共查询到20条相似文献,搜索用时 15 毫秒
1.
《Channels (Austin, Tex.)》2013,7(6):438-442
ZD7288 has been widely used as a tool in the study of hyperpolarization-activated cyclic nucleotide-gated channels (HCN channels), and to test the relationships between HCN channels and heart and brain function. ZD7288 is widely considered a selective blocker of HCN currents. Here we show that ZD7288 inhibits not only HCN channel currents, but also Na+ currents in DRG neurons and ZD7288 was confirmed to inhibit Na+ current in HEK293 cells transfected with Nav1.4 plasmids. Thus our findings challenge the view that ZD7288 is a selective blocker of HCN channels. Conclusions about the role of NCN channels in neuronal function should be re-evaluated if based exclusively on the effect of ZD7288. 相似文献
2.
Xing Wu Liping Liao Xiangming Liu Fang Luo Tianming Yang Chenhong Li 《Channels (Austin, Tex.)》2012,6(6):438-442
ZD7288 has been widely used as a tool in the study of hyperpolarization-activated cyclic nucleotide-gated channels (HCN channels), and to test the relationships between HCN channels and heart and brain function. ZD7288 is widely considered a selective blocker of HCN currents. Here we show that ZD7288 inhibits not only HCN channel currents, but also Na+ currents in DRG neurons and ZD7288 was confirmed to inhibit Na+ current in HEK293 cells transfected with Nav1.4 plasmids. Thus our findings challenge the view that ZD7288 is a selective blocker of HCN channels. Conclusions about the role of NCN channels in neuronal function should be re-evaluated if based exclusively on the effect of ZD7288. 相似文献
3.
Cai X 《Journal of molecular biology》2007,368(5):1284-1291
Depletion of intracellular Ca(2+) stores evokes Ca(2+) entry across the plasma membrane by inducing Ca(2+) release-activated Ca(2+) (CRAC) currents in many cell types. Recently, Orai and STIM proteins were identified as the molecular identities of the CRAC channel subunit and the endoplasmic reticulum Ca(2+) sensor, respectively. Here, extensive database searching and phylogenetic analysis revealed several lineage-specific duplication events in the Orai protein family, which may account for the evolutionary origins of distinct functional properties among mammalian Orai proteins. Based on similarity to key structural domains and essential residues for channel functions in Orai proteins, database searching also identifies a putative primordial Orai sequence in hyperthermophilic archaeons. Furthermore, modern Orai appears to acquire new structural domains as early as Urochodata, before divergence into vertebrates. The evolutionary patterns of structural domains might be related to distinct functional properties of Drosophila and mammalian CRAC currents. Interestingly, Orai proteins display two conserved internal repeats located at transmembrane segments 1 and 3, both of which contain key amino acids essential for channel function. These findings demonstrate biochemical and physiological relevance of Orai proteins in light of different evolutionary origins and will provide novel insights into future structural and functional studies of Orai proteins. 相似文献
4.
Alves GG Lima LM Fávero-Retto MP Lemos AP Peres-Sampaio CE Meyer-Fernandes JR Vieyra A Sola-Penna M 《Bioscience reports》2001,21(1):25-32
The plasma membrane (Ca2++Mg2+)ATPase hydrolyzes pseudo-substrates such as p-nitrophenylphosphate. Except when calmodulin is present, Ca2+ ions inhibit the p-nitrophenylphosphatase activity. In this report it is shown that, in the presence of glycerol, Ca2+ strongly stimulates phosphatase activity in a dose-dependent manner. The glycerol- and Ca2+-induced increase in activity is correlated with modifications in the spectral center of mass (average emission wavenumber) of the intrinsic fluorescence of the enzyme. It is concluded that the synergistic effect of glycerol and Ca2+ is related to opposite long-term hydration effects on the substrate binding domain and the Ca2+ binding domain. 相似文献
5.
