Use of lithium and SB-415286 to explore the role of glycogen synthase kinase-3 in the regulation of glucose transport and glycogen synthase Katrina MacAulay 1 , Eric Hajduch 1 , Anne S. Blair 1 , Matthew P. Coghlan 2 *, Stephen A. Smith 2 and Harinder S. Hundal 1 1 Division of Molecular Physiology, Faculty of Life Sciences, MSI/WTB Complex, University of Dundee, UK; 2 GlaxoSmithKline, Harlow, UK Glycogen synthase kinase 3 (GSK3) is inactivated by insulin and lithium and, like insulin, Li also activates glycogen synthase (GS) via inhibition of GSK3. Li also mimics insu- lin’s ability to stimulate glucose transport (GT), an obser- vation that has led to the suggestion that GSK3 may coordinate hormonal increases in GT and glycogen synthe- sis. Here we have used Li and SB-415286, a selective GSK3 inhibitor, to establish the importance of GSK3 in the hor- monal activation of GT in terms of its effect on GS in L6 myotubes and 3T3-L1 adipocytes. Insulin, Li and SB-415286 all induced a significant inhibition of GSK3, which was associated with a marked dephosphorylation and activation of GS. In L6 myotubes, SB-415286 induced a much greater activation of GS (6.8-fold) compared to that elicited by insulin (4.2-fold) or Li (4-fold). In adipocytes, insulin, Li and SB-415286 all caused a comparable activation of GS despite a substantial differentiation-linked reduction in GSK3 expression ( 85%) indicating that GSK3 remains an important determinant of GS activation in fat cells. Whilst Li and SB-415286 both inhibit GSK3 in muscle and fat cells, only Li stimulated GT. This increase in GT was not sensitive to inhibitors of PI3-kinase, MAP kinase or mTOR, but was suppressed by the p38 MAP kinase inhibitor, SB-203580. Consistent with this, phosphorylation of p38 MAP kinase induced by Li correlated with its stimulatory effect on GT. Our findings support a crucial role for GSK3 in the regula- tion of GS, but based on the differential effects of Li and SB-415286, it is unlikely that acute inhibition of GSK3 contributes towards the rapid stimulation of GT by insulin in muscle and fat cells. Keywords: adipocyte; muscle; GSK-3; insulin; p38 MAP kinase. One of the major physiological effects of insulin is to promote the uptake, metabolism and storage of glucose in adipose tissue and skeletal muscle [1]. The hormonal regulation of these cellular processes is initiated by the binding of insulin to its receptor and activation of the receptor kinase, which tyrosine phosphorylates intracellular target substrates, in particular insulin receptor substrate 1 (IRS-1) and its relatives IRS-2 and IRS-3 [2–5]. Of the numerous IRS binding proteins, the serine/lipid kinase phosphoinositide 3-kinase (PI3K) has been implicated strongly as a component of the signalling cascade that stimulates glucose transport and glycogen synthesis [6–9]. Another important component of this cascade is protein kinase B (PKB), which lies downstream of PI3K and whose activation is dependent upon phosphorylation of two key amino acid residues, Thr308 and Ser473 [10,11]. 3-Phosphoinositide-dependent kinase (PDK1) phosphory- lates Thr308 [12,13], whereas phosphorylation of Ser473 is thought to be mediated by a separate, as yet unidentified, upstream kinase that has been tentatively called Regulation of Fluid Volume and Composition Regulation of Fluid Volume and Composition Bởi: OpenStaxCollege The major hormones influencing total body water are ADH, aldosterone, and ANH Circumstances that lead to fluid depletion in the body include blood loss and dehydration Homeostasis requires that volume and osmolarity be preserved Blood volume is important in maintaining sufficient blood pressure, and there are nonrenal mechanisms involved in its preservation, including vasoconstriction, which can act within seconds of a drop in pressure Thirst mechanisms are also activated to promote the consumption of water lost through respiration, evaporation, or urination Hormonal mechanisms are activated to recover volume while maintaining a normal osmotic environment These mechanisms act principally on the kidney Volume-sensing Mechanisms The body cannot directly measure blood volume, but blood pressure can be measured Blood pressure often reflects blood volume and is measured by baroreceptors in the aorta and carotid sinuses When blood pressure increases, baroreceptors send more frequent action potentials to the central nervous system, leading to widespread vasodilation Included