perinatally from a skeletal myopathy. In addition, STIM1 haploinsufficiency confers a contractile defect only under conditions where rapid refilling of stores would be needed. These findings provide insight into the role of STIM1 in skeletal muscle and suggest that STIM1 has a universal role as an ER/SR calcium sensor in both excitable and non-excitable cells.

Short-term treatment with bFGF enhanced collateral development without increasing neointimal accumulation at sites of vascular injury. Although VEGF did not increase collateral development as administered in this study, it significantly exacerbated neointimal accumulation. These data provide support for the clinical investigation of bFGF in selected patients with ischemic heart disease.

The G protein-coupled receptor kinases (GRKs) and ␤-arrestins, families of molecules essential to the desensitization of G proteindependent signaling via seven-transmembrane receptors (7TMRs), have been recently shown to also transduce G protein-independent signals from receptors. However, the physiologic consequences of this G protein-independent, GRK͞␤-arrestin-dependent signaling are largely unknown. Here, we establish that GRK͞␤-arrestin-mediated signal transduction via the angiotensin II (ANG) type 1A receptor (AT1AR) results in positive inotropic and lusitropic effects in isolated adult mouse cardiomyocytes. We used the ''biased'' AT1AR agonist [Sar 1 , Ile 4 , Ile 8 ]-angiotensin II (SII), which is unable to stimulate G␣q-mediated signaling, but which has previously been shown to promote ␤-arrestin interaction with the AT1AR. Cardiomyocytes from WT, but not AT 1A R-deficient knockout (KO) mice, exhibited positive inotropic and lusitropic responses to both ANG and SII. Responses of WT cardiomyocytes to ANG were dramatically reduced by protein kinase C (PKC) inhibition, whereas those to SII were unaffected. In contrast, cardiomyocytes from ␤-arrestin2 KO and GRK6 KO mice failed to respond to SII, but displayed preserved responses to ANG. Cardiomyocytes from GRK2 heterozygous knockout mice (GRK2 ؉/؊ ) exhibited augmented responses to SII in comparison to ANG, whereas those from GRK5 KO mice did not differ from those from WT mice. These findings indicate the existence of independent G␣q͞PKC-and GRK6͞␤-arrestin2-dependent mechanisms by which stimulation of the AT1AR can modulate cardiomyocyte function, and which can be differentially activated by selective receptor ligands. Such ligands may have potential as a novel class of therapeutic agents.seven-transmembrane receptors ͉ G protein-coupled receptor kinase ͉ mice V irtually all physiologic processes in higher organisms are critically regulated by signal transduction through seventransmembrane receptors (7TMRs), the largest and most diverse family of cell surface receptors (1). The biological effects of 7TMR signal transduction have conventionally been attributed to activation of 7TMR-associated heterotrimeric G proteins, and consequent activation of second messenger-generating enzymes, production of second messengers, and activation of second messenger-dependent effector molecules (2). This classical understanding has been supported by numerous physiologic studies, and forms the basis for the utilization of drugs (agonists and antagonists) targeting 7TMRs, as therapeutic strategies for a vast array of diseases (1).However, this simple paradigm has been altered by an increasing appreciation of the biochemical importance of both negative regulation of G protein-dependent signaling, and G protein-independent signal transduction by 7TMRs (3, 4). G protein activation and dependent downstream processes are predominantly antagonized by the sequential actions of two families of molecules, the G protein-coupled receptor kinases (GRKs) and the ␤-arrestins. Agonist bin...