Netticadan T Temsah RM Kawabata K Dhalla NS 《Biochemical and biophysical research communications》2002,293(2):727-732
There is increasing evidence to suggest that Ca2+-calmodulin dependent protein kinase (CaMK) regulates the sarcoplasmic reticulum (SR) function and thus plays an important role in modulating the cardiac performance. Because intracellular Ca2+-overload is an important factor underlying cardiac dysfunction in a heart disease, its effect on SR CaMK was examined in the isolated rat heart preparations. Ca2+-depletion for 5 min followed by Ca2+-repletion for 30 min, which is known to produce intracellular Ca2+-overload, was observed to attenuate cardiac function as well as SR Ca2+-uptake and Ca2+-release activities. Attenuated SR function in the heart was associated with reduced CaMK phosphorylation of the SR Ca2+-cycling proteins such as Ca2+-release channel, Ca2+-pump ATPase, and phospholamban, decreased CaMK activity, and depressed levels of SR Ca2+-cycling proteins. These results indicate that alterations in cardiac performance and SR function following the occurrence of intracellular Ca2+-overload may partly be due to changes in the SR CaMK activity. 相似文献
6.
Creation by mutagenesis of a mammalian Ca(2+) channel beta subunit that confers praziquantel sensitivity to a mammalian Ca(2+) channel 总被引:2,自引:0,他引:2
Kohn AB Roberts-Misterly JM Anderson PA Greenberg RM 《International journal for parasitology》2003,33(12):1303-1308
Voltage-gated Ca(2+) channel beta subunits are important modulators of the pore-forming alpha(1) subunit. We have cloned two schistosome beta subunits that confer sensitivity to the antischistosomal drug praziquantel (PZQ) to an otherwise insensitive mammalian alpha(1) subunit. The primary site of beta subunit interaction with alpha(1) subunits is the beta interaction domain (BID). The BID contains two conserved serines (225, 235 in rat beta2a) that constitute consensus sites for protein kinase C phosphorylation. However, these serines are absent in these schistosome beta subunits. Here we show that the capability to confer PZQ sensitivity can be created in the rat beta2a subunit by eliminating both serines in the BID. These results are consistent with, and should help our understanding of, the selective toxicity of PZQ. 相似文献
7.
Yamashita M 《FEBS letters》2006,580(21):4979-4983
Ca(2+) release from intracellular Ca(2+) stores, a pivotal event in Ca(2+) signaling, is a 'quantal' process; it terminates after a rapid release of a fraction of stored Ca(2+). To explain the 'quantal' nature, 'all-or-none' model and 'steady-state' model were proposed. This article shortly reviews these hypotheses and considers a recently proposed mechanism, 'luminal potential' model, in which the membrane potential of Ca(2+) store regulates Ca(2+) efflux. By reassessing the 'quantal' nature, other important features of Ca(2+) signaling, oscillation and synchronization, are highlighted. The mechanism for 'quantal' Ca(2+) release may underlie the temporal and spatial control of Ca(2+) signaling. 相似文献
8.
Testosterone is a potent inhibitor of L-type Ca(2+) channels 总被引:3,自引:0,他引:3
Scragg JL Jones RD Channer KS Jones TH Peers C 《Biochemical and biophysical research communications》2004,318(2):503-506
Testosterone administration is beneficial in alleviating myocardial ischaemia in men with significant coronary artery disease (CAD), a condition which is associated with hypotestosteronaemia. Infusion of physiological concentrations of testosterone into coronary arteries at angiography results in rapid vasodilatation in patients with CAD. Whilst the cardiovascular benefits of testosterone have long been documented, the underlying mechanism(s) have not yet been revealed. Here, we have investigated whether testosterone might act like widely prescribed antihypertensive dihydropyridines, as an endogenous Ca(2+) channel antagonist. To do this, we used the whole-cell patch-clamp technique to record Ca(2+) currents from the A7r5 smooth muscle cell line and HEK 293 cells stably expressing either L- or T-type Ca(2+) channels. We demonstrate that testosterone directly inhibited both native and human recombinant vascular L-type Ca(2+) channels in a manner that was voltage-independent and, crucially, displayed an IC(50) value of 38 nM, a value within the physiological range. At higher (supraphysiological) concentrations both native and human recombinant T-type channels were also inhibited by testosterone. Our data indicate that testosterone acts like widely prescribed antihypertensive dihydropyridines to reduce Ca(2+) influx into vascular smooth muscle and so promote vasodilation. This effect is likely to account for its beneficial cardiovascular actions. 相似文献
9.
