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Activation Rate (activation + rate)
Selected AbstractsRegulatory Mechanisms and Physiological Relevance of a Voltage-Gated H+ Channel in Murine Osteoclasts: Phorbol Myristate Acetate Induces Cell Acidosis and the Channel Activation,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 11 2003Hiroyuki Mori Abstract The voltage-gated H+ channel is a powerful H+ extruding mechanism of osteoclasts, but its functional roles and regulatory mechanisms remain unclear. Electrophysiological recordings revealed that the H+ channel operated on activation of protein kinase C together with cell acidosis. Introduction: H+ is a key signaling ion in bone resorption. In addition to H+ pumps and exchangers, osteoclasts are equipped with H+ conductive pathways to compensate rapidly for pH imbalance. The H+ channel is distinct in its strong H+ extrusion ability and voltage-dependent gatings. Methods: To investigate how and when the H+ channel is available in functional osteoclasts, the effects of phorbol 12-myristate 13-acetate (PMA), an activator for protein kinase C, on the H+ channel were examined in murine osteoclasts generated in the presence of soluble RANKL (sRANKL) and macrophage-colony stimulating factor (M-CSF). Results and Conclusions: Whole cell recordings clearly showed that the H+ current was enhanced by increasing the pH gradient across the plasma membrane (,pH), indicating that the H+ channel changed its activity by sensing ,pH. The reversal potential (Vrev) was a valuable tool for the real-time monitoring of ,pH in clamped cells. In the permeabilized patch, PMA (10 nM-1.6 ,M) increased the current density and the activation rate, slowed decay of tail currents, and shifted the threshold toward more negative voltages. In addition, PMA caused a negative shift of Vrev, suggesting that intracellular acidification occurred. The PMA-induced cell acidosis was confirmed using a fluorescent pH indicator (BCECF), which recovered quickly in a K+ -rich alkaline solution, probably through the activated H+ channel. Both cell acidosis and activation of the H+ channel by PMA were inhibited by staurosporine. In ,80% of cells, the PMA-induced augmentation in the current activity remained after compensating for the ,pH changes, implying that both ,pH-dependent and -independent mechanisms mediated the channel activation. Activation of the H+ channel shifted the membrane potential toward Vrev. These data suggest that the H+ channel may contribute to regulation of the pH environments and the membrane potential in osteoclasts activated by protein kinase C. [source] The Mechanisms of Atrial FibrillationJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 2006PENG-SHENG CHEN M.D. In this article we have reviewed the mechanisms of atrial fibrillation (AF) with special emphasis on the thoracic veins. Based on a number of features, the thoracic veins are highly arrhythmogenic. The pulmonary vein (PV)-left atrial (LA) junction has discontinuous myocardial fibers separated by fibrotic tissues. The PV muscle sleeve is highly anisotropic. The vein of Marshall (VOM) in humans has multiple small muscle bundles separated by fibrosis and fat. Insulated muscle fibers can promote reentrant excitation, automaticity, and triggered activity. The PV muscle sleeves contain periodic acid-Schiff (PAS)-positive large pale cells that are morphologically reminiscent of Purkinje cells. These special cells could be the sources of focal discharge. Antiarrhythmic drugs have significant effects on PV muscle sleeves both at baseline and during AF. Both class I and III drugs have effects on wavefront traveling from PV to LA and from LA to PV. Separating the thoracic veins and the LA with ablation techniques also prevents PV-LA interaction. By reducing PV-LA interaction, pharmacological therapy and PV isolation reduce the activation rate in PV, intracellular calcium accumulation, and triggered activity. Therefore, thoracic vein isolation is an important technique in AF control. We conclude that thoracic veins are important in the generation and maintenance of AF. [source] Reentry Site During Fibrillation Induction in Relation to Defibrillation Efficacy for Different Shock WaveformsJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 5 2001Ph.D., RAYMOND E. IDEKER M.D. Reentry Site and Defibrillation Waveform Efficacy.Introduction: Unsuccessful defibrillation shocks may reinitiate fibrillation by causing postshock reentry. Methods and Results: To better understand why some waveforms are more efficacious for defibrillation, reentry was induced in six dogs with 1-, 2-, 4-, 8-, and 16-msec monophasic and 1/1- (both phases 1 msec) 2/2-, 4/4-, and 8/8-msec biphasic shocks. Reentry was initiated by 141 ± 15 V shocks delivered from a defibrillator with