in this vasodilation are the afferent arterioles supplying the glomerulus, resulting in increased GFR, and water loss by the kidneys If pressure decreases, fewer action potentials travel to the central nervous system, resulting in more sympathetic stimulation-producing vasoconstriction, which will result in decreased filtration and GFR, and water loss Decreased blood pressure is also sensed by the granular cells in the afferent arteriole of the JGA In response, the enzyme renin is released You saw earlier in the chapter that renin activity leads to an almost immediate rise in blood pressure as activated angiotensin II produces vasoconstriction The rise in pressure is sustained by the aldosterone effects initiated by angiotensin II; this includes an increase in Na+ retention and water volume As an aside, late in the menstrual cycle, progesterone has a modest influence on water retention Due to its structural similarity to aldosterone, progesterone 1/7 Regulation of Fluid Volume and Composition binds to the aldosterone receptor in the collecting duct of the kidney, causing the same, albeit weaker, effect on Na+ and water retention Cardiomyocytes of the atria also respond to greater stretch (as blood pressure rises) by secreting ANH ANH opposes the action of aldosterone by inhibiting the recovery of Na+ by the DCT and collecting ducts More Na+ is lost, and as water follows, total blood volume and pressure decline In low-pressure states, ANH does not seem to have much effect ADH is also called vasopressin Early researchers found that in cases of unusually high secretion of ADH, the hormone caused vasoconstriction (vasopressor activity, hence the name) Only later were its antidiuretic properties identified Synthetic ADH is still used occasionally to stem life-threatening esophagus bleeding in alcoholics When blood volume drops 5–10 percent, causing a decrease in blood pressure, there is a rapid and significant increase in ADH release from the posterior pituitary Immediate vasoconstriction to increase blood pressure is the result ADH also causes activation of aquaporin channels in the collecting ducts to affect the recovery of water to help restore vascular volume Diuretics and Fluid Volume A diuretic is a compound that increases urine volume Three familiar drinks contain diuretic compounds: coffee, tea, and alcohol The caffeine in coffee and tea works by promoting vasodilation in the nephron, which increases GFR Alcohol increases GFR by inhibiting ADH release from the posterior pituitary, resulting in less water recovery by the collecting duct In cases of high blood pressure, diuretics may be prescribed to reduce blood volume and, thereby, reduce blood pressure The most frequently prescribed anti-hypertensive diuretic is hydrochlorothiazide It inhibits the Na+/ Cl– symporter in the DCT and collecting duct The result is a loss of Na+ with water following passively by osmosis Osmotic diuretics promote water loss by osmosis An example is the indigestible sugar mannitol, which is most often administered to reduce brain swelling after head injury However, it is not the only sugar that can produce a diuretic effect In cases of poorly controlled diabetes mellitus, glucose levels exceed the capacity of the tubular glucose symporters, resulting in glucose in the urine The unrecovered glucose becomes a powerful osmotic diuretic Classically, in the days before glucose could be detected in the blood and urine, clinicians identified diabetes mellitus by the three Ps: polyuria (diuresis), polydipsia (increased thirst), and polyphagia (increased hunger) 2/7 Regulation of Fluid Volume and Composition Regulation of Extracellular Na+ Sodium has a very strong osmotic effect and attracts water It plays a larger role in the osmolarity of the plasma than any other ... BANK HOLDING COMPANY ACT Characteristics and Regulation of Exempt Institutions and the Implications of Removing the Exemptions Report to Congressional Committees January 2012 GAO-12-160 United States Government Accountability Office GAO United States Government Accountability Office Highlights of GAO-12-160, a report to congressional committees January 2012 BANK HOLDING COMPANY ACT Characteristics and Regulation of Exempt Institutions and the Implications of Removing the Exemptions Why GAO Did This Study The Bank Holding Company Act of 1956 (BHC Act) establishes the legal framework under which bank holding companies—that is, companies which own or control