Komori Y Tanaka M Kuba M Ishii M Abe M Kitamura N Verkhratsky A Shibuya I Dayanithi G 《Cell calcium》2010,48(6):324-332
Multiple mechanisms that maintain Ca(2+) homeostasis and provide for Ca(2+) signalling operate in the somatas and neurohypophysial nerve terminals of supraoptic nucleus (SON) neurones. Here, we examined the Ca(2+) clearance mechanisms of SON neurones from adult rats by monitoring the effects of the selective inhibition of different Ca(2+) homeostatic molecules on cytosolic Ca(2+) ([Ca(2+)](i)) transients in isolated SON neurones. In addition, we measured somatodendritic vasopressin (AVP) release from intact SON tissue in an attempt to correlate it with [Ca(2+)](i) dynamics. When bathing the cells in a Na(+)-free extracellular solution, thapsigargin, cyclopiazonic acid (CPA), carbonyl cyanide 3-chlorophenylhydrazone (CCCP), and the inhibitor of plasma membrane Ca(2+)-ATPase (PMCA), La(3+), all significantly slowed down the recovery of depolarisation (50 mM KCl)-induced [Ca(2+)](i) transients. The release of AVP was stimulated by 50 mM KCl, and the decline in the peptide release was slowed by Ca(2+) transport inhibitors. In contrast to previous reports, our results show that in the fully mature adult rats: (i) all four Ca(2+) homeostatic pathways, the Na(+)/Ca(2+) exchanger, the endoplasmic reticulum Ca(2+) pump, the plasmalemmal Ca(2+) pump and mitochondria, are complementary in actively clearing Ca(2+) from SON neurones; (ii) somatodendritic AVP release closely correlates with intracellular [Ca(2+)](i) dynamics; (iii) there is (are) Ca(2+) clearance mechanism(s) distinct from the four outlined above; and (iv) Ca(2+) homeostatic systems in the somatas of SON neurones differ from those expressed in their terminals. 相似文献
10.
We report here a combination of site-directed mutations that eliminate the high-affinity Ca(2+) response of the large-conductance Ca(2+)-activated K(+) channel (BK(Ca)), leaving only a low-affinity response blocked by high concentrations of Mg(2+). Mutations at two sites are required, the "Ca(2+) bowl," which has been implicated previously in Ca(2+) binding, and M513, at the end of the channel's seventh hydrophobic segment. Energetic analyses of mutations at these positions, alone and in combination, argue that the BK(Ca) channel contains three types of Ca(2+) binding sites, one of low affinity that is Mg(2+) sensitive (as has been suggested previously) and two of higher affinity that have similar binding characteristics and contribute approximately equally to the power of Ca(2+) to influence channel opening. Estimates of the binding characteristics of the BK(Ca) channel's high-affinity Ca(2+)-binding sites are provided. 相似文献
11.
Leung YM Ahmed I Sheu L Tsushima RG Diamant NE Gaisano HY 《Biochemical and biophysical research communications》2006,345(1):340-344
In low or absence of glucose, alpha-cells generate rhythmic action potentials and secrete glucagon. alpha-Cell T-type Ca(2+) channels are believed to be pacemaker channels, which are expected to open near the resting membrane potential (around -60 mV) to initiate a small depolarization. A previous publication, however, showed that alpha-cell T-type Ca(2+) channels have an activation threshold of -40 mV, which does not appear to fulfill their role as pacemakers. In this work, we investigated the Ca(2+) channel characteristics in alpha-cells of mouse-insulin-promoter green-fluorescent-protein (MIP-GFP) mouse. The beta-cells of MIP-GFP were conveniently distinguished as green cells, while immunostaining indicated that the majority of non-green cells were alpha-cells. We found that majority of alpha-cells possessed T-type Ca(2+) channels having an activation threshold of -40 mV; these cells also had high-voltage-activated (HVA) Ca(2+) channels (activation threshold of -20 mV). A novel finding here is that a minority of alpha-cells had T-type Ca(2+) channels with an activation threshold of -60 mV. This minor population of alpha-cells was, surprisingly, devoid of HVA Ca(2+) channels. We suggest that this alpha-cell subpopulation may act as pacemaker cells in low or absence of glucose. 相似文献
12.
Summary The two high affinity calcium binding sites of the cardiac (Ca2+ + Mg2+)-ATPase have been identified with the use of Eu3+. Eu3+ competes for the two high affinity calcium sites on the enzyme. With the use of laser-pulsed fluorescent spectroscopy, the environment of the two sites appear to be heterogeneous and contain different numbers of H2O molecules coordinated to the ion. The ion appears to be occluded even further in the presence of ATP. Using non-radiative energy transfer studies, we were able to estimate the distance between the two Ca2+ sites to be between 9.4 to 10.2 A in the presence of ATP. Finally, from the assumption that the calcium site must contain four carboxylic side chains to provide the 6–8 ligands needed to coordinate calcium, and based on our recently published data, we predict the peptidic backbone of the two sites. 相似文献
13.