a 150- , F capacitance during the vulnerable period of paced rhythm (183 ± 12 msec after the last pacing stimulus). The shock potential gradient field was orthogonal to the dispersion of refractoriness. Activation was mapped with 121 electrodes covering 4 × 4 cm of the right ventricular epicardium, and potential gradient and degree of recovery of excitability were estimated at the sites of reentry. Defibrillation thresholds (DFTs) were estimated by an up-down protocol for the same nine waveforms in eight dogs internally and in nine other dogs externally. DFT voltages for the different waveforms were positively correlated with the magnitude of shock potential gradient and negatively correlated with the recovery interval at the site at which reentry was induced by the waveform during paced rhythm for both internal (DFT = 1719 + 64.5 , V , 11.1RI; R2= 0.93) and external defibrillation (DFT = 3445 + 150 , V , 22RI; R2= 0.93). Conclusion: The defibrillation waveforms with the lowest DFTs were those that induced reentry at sites of low shock potential gradient, indicating efficacious stimulation of myocardium. Additionally, the site of reentry induced by waveforms with the lowest DFTs was in myocardium that was more highly recovered just before the shock, perhaps because this high degree of recovery seldom occurs during defibrillation due to the rapid activation rate during fibrillation. [source] Chain Length Distributions of Polyolefins Made in Stopped-Flow Reactors for Non-Instantaneous Site ActivationMACROMOLECULAR REACTION ENGINEERING, Issue 2 2008Joćo B. P. Soares Abstract We developed an analytical solution to describe how the chain length distribution (CLD) of polymers made with coordination polymerization catalysts vary as a function of time for very short polymerizations considering non-instantaneous site activation. This solution is an extension of our previous analytical expression for instantaneous site activation. We validated the analytical solution with dynamic Monte Carlo simulation and obtained excellent agreement. Simulation results indicate that, unless the catalyst activation rate is much lower than the propagation rate, it will have only a minor effect on the initial shape of the CLD of polymers made in stopped-flow reactors (SFR). We also show how incorrect polymerization kinetic parameters may be estimated when assuming instantaneous site activation when this hypothesis is not applicable to the polymerization data under investigation. [source] Effect of maturation stage at cryopreservation on post-thaw cytoskeleton quality and fertilizability of equine oocytesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 5 2006T. Tharasanit Abstract Oocyte cryopreservation is a potentially valuable technique for salvaging the germ-line when a valuable mare dies, but facilities for in vitro embryo production or oocyte transfer are not immediately available. This study examined the influence of maturation stage and freezing technique on the cryopreservability of equine oocytes. Cumulus oocyte complexes were frozen at the immature stage (GV) or after maturation in vitro for 30 hr (MII), using either conventional slow freezing (CF) or open pulled straw vitrification (OPS); cryoprotectant-exposed and untreated nonfrozen oocytes served as controls. After thawing, GV oocytes were matured in vitro, and MII oocytes were incubated for 0 or 6 hr, before staining to examine meiotic spindle quality by confocal microscopy. To assess fertilizability, CF MII oocytes were subjected to intracytoplasmic sperm injection (ICSI) and cultured in vitro. At 12, 24, and 48 hr after ICSI, injected oocytes were fixed to examine their progression through fertilization. Both maturation stage and freezing technique affected oocyte survival. The meiosis resumption rate was higher for OPS than CF for GV oocytes (28% vs. 1.2%; P,<,0.05), but still much lower than for controls (66%). Cryopreserving oocytes at either stage induced meiotic spindle disruption (37%,67% normal spindles vs. 99% in controls; P,<,0.05). Among frozen oocytes, however, spindle quality was best for oocytes frozen by CF at the MII stage and incubated for 6 hr post-thaw (67% normal); since this combination of cryopreservation/IVM yielded the highest proportion of oocytes reaching MII with a normal spindle (35% compared to <20% for other groups), it was used when examining the effects of cryopreservation on fertilizability. In this respect, the rate of normal fertilization for CF MII oocytes after ICSI was much lower than for controls (total oocyte activation rate, 26% vs. 56%; cleavage rate at 48 hr, 8% vs. 42%: P,<,0.05). Thus, although IVM followed by CF yields a respectable percentage of normal-looking MII oocytes (35%), their ability to support fertilization is severely compromised. Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. [source] Alpha-retinals as Rhodopsin Chromophores,Preference for the 9- Z Configuration and Partial Agonist Activity,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2008Yajie Wang The visual pigment rhodopsin, the photosensory element of the rod photoreceptor cell in the vertebrate retina, shows in combination with an endogenous ligand, 11- Z retinal, an astonishing photochemical performance. It exhibits an unprecedented quantum yield (0.67) in a highly defined and ultrafast photoisomerization process. This triggers the conformational changes leading to the active state Meta(rhodopsin) II. Retinal is covalently bound to Lys-296 of the protein opsin in a protonated Schiff base. The resulting positive charge delocalization over the terminal part of the polyene chain of retinal creates a conjugation defect that upon photoexcitation moves to the opposite end of the polyene. Shortening the polyene as in 4,5-dehydro,5,6-dihydro (alpha), 5,6-dihydro or 7,8-dihydro-analogs might facilitate photoisomerization of a 9- Z and a 11- Z bond. Here we describe pigment analogs generated with bovine opsin and 11- Z or 9- Z 4,5-dehydro,5,6-dihydro-retinal that were further characterized by UV,Vis and FTIR spectroscopy. The preference of opsin for native 11- Z retinal over the 9- Z isomer is reversed in 4,5-dehydro,5,6-dihydro-retinal. 9- Z 4,5-dehydro,5,6-dihydro-retinal readily generated a photosensitive pigment. This modification has no effect on the quantum yield, but affects the Batho,blueshifted intermediate (BSI) equilibrium and leads to a strong decrease in the G-protein activation rate because of a downshift of the pKa of the Meta I,Meta II equilibrium. [source] The ,1 and ,6 subunit subtypes of the mammalian GABAA receptor confer distinct channel gating kineticsTHE JOURNAL OF PHYSIOLOGY, Issue 2 2004Janet L. Fisher The GABAA receptors show a large degree of structural heterogeneity, with seven different subunit families, and 16 different subtypes in mammalian species. The , family is the largest, with six different subtypes. The ,1 and ,6 subtypes are among the most diverse within this family and confer distinct pharmacological properties to recombinant and neuronal receptors. To determine whether different single channel and macroscopic kinetic properties were also associated with these subtypes, the ,1 or ,6 subunit was expressed in mammalian cells along with ,3 and ,2L subunits and the kinetic properties examined with outside-out patch recordings. The ,1,3,2L receptors responded to GABA with long-duration openings organized into multi-opening bursts. In contrast, channel openings of the ,6,3,2L receptors were predominately short in duration and occurred as isolated, single openings. The subunit subtype also affected the deactivation rate of the receptor, which was almost 2-fold slower for ,6,3,2L, compared with the ,1,3,2L isoform. Onset of fast desensitization did not differ between the isoforms. To determine the structural domains responsible for these differences in kinetic properties, we constructed six chimeric subunits, combining different regions of the ,1 and ,6 subunits. The properties of the chimeric subunits indicated that structures within the third transmembrane domain (TM3) and the TM3,TM4 intracellular loop conferred differences in single channel gating kinetics that subsequently affected the deactivation rate and GABA EC50. The effect of agonist concentration on the rise time of the current showed that the extracellular N-terminal domain was largely responsible for binding characteristics, while the transmembrane domains determined the activation rate at saturating GABA concentrations. This suggests that subunit structures outside of the agonist binding and pore-lining domains are responsible for the kinetic differences conferred by the ,1 and ,6 subtypes. Structural heterogeneity within these transmembrane and intracellular regions can therefore influence the characteristics of the postsynaptic response of GABAA receptors with different subunit composition. [source] A ,bottom-up' approach for endo-PK/PD analysisBIOPHARMACEUTICS AND DRUG DISPOSITION, Issue 7 2006S. Neelakantan Abstract A ,bottom-up' PK/PD analysis approach employing system analysis principles of convolution/deconvolution and special nonparametric estimation procedures is presented to resolve the complex ,endo-PK/PD' of the endogenous form of recombinant drugs using erythropoietin (EPO) as an example. A novel cellular deconvolution algorithm is presented that facilitates the identification of the functional relationship between the variables involved in EPO's complex PK/PD. Five sheep each underwent two phlebotomies spaced 4,6 weeks apart when their hemoglobin levels were reduced from 12 g/dl to 3,4 g/dl. EPO