Rationale: Cardiac muscle adapts to increase workload by altering cardiomyocyte size and function resulting in cardiac hypertrophy. G protein-coupled receptor signaling is known to govern the hypertrophic response through the regulation of ion channel activity and downstream signaling in failing cardiomyocytes. Objective: Transient receptor potential canonical (TRPC) channels are G protein-coupled receptor operated channels previously implicated in cardiac hypertrophy. Our objective of this study is to better understand how TRPC channels influence cardiomyocyte calcium signaling. Methods and Results: Here, we used whole cell patch clamp of adult cardiomyocytes to show upregulation of a nonselective cation current reminiscent of TRPC channels subjected to pressure overload. This TRPC current corresponds to the increased TRPC channel expression noted in hearts of mice subjected to pressure overload. Importantly, we show that mice lacking TRPC1 channels are missing this putative TRPC current. Moreover, Trpc1 ؊/؊ mice fail to manifest evidence of maladaptive cardiac hypertrophy and maintain preserved cardiac function when subjected to hemodynamic stress and neurohormonal excess. In addition, we provide a mechanistic basis for the protection conferred to Trpc1 ؊/؊ mice as mechanosensitive signaling through calcineurin/NFAT, mTOR and Akt is altered in Trpc1 ؊/؊ mice. Conclusions: From these studies, we suggest that TRPC1 channels are critical for the adaptation to biomechanical stress and TRPC dysregulation leads to maladaptive cardiac hypertrophy and failure. (Circ Res. 2009;105:1023-1030.)Key Words: transient receptor potential channels Ⅲ G protein receptor signaling Ⅲ cardiac hypertrophy C ardiac myocytes respond to changing mechanical workloads by altering the frequency and amplitude of their calcium transients. 1,2 Encoded in these calcium transients are signals that alter not only the immediate contractile response but also initiate and maintain a remodeling response that adjusts cellular mass, ionic currents, kinetic properties of contractile proteins, and metabolic capacity. 3 It is likely that persistence of these signals modulate the calcium signaling events resulting in a hypertrophic response and adverse remodeling. To identify the proximal signals that regulate cardiac hypertrophy, attention has begun to focus on ion channels because they may link mechanical activity to cell signaling. 4 Recent work has raised the possibility that hypertrophic agonists linked to G-protein coupled receptors activate calcium entry through transient receptor potential canonical (TRPC) channels. [5][6][7][8][9][10] TRPC channels encompass a large family of nonselective cation channels found in many different cell types. 11 TRPC channels are activated downstream of G-protein receptor through the phospholipase C signaling by inositol trisphosphate (TRPC1/4/5) or by diacyl glycerol (TRPC3/6/7). 5,12 More recently TRPC1/6 channels were found to be mechanosensitive channels that mediate nonselective cation entry in response to i...

Ryanodine receptors (RyRs), intracellular calcium release channels essential for skeletal and cardiac muscle contraction, are also expressed in various types of smooth muscle cells. In particular, recent studies have suggested that in airway smooth muscle cells (ASMCs) provoked by spasmogens, stored calcium release by the cardiac isoform of RyR (RyR2) contributes to the calcium response that leads to airway constriction (bronchoconstriction). Here we report that mouse ASMCs also express the skeletal muscle and brain isoforms of RyRs (RyR1 and RyR3, respectively). In these cells, RyR1 is localized to the periphery near the cell membrane, whereas RyR3 is more centrally localized. Moreover, RyR1 and/or RyR3 in mouse airway smooth muscle also appear to mediate bronchoconstriction caused by the muscarinic receptor agonist carbachol. Inhibiting all RyR isoforms with >200 M ryanodine attenuated the graded carbachol-induced contractile responses of mouse bronchial rings and calcium responses of ASMCs throughout the range of carbachol used (50 nM to >3 M). In contrast, inhibiting only RyR1 and RyR3 with 25 M dantrolene attenuated these responses caused by high (>500 nM) but not by low concentrations of carbachol. These data suggest that, as the stimulation of muscarinic receptor in the airway smooth muscle increases, RyR1 and/or RyR3 also mediate the calcium response and thus bronchoconstriction. Our findings provide new insights into the complex calcium signaling in ASMCs and suggest that RyRs are potential therapeutic targets in bronchospastic disorders such as asthma.