banks—operate and restricts the type of activities that these companies may conduct. The BHC Act excludes from these restrictions certain companies because the financial institutions they own are exempt from the BHC Act definition of “bank”. However, these exempt institutions are eligible for FDIC insurance raising questions about continuing to exempt their holding companies from BHC Act requirements. The Dodd-Frank Wall Street Reform and Consumer Protection Act directs GAO to study the implications of removing the exemptions. This report examines (1) the number and general characteristics of certain institutions in the U.S. banking system that are exempt from the definition of bank in the BHC Act, (2) the federal regulatory system for exempt financial institutions, and (3) potential implications of subjecting the holding companies of exempt institutions to BHC Act requirements. GAO analyzed data and exams from exempt institutions and regulators, and examined regulators’ guidance and policies. GAO also interviewed regulators and officials from 31 exempt financial institutions. We provided a draft of this report to the relevant agencies. Treasury provided written comments and we received technical comments from other agencies which we incorporated as appropriate. What GAO Found The 1,002 exempt financial institutions make up a small percentage of the assets of the overall banking system—about 7 percent—and include industrial loan corporations (ILC), limited-purpose credit card banks, municipal deposit banks, trust banks with insured deposits, and savings and loans (S&L). Although exempt from the BHC Act, S&L holding companies are regulated by the Federal Reserve System Board of Governors (Federal Reserve) under the Home Owners’ Loan Act as amended. Excluding S&Ls, the number of exempt institutions drops to 57 that comprise less than 1 percent of banking system assets and there is a 3-year moratorium on the approval of federal deposit insurance on select exempt institutions that ends in 2013. These institutions vary by size, activities, and risks. Larger institutions such as ILCs provide banking services similar to those of commercial banks and carry many of the same risks. Other exempt institutions are smaller, provide only a few services such as credit card loans and related services, and thus have lower risk profiles. Federal regulation of the holding companies of exempt institutions and their affiliates varies. The Federal Deposit Insurance Corporation (FDIC) and Office of the E2A participates in a fine control of pre-mature B-cell apoptosis mediated by B-cell receptor signaling via transcriptional regulation of survivin, IAP2 and caspase-8 genes Kenji Toyonaga 1,2, *, Hidehiko Kikuchi 1,3, *, Koki Yamashita 1 , Masami Nakayama 1 , Kazuo Chijiiwa 2 and Tatsuo Nakayama 1,3 1 Section of Biochemistry and Molecular Biology, Department of Medical Sciences, Miyazaki Medical College, University of Miyazaki, Japan 2 Section of Surgical Oncology and Regulation of Organ Function, Department of Medical Science, Miyazaki Medical College, University of Miyazaki, Japan 3 Department of Life Science, Frontier Science Research Center, University of Miyazaki, Japan It is widely known that B lymphocytes are essential to immune responses in health and disease, and maintain homeostasis by balancing cell viability and cell death [1]. The molecular mechanisms of B-lymphocyte devel- opment have mostly been studied in mammalian bone marrow and/or peripheral lymphoid tissue (e.g. spleen). Their development requires not only controlled lineage- and locus-specific immunoglobulin gene recombination, establishing unique antigen specificity of the B lympho- cytes, but also developmental stage-specific gene expres- sion participating in lymphoid cell proliferation and synthesis of immune mediators [1–3]. In addition, vari- ous factors and/or signals control various aspects of the normal development of B lymphocytes and func- tion of the immune system. Such developmental activi- ties require numerous transcription factors, i.e. the Keywords apoptosis; B cell; caspase; E2A; survivin Correspondence T. Nakayama, Department of Life Science, Frontier Science Research Center, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan Fax: +81 985 85 6503 Tel: +81 985 85 3127 E-mail: tnakayam@med.miyazaki-u.ac.jp *These authors contributed equally to this work (Received 8 May 2008, revised 16 December 2008, accepted 24 December 