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Tora Mitra-Ganguli, Iuliia Vitko, Edward Perez-Reyes, Ann R Rittenhouse
The G(q)-coupled tachykinin receptor (neurokinin-1 receptor [NK-1R]) modulates N-type Ca(2+) channel (Ca(V)2.2 or N channel) activity at two distinct sites by a pathway involving a lipid metabolite, most likely arachidonic acid (AA). In another study published in this issue (Heneghan et al. 2009. J. Gen Physiol. doi:10.1085/jgp.200910203), we found that the form of modulation observed depends on which Ca(V)beta is coexpressed with Ca(V)2.2. When palmitoylated Ca(V)beta2a is coexpressed, activation of NK-1Rs by substance P (SP) enhances N current...
November 2009: Journal of General Physiology
John F Heneghan, Tora Mitra-Ganguli, Lee F Stanish, Liwang Liu, Rubing Zhao, Ann R Rittenhouse
In superior cervical ganglion (SCG) neurons, stimulation of M(1) receptors (M(1)Rs) produces a distinct pattern of modulation of N-type calcium (N-) channel activity, enhancing currents elicited with negative test potentials and inhibiting currents elicited with positive test potentials. Exogenously applied arachidonic acid (AA) reproduces this profile of modulation, suggesting AA functions as a downstream messenger of M(1)Rs. In addition, techniques that diminish AA's concentration during M(1)R stimulation minimize N-current modulation...
November 2009: Journal of General Physiology
Marino DiFranco, Marbella Quinonez, Joana Capote, Julio Vergara
A growing interest in cell biology is to express transgenically modified forms of essential proteins (e.g. fluorescently tagged constructs and/or mutant variants) in order to investigate their endogenous distribution and functional relevance. An interesting approach that has been implemented to fulfill this objective in fully differentiated cells is the in vivo transfection of plasmids by various methods into specific tissues such as liver, skeletal muscle, and even the brain. We present here a detailed description of the steps that must be followed in order to efficiently transfect genetic material into fibers of the flexor digitorum brevis (FDB) and interosseus (IO) muscles of adult mice using an in vivo electroporation approach...
October 19, 2009: Journal of Visualized Experiments: JoVE
Viktor Yarotskyy, Keith S Elmslie
omega-conotoxin GVIA (omegaCTX) is a specific blocker of N-type calcium (CaV2.2) channels that inhibits neuropathic pain. While the toxin appears to be an open channel blocker, we show that N-channel gating charge movement is modulated. Gating currents were recorded from N-channels expressed along with beta2a and alpha2delta subunits in HEK293 cells in external solutions containing either lanthanum and magnesium (La-Mg) or 5 mM Ca2+ plus omegaCTX (omegaCTX-Ca). A comparison showed that omegaCTX induced a 10-mV right shift in the gating charge versus voltage (Q-V) relationship, smaller off-gating current time constant (tau Q(Off)), a lower tau Q(Off) voltage dependence, and smaller on-gating current (Q(On)) tau...
January 2009: Journal of Neurophysiology
Iuliia Vitko, Aleksandr Shcheglovitov, Joel P Baumgart, Imilla I Arias-Olguín, Janet Murbartián, Juan Manuel Arias, Edward Perez-Reyes
BACKGROUND: The Ca(v)beta subunits of high voltage-activated Ca(2+) channels control the trafficking and biophysical properties of the alpha(1) subunit. The Ca(v)beta-alpha(1) interaction site has been mapped by crystallographic studies. Nevertheless, how this interaction leads to channel regulation has not been determined. One hypothesis is that betas regulate channel gating by modulating movements of IS6. A key requirement for this direct-coupling model is that the linker connecting IS6 to the alpha-interaction domain (AID) be a rigid structure...
2008: PloS One
Naser Jaleel, Hiroyuki Nakayama, Xiongwen Chen, Hajime Kubo, Scott MacDonnell, Hongyu Zhang, Remus Berretta, Jeffrey Robbins, Leanne Cribbs, Jeffery D Molkentin, Steven R Houser
T-type Ca(2+) channels (TTCCs) are expressed in the developing heart, are not present in the adult ventricle, and are reexpressed in cardiac diseases involving cardiac dysfunction and premature, arrhythmogenic death. The goal of this study was to determine the functional role of increased Ca(2+) influx through reexpressed TTCCs in the adult heart. A mouse line with cardiac-specific, conditional expression of the alpha1G-TTCC was used to increase Ca(2+) influx through TTCCs. alpha1G hearts had mild increases in contractility but no cardiac histopathology or premature death...
November 7, 2008: Circulation Research
Andriy V Dresviannikov, Karen M Page, Jerôme Leroy, Wendy S Pratt, Annette C Dolphin
CaVbeta subunits of voltage-gated calcium channels contain two conserved domains, a src-homology-3 (SH3) domain and a guanylate kinase-like (GK) domain with an intervening HOOK domain. We have shown in a previous study that, although Gbetagamma-mediated inhibitory modulation of CaV2.2 channels did not require the interaction of a CaVbeta subunit with the CaValpha1 subunit, when such interaction was prevented by a mutation in the alpha1 subunit, G protein modulation could not be removed by a large depolarization and showed voltage-independent properties (Leroy et al...
