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CaV2.2 (N-type) voltage-gated calcium channels (Ca2+ channels) play key roles in neurons and neuroendocrine cells including the control of cellular excitability, neurotransmitter / hormone secretion, and gene expression. Calcium entry is precisely controlled by channel gating properties including multiple forms of inactivation. \"Fast\" voltage-dependent inactivation is relatively well-characterized and occurs over the tens-to- hundreds of milliseconds timeframe. Superimposed on this is the molecularly distinct, but poorly understood process of \"slow\" voltage-dependent inactivation, which develops / recovers over seconds-to-minutes. Protein kinases can modulate \"slow\" inactivation of sodium channels, but little is known about if/how second messengers control \"slow\" inactivation of Ca2+ channels. We investigated this using recombinant CaV2.2 channels expressed in HEK293 cells and native CaV2 channels endogenously expressed in adrenal chromaffin cells. The PKC activator phorbol 12-myristate 13-acetate (PMA) dramatically prolonged recovery from \"slow\" inactivation, but an inactive control (4α-PMA) had no effect. This effect of PMA was prevented by calphostin C, which targets the C1-domain on PKC, but only partially reduced by inhibitors that target the catalytic domain of PKC. The subtype of the channel β-subunit altered the kinetics of inactivation but not the magnitude of slowing produced by PMA. Intracellular GDP-β-S reduced the effect of PMA suggesting a role for G proteins in modulating \"slow\" inactivation. We postulate that the kinetics of recovery from \"slow\" inactivation could provide a molecular memory of recent cellular activity and help control CaV2 channel availability, electrical excitability, and neurotransmission in the seconds-to-minutes timeframe.

Butanol (C4H10OH) has been used both to dissect the molecular targets of alcohols/general anesthetics and to implicate phospholipase D (PLD) signaling in a variety of cellular functions including neurotransmitter and hormone exocytosis. Like other primary alcohols, 1-butanol is a substrate for PLD and thereby disrupts formation of the intracellular signaling lipid phosphatidic acid. Because secondary and tertiary butanols do not undergo this transphosphatidylation, they have been used as controls for 1-butanol to implicate PLD signaling. Recently, selective pharmacological inhibitors of PLD have been developed and, in some cases, fail to block cellular functions previously ascribed to PLD using primary alcohols. For example, exocytosis of insulin and degranulation of mast cells are blocked by primary alcohols, but not by the PLD inhibitor FIPI. In this study we show that 1-butanol reduces catecholamine secretion from adrenal chromaffin cells to a much greater extent than tert-butanol, and that the PLD inhibitor VU0155056 has no effect. Using fluorescent imaging we show the effect of these drugs on depolarization-evoked calcium entry parallel those on secretion. Patch-clamp electrophysiology confirmed the peak amplitude of voltage-gated calcium channel currents (I(Ca)) is inhibited by 1-butanol, with little or no block by secondary or tert-butanol. Detailed comparison shows for the first time that the different butanol isomers exert distinct, and sometimes opposing, effects on the voltage-dependence and gating kinetics of I(Ca). We discuss these data with regard to PLD signaling in cellular physiology and the molecular targets of general anesthetics.

The dorsal root ganglia (DRG) house the primary afferent neurons responsible for somatosensation, including pain. We previously identified Jedi-1 (PEAR1/MEGF12) as a phagocytic receptor expressed by satellite glia in the DRG involved in clearing apoptotic neurons during development. Here, we further investigated the function of this receptor in vivo using Jedi-1 null mice. In addition to satellite glia, we found Jedi-1 expression in perineurial glia and endothelial cells, but not in sensory neurons. We did not detect any morphological or functional changes in the glial cells or vasculature of Jedi-1 knockout mice. Surprisingly, we did observe changes in DRG neuron activity. In neurons from Jedi-1 knockout (KO) mice, there was an increase in the fraction of capsaicin-sensitive cells relative to wild type (WT) controls. Patch-clamp electrophysiology revealed an increase in excitability, with a shift from phasic to tonic action potential firing patterns in KO neurons. We also found alterations in the properties of voltage-gated sodium channel currents in Jedi-1 null neurons. These results provide new insight into the expression pattern of Jedi-1 in the peripheral nervous system and indicate that loss of Jedi-1 alters DRG neuron activity indirectly through an intercellular interaction between non-neuronal cells and sensory neurons.