2008) doi:10.1111/j.1742-4658.2009.06881.x Antigen binding to the B-cell receptor (BCR) of pre-mature B lymphocytes induces their apoptotic cell death, but binding to the BCR of mature B lymphocytes triggers activation and proliferation. Binding to pre-mature B lymphocytes is thought not only to function as a mechanism to exclude B-cell clones that possess the ability to react with self-antigen, but also to act as a defense mechanism in auto-immune diseases. Cross-linking of BCR of pre-mature B-cell lines, including the chicken DT40 cell line, with anti-immunoglobulin IgG induces apoptotic cell death. Treatment with phorbol 12-myristate 13-acetate/ionomycin, which mimics BCR stimula- tion, is used to study intracellular signal transduction of B lymphocytes. Here, by analyzing the E2A-deficient DT40 cell line, E2A )/) , we show that E2A deficiency prevents certain levels of apoptotic cell death mediated by BCR signaling. In addition, E2A deficiency-linked BCR signaling controls the mimicked pre-mature B-cell apoptosis by PMA/ionomycin through ele- vated survivin plus inhibitor of apoptosis 2 levels, and reduced caspase-3 and caspase-8 activities, resulting in increased amounts of ICAD (inhibitor of caspase-activated DNase), compared with those in the presence of E2A, followed by reduction of DNA fragmentation. These findings will contrib- ute to the resolution of molecular mechanisms of negative selection of B cells and also auto-immune MINIREVIEW Tec family kinases Itk and Rlk ⁄ Txk in T lymphocytes: cross-regulation of cytokine production and T-cell fates Julio Gomez-Rodriguez, Zachary J. Kraus and Pamela L. Schwartzberg National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, USA Introduction Among the key players in intracellular signaling in lym- phocytes are the Tec family kinases (TFKs), which include Tec, Bruton’s tyrosine kinase (Btk), IL-2 induc- ible T-cell kinase (Itk, also known as EMT or TSK), resting lymphocyte kinase (Rlk, also known as Txk) and Bmx (Etk). These kinases are activated by a wide variety of surface receptors including antigen, cytokine, chemokine, G-protein coupled and Toll-like receptors, as well as integrins [1]. Three TFKs are expressed in the T-cell lineage, Itk, Rlk ⁄ Txk and Tec, which are found in both thymocytes and mature T cells. Itk is expressed at the highest levels, followed by Rlk ⁄ Txk and then Tec. Consistent with these levels of expression, Itk has the greatest effects on T-cell function, where it plays a major role in T-cell receptor (TCR) signaling. Although BTK was the first tyrosine kinase associated with a primary immunodeficiency, X-linked agamma- globulinemia in humans and X-linked immunodefi- ciency in mice [1], IL-2 inducible T-cell kinase has only recently been implicated in a human primary genetic immune disorder. A homozygous missense mutation in ITK was found in two patients with a fatal Epstein- Barr Virus-associated lymphoproliferative disorder [2]. Nonetheless, mice deficient in the TFKs Itk or Itk and Rlk ⁄ Txk show altered T-cell development and impaired mature T-cell effector function, highlighting the importance of this family in T cells [1]. In addition, altered expression of Tec kinases has been found in pathological states. Patients with atopic dermatitis, a Th2-mediated disease, exhibit increased Itk expression Keywords cytokines; innate lymphocytes; Itk; PLZF; Rlk ⁄ Txk; T-helper cells; Th1; Th2; Th17; thymus Correspondence P. L. Schwartzberg, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA Fax: +1 301 402-2170 Tel: +1 301 435-1906 E-mail: pams@nhgri.nih.gov (Received 1 September 2010, revised 2 December 2010, accepted 25 February 2011) doi:10.1111/j.1742-4658.2011.08072.x Developing thymocytes and T cells express the Tec kinases Itk, Rlk ⁄ Txk and Tec, which are critical modulators of T-cell receptor signaling, required for full activation of phospholipase Cc, and downstream Ca 2+ and ERK- mediated signaling pathways. Over the last 10 years, data have implicated the Tec family kinases Itk and Rlk ⁄ Txk as important regulators of cyto- kine production by CD4 + effector T-cell populations. Emerging data now suggest that the Tec family kinases not only influence cytokine-producing T-cell populations in the periphery, but also regulate the development of distinct innate-type cytokine-producing T-cell populations in the thymus. Together, these results suggest that the Tec family kinases play critical roles in helping shape