levels and reticulocyte counts were frequently sampled. The data were analysed using end-constrained cubic splines. The rate of reticulocyte production was determined using the novel deconvolution methodology. The erythroid progenitor cells activation rate by EPO was estimated from the reticulocyte production rate using a lag-time parameter which determines the delay in the reticulocyte appearance in the blood relative to the activation of erythroid progenitors. Hysteresis minimization combined with cellular deconvolution was employed to determine the population PK/PD transduction function relating the progenitor activation rate to EPO concentrations in a nonparametric manner without assuming a specific structure. The proposed approach provides a rational informative starting point for developing parametric PK/PD models to resolve the complex endo-PK/PD of recombinant drugs. Copyright © 2006 John Wiley & Sons, Ltd. [source] Functional assessment of centrosomes of spermatozoa and spermatids microinjected into rabbit oocytes,MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2009Masahito Tachibana Abstract Although intracytoplasmic sperm injection (ICSI) is a widely used assisted reproductive technique, the fertilization rates and pregnancy rates of immature spermatids especially in round spermatid injection (ROSI) remain very low. During mammalian fertilization, the sperm typically introduces its own centrosome which then acts as a microtubule organizing center (MTOC) and is essential for the male and female genome union. In order to evaluate the function of immature germ cell centrosomes, we used the rabbit gamete model because rabbit fertilization follows paternal pattern of centrosome inheritance. First, rabbit spermatids and spermatozoa were injected into oocytes using a piezo-micromanipulator. Next, the centrosomal function to form a sperm aster was determined. Furthermore, two functional centrosome proteins (,-tubulin and centrin) of the rabbit spermatogenic cells were examined. Our results show that the oocyte activation rates by spermatozoa, elongated spermatids, and round spermatids were 86% (30/35), 30% (11/36), and 5% (1/22), respectively. Sperm aster formation rates after spermatozoa, elongated spermatids, and round spermatids injections were 47% (14/30), 27% (3/11), and 0% (0/1), respectively. The aster formation rate of the injected elongating/elongated spermatids was significantly lower than that of the mature spermatozoa (P,=,0.0242). Moreover, sperm asters were not observed in round spermatid injection even after artificial activation. These data suggest that poor centrosomal function, as measured by diminished aster formation rates, is related to the poor fertilization rates when immature spermatogenic cells are injected. Mol. Reprod. Dev. 76: 270,277, 2009. © 2008 Wiley-Liss, Inc. [source] Elevated Histone H1 (MPF) and Mitogen-activated Protein Kinase Activities in Pig Oocytes Following In Vitro Maturation do not Indicate Cytoplasmic MaturationREPRODUCTION IN DOMESTIC ANIMALS, Issue 2 2009MA Setiadi Contents Effects of different media (TCM 199 + BSA, TCM 199 + FCS, TCM 199 + NBCS, Whitten's medium + BSA) supplemented with estradiol-17, and two isolated and everted follicle shells on MPF and MAP kinase activities and the sensitivity to parthenogenetic activation of pig oocytes were examined at the end of culture (48 h). Elevated (P < 0.05) activities of MAP kinase were recorded in metaphase II oocytes following culture in Whitten's medium, whereas MPF levels were lowest (P < 0.05) in MII oocytes matured in TCM 199 supplemented with BSA. Oocytes matured in TCM 199 based media showed higher (P < 0.05) activation rates when compared to oocytes incubated in Whitten's medium. Whitten's medium supplemented with different protein sources (amino acids, FCS, BSA) was used to study the effects of different exposure periods to eCG/hCG stimulation on MPF and MAP kinase activities and in vivo fertilisability following culture for 48 h. MPF and MAP kinase activities were significantly increased by eCG/hCG stimulation of COCs during maturation. Further, the continuous presence of eCG/hCG during culture (48 h) significantly increased the levels of both kinases in comparison to stimulation by gonadotrophins alone during the first 24 h of incubation. In vivo fertilisation of oocytes matured in Whitten's medium supplemented with eCG/hCG for 24 or 48 h led to a significant retardation of early embryonic development compared to ovulated oocytes. In conclusion, media composition and gonadotrophin stimulation affect MPF/MAP kinase activities and the susceptibility to parthenogenetic activation of IVM oocytes. However, elevated kinase levels in pig oocytes following culture do not indicate complete cytoplasmic maturation. [source] | |||||||||||||