In this collection, senior experts explore all aspects of extreme right wing political violence, from the nature of the threat, processes of engagement, and ideology to the lessons that can be drawn from exiting such engagement. Further, right wing activism and political violence are compared with Jihadi violence and engagement. Also, the European experience is placed within a greater framework, including that of the United States and the Arab Spring. The book opens with an essay on U.S. far right groups, investigating their origins and processes of recruitment. It then delves into violence against UK Mosques and Islamic centers, the relationship between Ulster loyalism and far right extremism, the Dutch extremist landscape, and the July 2011 Norway attacks. Also discussed are how narratives of violence are built and justified, at what point do individuals join into violence, and how differently states respond to left-wing vs. right-wing extremism. This comparative work offers a unique look into the very nature of right wing extremism and will be a must-read for anyone studying political violence and terrorism

Problem Atrial fibrillation is the most common persistent arrhythmia in adults and carries an increased risk of thromboembolism and stroke. Electrical (DC) cardioversion is an effective treatment, but logistical difficulties in many institutions lead to problems providing a prompt service. This reduces the rate of long term success, delays relief of symptoms, and increases the burden on anticoagulation clinics. Design Prospective audit of introduction of a collaborative, nurse led DC cardioversion service in a day surgery unit. Setting Day surgery unit 5 km from an acute hospital in southeast London. Key measures for improvement Waiting times, success of procedures, and complication rates. Strategies for change Collaborative working across traditional specialty boundaries; empowerment of patients within the process; using a nurse consultant as a single point of reference to coordinate the service. Effects of change Sinus rhythm was restored in 131 (92%) of the first 143 patients treated. Three patients needed hospital admission; all were discharged uneventfully within 24 hours. No important complications occurred. Waiting times were reduced from 27 weeks to eight weeks for patients eligible for the service. Lessons learnt Elective DC cardioversion under general anaesthesia can be safely done by an appropriately trained nurse in a day surgery unit remote from an acute general hospital. This model of care is effective and can reduce waiting times and relieve pressure on acute beds and junior doctors.

N- and P/Q-type Ca2+channels regulate a number of critical physiological processes including synaptic transmission and hormone secretion. These Ca2+channels are multisubunit proteins, consisting of a pore-forming α1and accessory β and α2δ subunits each encoded by multiple genes and splice variants. β subunits alter current amplitude and kinetics. The β2asubunit is associated with slowed inactivation, an effect that requires the palmitoylation of two N-terminal cysteine residues in β2a. In the current manuscript, we studied steady state inactivation properties of native N-and P/Q-type Ca2+channels and recombinant N-type Ca2+channels. When bovine α1Band β2aand human α2δ were coexpressed in tsA 201 cells, we observed significant variations in inactivation; some cells exhibited virtually no inactivation as the holding potential was altered whereas others exhibited significant inactivation. A similar variability in inactivation was observed in native channels from bovine chromaffin cells. In individual chromaffin cells, the amount of inactivation exhibited by N-type channels was correlated with the inactivation of P/Q-type channels, suggesting a shared mechanism. Our results with recombinant channels with known β subunit composition indicated that inactivation could be dynamically regulated, possibly by alterations in β subunit palmitoylation. Tunicamycin, which inhibits palmitoylation, increased steady-state inactivation of Ca2+channels in chromaffin cells. Cerulenin, another drug that inhibits palmitoylation, also increased inactivation. Tunicamycin produced a similar effect on recombinant N-type Ca2+channels containing β2abut not β2bor β2asubunits mutated to be palmitoylation deficient. Our results suggest that Ca2+channels containing β2asubunits may be regulated by dynamic palmitoylation.