February 2009: Pflügers Archiv: European Journal of Physiology
Chad E Grueter, Sunday A Abiria, Yunji Wu, Mark E Anderson, Roger J Colbran
Ca2+/calmodulin-dependent protein kinase II (CaMKII) phosphorylates the beta2a subunit of voltage-gated Ca2+ channels at Thr498 to facilitate cardiac L-type Ca2+ channels. CaMKII colocalizes with beta2a in cardiomyocytes and also binds to a domain in beta2a that contains Thr498 and exhibits an amino acid sequence similarity to the CaMKII autoinhibitory domain and to a CaMKII binding domain in the NMDA receptor NR2B subunit (Grueter, C. E. et al. (2006) Mol. Cell 23, 641). Here, we explore the selectivity of the actions of CaMKII among Ca2+ channel beta subunit isoforms...
February 12, 2008: Biochemistry
Roman Laszlo, Christoph Winkler, Stefan Wöhrl, Ralf E Wessel, Sara Laszlo, Mathias C Busch, Jürgen Schreieck, Ralph F Bosch
We investigated the effects of a 7-day verapamil pretreatment (VPT, 7.5 mg/kg bodyweight subcutaneously every 12 h) on ionic currents and molecular mechanisms underlying tachycardia-induced early electrical remodeling after 24-h rapid atrial pacing (RAP, 600 bpm) in rabbit atrium. Animals were divided into four groups (n = 6 each group): control (not paced, no verapamil), paced only, verapamil only and verapamil and paced, respectively. VPT doubled ICa,L [7.0 +/- 0.7 pA/pF (control) vs 14.2 +/- 0.6 pA/pF (verapamil only)]...
December 2007: Naunyn-Schmiedeberg's Archives of Pharmacology
Allen W Chan, Steven Owens, Connie Tung, Elise F Stanley
Presynaptic CaV2.2 (N type) calcium channels gate the influx of calcium ions to trigger transmitter release. We have previously demonstrated at the chick ciliary ganglion presynaptic calyx terminal that the bulk of these channels are highly resistant to voltage dependent inactivation [E.F. Stanley, G. Goping, Characterization of a calcium current in a vertebrate cholinergic presynaptic nerve terminal, J. Neurosci. 11 (1991) 985-993; E.F. Stanley, Syntaxin I modulation of presynaptic calcium channel inactivation revealed by botulinum toxin C1, Eur...
October 2007: Cell Calcium
Liansheng Sun, Guangfen Mao, Satya P Kunapuli, Danny N Dhanasekaran, A Koneti Rao
Phospholipase C (PLC) beta2 plays a pivotal role in G-protein dependent signal transduction in platelets. We have previously demonstrated in platelets, leukocytes and human erythroleukemia cells the presence of transcripts of two forms of PLC-beta2 generated by alternative splicing. They differ by 45 nucleotides in the carboxyl-terminal region and are designated as PLC-beta2a and PLC-beta2b, with and without by 15 amino acid residues (corresponding to 864-878). The presence of the two variants has not been shown at the protein level in cells...
May 2007: Platelets
Stefan Herzig, Ismail F Y Khan, Dirk Gründemann, Jan Matthes, Andreas Ludwig, Guido Michels, Uta C Hoppe, Dipayan Chaudhuri, Arnold Schwartz, David T Yue, Roger Hullin
L-type calcium channels are composed of a pore, alpha1c (Ca(V)1.2), and accessory beta- and alpha2delta-subunits. The beta-subunit core structure was recently resolved at high resolution, providing important information on many functional aspects of channel modulation. In this study we reveal differential novel effects of five beta2-subunits isoforms expressed in human heart (beta(2a-e)) on the single L-type calcium channel current. These splice variants differ only by amino-terminal length and amino acid composition...
May 2007: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
Xifu Liu, Xinhua Gu, Zhaoming Li, Xinyan Li, Hui Li, Jianjie Chang, Ping Chen, Jing Jin, Bing Xi, Denghong Chen, Donna Lai, Robert M Graham, Mingdong Zhou
OBJECTIVES: We evaluated the therapeutic potential of a recombinant 61-residue neuregulin-1 (beta2a isoform) receptor-active peptide (rhNRG-1) in multiple animal models of heart disease. BACKGROUND: Activation of the erbB family of receptor tyrosine kinases by rhNRG-1 could provide a treatment option for heart failure, because neuregulin-stimulated erbB2/erbB4 heterodimerization is not only critical for myocardium formation in early heart development but prevents severe dysfunction of the adult heart and premature death...