immune responses via their effects on the differentiation and function of distinct cytokine-producing, effector T-cell populations. Abbreviations BTK, Bruton’s tyrosine kinase; IFN, interferon; IL, interleukin; iNKT, invariant natural killer T cell; MHC, major histocompatibility complex; NFAT, nuclear MINIREVIEW Galanin-like peptide and the regulation of feeding behavior and energy metabolism Kanako Shiba 1 , Haruaki Kageyama 1 , Fumiko Takenoya 1,2 and Seiji Shioda 1 1 Department of Anatomy, Showa University School of Medicine, Tokyo, Japan 2 Department of Physical Education, Hoshi University School of Pharmacy and Pharmaceutical Science, Tokyo, Japan Introduction Neuropeptides of G protein-coupled receptor (GPCR) ligands are shown to perform a range of physiological functions. Subsequent to the discovery of leptin [1] and ghrelin [2], a number of studies have demonstrated structural and functional characters of appetite-regu- lating neuropeptides, such as orexin, melanin-concen- trating hormone (MCH), neuropeptide Y (NPY), a-melanocyte stimulating hormone (a-MSH) derived from pro-opiomelanocortin (POMC) [3], neuropeptide W [4], relaxin-3 [5] and prolactin-releasing peptide [6]. Galanin is a 29 amino acid peptide that was dis- covered by the detection of its C-terminal amide sequence in porcine intestinal extract in 1983 [7]. The galanin receptors (GALRs) belong to one of the GPCR families and have three known subtypes: GALR1, GALR2 and GALR3. Sixteen years after the discovery of galanin, a galanin-like peptide (GALP) that consists of 60 amino acids was isolated from porcine hypothalamus using a binding assay for GALRs [8]. The 9–21 amino acid sequence of GALP is identical to that of the first 13 amino acids of gala- nin (Fig. 1). However, galanin and GALP are encoded by separate genes that are typically located on separate chromosomes: the GALP gene is located Keywords clinical implication; feeding regulation; galanin; GPCRs leptin; mouse; neuronal network; obesity; rat; thermogenesis Correspondence S. Shioda, Department of Anatomy, Showa University School of Medicine, 1-5-8 Hatanodai, Shinagawa-ku, Tokyo 142- 8555, Japan Fax: +81 3 3784 6815 Tel: +81 3 3784 8103 E-mail: shioda@med.showa-u.ac.jp (Received 14 June 2010, revised 5 September 2010, accepted 12 October 2010) doi:10.1111/j.1742-4658.2010.07933.x The hypothalamic neuropeptides modulate physiological activity via G pro- tein-coupled receptors (GPCRs). Galanin-like peptide (GALP) is a 60 amino acid neuropeptide that was originally isolated from porcine hypo- thalamus using a binding assay for galanin receptors, which belong to the GPCR family. GALP is mainly produced in neurons in the hypothalamic arcuate nucleus. GALP-containing neurons form neuronal networks with several other types of peptide-containing neurons and then regulate feeding behavior and energy metabolism. In rats, the central injection of GALP produces a dichotomous action that involves transient hyperphasia fol- lowed by hypophasia and a reduction in body weight, whereas, in mice, it has only one action that reduces both food intake and body weight. In the present minireview, we discuss current evidence regarding the function of GALP, particularly in relation to feeding and energy metabolism. We also examine the effects of GALP activity on food intake, body weight and locomotor activity after intranasal infusion, a clinically viable mode of delivery. We conclude that GALP may be of therapeutic value for obesity and life-style-related diseases in the near future. Abbreviations ARC, arcuate nuclei; a-MSH, a-melanocyte stimulating hormone; DMH, dorsomedial hypothalamus; GALP, galanin-like peptide; GALR, galanin receptor; GPCR, G protein-coupled receptors; IL-1, interleukin-1; LH, lateral hypothalamus; MCH, melanin-concentrating hormone; MPA, medial preoptic area; NPY, neuropeptide Y; NTS, ... dominant anion of this compartment, and its regulation closely mirrors that of Na+ Regulation of Ca++ and Phosphate The parathyroid glands monitor and respond to circulating levels of Ca++ in the... expiration of CO2, and excretion of acid or base by the kidneys The breakdown of amino acids produces ammonia Most ammonia is converted into less-toxic urea 5/7 Regulation of Fluid Volume and Composition. . .Regulation of Fluid Volume and Composition binds to the aldosterone receptor in the collecting duct of the kidney, causing the same, albeit weaker, effect on Na+ and water retention