Calpains Play a Role in Insulin Secretion and Action Seamus K. Sreenan 1 , Yun-Ping Zhou 1 , Kenichi Otani 1 8 , Polly A. Hansen 6 , Kevin P.M. Currie 2 , Chien-Yuan Pan 2 , Jean-Pyo Lee 1 , Diane M. Ostrega 1 , William Pugh 1 , Yukio Horikawa 3 5 , Nancy J. Cox 1 4 , Craig L. Hanis 7 , Charles F. Burant 6 , Aaron P. Fox 2 , Graeme I. Bell 1 3 4 5 and Kenneth S. Polonsky 8 1 Medicine 2 Neurobiology, Pharmacology and Physiology 3 Biochemistry and Molecular Biology, and 4 Human Genetics, and the 5 Howard Hughes Medical Institute, the University of Chicago, Chicago, Illinois 6 Department of Cell Biology, Parke-Davis Pharmaceutical Research Division/Warner-Lambert Company, Ann Arbor, Michigan 7 Human Genetics Center, the University of Texas Health Science Center at Houston, Houston, Texas 8 Department of Medicine, Washington University, St. Louis, Missouri Abstract Studies of the genetic basis of type 2 diabetes suggest that variation in the calpain-10 gene affects susceptibility to this common disorder, raising the possibility that calpain-sensitive pathways may play a role in regulating insulin secretion and/or action. Calpains are ubiquitously expressed cysteine proteases that are thought to regulate a variety of normal cellular functions. Here, we report that short-term (4-h) exposure to the cell-permeable calpain inhibitors calpain inhibitor II and E-64-d increases the insulin secretory response to glucose in mouse pancreatic islets. This dose-dependent effect is observed at glucose concentrations above 8 mmol/l. This effect was also seen with other calpain inhibitors with different mechanisms of action but not with cathepsin inhibitors or other protease inhibitors. Enhancement of insulin secretion with short-term exposure to calpain inhibitors is not mediated by increased responses in intracellular Ca 2+ or increased glucose metabolism in islets but by accelerated exocytosis of insulin granules. In muscle strips and adipocytes, exposure to both calpain inhibitor II and E-64-d reduced insulin-mediated glucose transport. Incorporation of glucose into glycogen in muscle also was reduced. These results are consistent with a role for calpains in the regulation of insulin secretion and insulin action. Footnotes Address correspondence and reprint requests to Kenneth S. Polonsky, Department of Medicine, Washington University, 660 S. Euclid, Campus Box 8066, St. Louis, MO 63110. E-mail: polonsky{at}im.wustl.edu . Received for publication 21 June 2000 and accepted in revised form 24 May 2001. S.K.S. is currently affiliated with the Department of Endocrinology, Royal College of Surgeons, James Connolly Memorial Hospital, Blanchardstown, Dublin, Ireland. Y.H. is currently affiliated with the Laboratory of Molecular Genetics, Institute for Molecular and Cellular Regulation, Gunma University, Gunma, Japan. ANOVA, analysis of varience; AUC, area under the curve; 2-DOG, 2-deoxyglucose; KHB, Krebs-Henseleit buffer; KRB, Krebs-Ringer’s buffer; 3-MG, 3- O -[ 3 H]methyl- d -glucose; TEA, tetraethylammonium.

CaV2.2 (N-type) voltage-gated calcium channels (Ca2+ channels) play key roles in neurons and neuroendocrine cells including the control of cellular excitability, neurotransmitter / hormone secretion, and gene expression. Calcium entry is precisely controlled by channel gating properties including multiple forms of inactivation. \"Fast\" voltage-dependent inactivation is relatively well-characterized and occurs over the tens-to- hundreds of milliseconds timeframe. Superimposed on this is the molecularly distinct, but poorly understood process of \"slow\" voltage-dependent inactivation, which develops / recovers over seconds-to-minutes. Protein kinases can modulate \"slow\" inactivation of sodium channels, but little is known about if/how second messengers control \"slow\" inactivation of Ca2+ channels. We investigated this using recombinant CaV2.2 channels expressed in HEK293 cells and native CaV2 channels endogenously expressed in adrenal chromaffin cells. The PKC activator phorbol 12-myristate 13-acetate (PMA) dramatically prolonged recovery from \"slow\" inactivation, but an inactive control (4 alpha -PMA) had no effect. This effect of PMA was prevented by calphostin C, which targets the C1-domain on PKC, but only partially reduced by inhibitors that target the catalytic domain of PKC. The subtype of the channel beta -subunit altered the kinetics of inactivation but not the magnitude of slowing produced by PMA. Intracellular GDP- beta -S reduced the effect of PMA suggesting a role for G proteins in modulating \"slow\" inactivation. We postulate that the kinetics of recovery from \"slow\" inactivation could provide a molecular memory of recent cellular activity and help control CaV2 channel availability, electrical excitability, and neurotransmission in the seconds-to-minutes timeframe.