October 3, 2006: Journal of the American College of Cardiology
Tae-Seong Lee, Rosi Karl, Sven Moosmang, Peter Lenhardt, Norbert Klugbauer, Franz Hofmann, Thomas Kleppisch, Andrea Welling
Calcium-dependent facilitation of L-type calcium channels has been reported to depend on the function of calmodulin kinase II. In contrast, the mechanism for voltage-dependent facilitation is not clear. In HEK 293 cells expressing Ca(v)1.2, Ca(v)beta2a, and calmodulin kinase II, the calcium current measured at +30 mV was facilitated up to 1.5-fold by a 200-ms-long prepulse to +160 mV. This voltage-dependent facilitation was prevented by the calmodulin kinase II inhibitors KN93 and the autocamtide-2-related peptide...
September 1, 2006: Journal of Biological Chemistry
Brian S Finlin, Robert N Correll, Chunyan Pang, Shawn M Crump, Jonathan Satin, Douglas A Andres
Voltage-gated calcium channels are multiprotein complexes that regulate calcium influx and are important contributors to cardiac excitability and contractility. The auxiliary beta-subunit (CaV beta) binds a conserved domain (the alpha-interaction domain (AID)) of the pore-forming CaV alpha1 subunit to modulate channel gating properties and promote cell surface trafficking. Recently, members of the RGK family of small GTPases (Rem, Rem2, Rad, Gem/Kir) have been identified as novel contributors to the regulation of L-type calcium channel activity...
August 18, 2006: Journal of Biological Chemistry
Shawn M Crump, Robert N Correll, Elizabeth A Schroder, William C Lester, Brian S Finlin, Douglas A Andres, Jonathan Satin
Cardiac voltage-gated L-type Ca channels (Ca(V)) are multiprotein complexes, including accessory subunits such as Ca(V)beta2 that increase current expression. Recently, members of the Rad and Gem/Kir-related family of small GTPases have been shown to decrease current, although the mechanism remains poorly defined. In this study, we evaluated the contribution of the L-type Ca channel alpha-subunit (Ca(V)1.2) to Ca(V)beta2-Rem inhibition of Ca channel current. Specifically, we addressed whether protein kinase A (PKA) modulation of the Ca channel modifies Ca(V)beta2-Rem inhibition of Ca channel current...
October 2006: American Journal of Physiology. Heart and Circulatory Physiology
N A Ramakrishnan, M J Drescher, S A Sheikhali, K M Khan, J S Hatfield, M J Dickson, D G Drescher
We report new molecular evidence for the presence of an N-type (Ca(v)2.2, alpha1B) voltage-gated Ca(2+) channel in hair cells of the saccular epithelium of the rainbow trout. The Ca(v)2.2 amino-acid sequence shows 68% and 63% identity compared with chick and human Ca(v)2.2, respectively. This channel reveals features that are characteristic of an N-type Ca(2+) channel: an omega-conotoxin GVIA binding domain, G(betagamma) binding regions, and a synaptic protein interaction site. Immunohistochemical studies with a custom antibody show that immunoreactivity for the Ca(v)2...
2006: Neuroscience
A Livneh, R Cohen, D Atlas
The inactivation of voltage-gated L-type Ca(2+) channels (Ca(V)1) regulates Ca(2+) entry and controls intracellular Ca(2+) levels that are essential for cellular activity. The molecular entities implicated in L-channel (Ca(V)1.2) inactivation are not fully identified. Here we show for the first time the functional impact of one of the two highly conserved clusters of six negatively charged glutamates and aspartate (802-807; poly ED motif) at the II-III loop of the alpha 1 subunits of rabbit of Ca(v)1.2, alpha(1)1...
2006: Neuroscience
Yu-Jin Won, Kum Whang, In Deok Kong, Kyu-Sang Park, Joong-Woo Lee, Seong-Woo Jeong
Among the autonomic ganglia, major pelvic ganglia (MPG) innervating the urogenital system are unique because both sympathetic and parasympathetic neurons are colocalized within one ganglion capsule. Sympathetic MPG neurons are discriminated from parasympathetic ones by expression of low voltage-activated Ca2+ channels that primarily arise from T-type alpha1H isoform and contribute to the generation of low-threshold spikes. Until now, however, expression profiles of high voltage-activated (HVA) Ca2+ channels in these two populations of MPG neurons remain unknown...
June 2006: Journal of Pharmacology and Experimental Therapeutics
Kevin J Gingrich, Son Tran, Igor M Nikonorov, Thomas J Blanck
BACKGROUND: Volatile anesthetics depress cardiac contractility, which involves inhibition of cardiac L-type calcium channels. To explore the role of voltage-dependent inactivation, the authors analyzed halothane effects on recombinant cardiac L-type calcium channels (alpha1Cbeta2a and alpha1Cbeta2aalpha2/delta1), which differ by the alpha2/delta1 subunit and consequently voltage-dependent inactivation. METHODS: HEK-293 cells were transiently cotransfected with complementary DNAs encoding alpha1C tagged with green fluorescent protein and beta2a, with and without alpha2/delta1...
December 2005: Anesthesiology
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