Fluorescent ryanodine revealed the distribution of ryanodine receptors in the submembrane cytoplasm (less than a few micrometers) of cultured bullfrog sympathetic ganglion cells. Rises in cytosolic Ca(2+) ([Ca(2+)](i)) elicited by single or repetitive action potentials (APs) propagated at a high speed (150 microm/s) in constant amplitude and rate of rise in the cytoplasm bearing ryanodine receptors, and then in the slower, waning manner in the deeper region. Ryanodine (10 microM), a ryanodine receptor blocker (and/or a half opener), or thapsigargin (1-2 microM), a Ca(2+)-pump blocker, or omega-conotoxin GVIA (omega-CgTx, 1 microM), a N-type Ca(2+) channel blocker, blocked the fast propagation, but did not affect the slower spread. Ca(2+) entry thus triggered the regenerative activation of Ca(2+)-induced Ca(2+) release (CICR) in the submembrane region, followed by buffered Ca(2+) diffusion in the deeper cytoplasm. Computer simulation assuming Ca(2+) release in the submembrane region reproduced the Ca(2+) dynamics. Ryanodine or thapsigargin decreased the rate of spike repolarization of an AP to 80%, but not in the presence of iberiotoxin (IbTx, 100 nM), a BK-type Ca(2+)-activated K(+) channel blocker, or omega-CgTx, both of which decreased the rate to 50%. The spike repolarization rate and the amplitude of a single AP-induced rise in [Ca(2+)](i) gradually decreased to a plateau during repetition of APs at 50 Hz, but reduced less in the presence of ryanodine or thapsigargin. The amplitude of each of the [Ca(2+)](i) rise correlated well with the reduction in the IbTx-sensitive component of spike repolarization. The apamin-sensitive SK-type Ca(2+)-activated K(+) current, underlying the afterhyperpolarization of APs, increased during repetitive APs, decayed faster than the accompanying rise in [Ca(2+)](i), and was suppressed by CICR blockers. Thus, ryanodine receptors form a functional triad with N-type Ca(2+) channels and BK channels, and a loose coupling with SK channels in bullfrog sympathetic neurons, plastically modulating AP. 相似文献
14.
Scorpion toxins that block T-type Ca2+ channels in spermatogenic cells inhibit the sperm acrosome reaction 总被引:3,自引:0,他引:3
López-González I Olamendi-Portugal T De la Vega-Beltrán JL Van der Walt J Dyason K Possani LD Felix R Darszon A 《Biochemical and biophysical research communications》2003,300(2):408-414
The acrosome reaction (AR) is a Ca(2+)-dependent event required for sperm to fertilize the egg. The activation of T-type voltage-gated Ca(2+) channels plays a key role in the induction of this process. This report describes the actions of two toxins from the scorpion Parabuthus granulatus named kurtoxin-like I and II (KLI and KLII, respectively) on sperm Ca(2+) channels. Both toxins decrease T-type Ca(2+) channel activity in mouse spermatogenic cells and inhibit the AR in mature sperm. Saturating concentrations of the toxins inhibited at most approximately 70% of the whole-cell Ca(2+) current, suggesting the presence of a toxin-resistant component. In addition, both toxins inhibited approximately 60% of the AR, which is consistent with the participation of T-type Ca(2+) channels in the sperm AR. 相似文献
15.
da-Silva WS Bomfim FM Galina A de Meis L 《Biochemical and biophysical research communications》2003,307(3):472-476
Maize root tonoplasts are able to accumulate Ca(2+) using the energy derived from the H(+) gradient formed during PP(i) hydrolysis. Oxalate increases 6- to 10-fold the amount of Ca(2+) accumulated by tonoplast. Two apparently different K(s) values for Ca(2+) with values of 0.36 and 4.70 microM were detected when oxalate was included in the medium and the free Ca(2+) concentration in the medium was buffered with the use of EGTA. Binding of Ca(2+) to the outer surface of tonoplasts inhibits the outflow of Ca(2+) previously accumulated by the tonoplast, half-maximal inhibition being observed in presence of 1 microM Ca(2+). Thapsigargin, a specific inhibitor of Ca(2+)-ATPase, inhibits the Ca(2+) uptake driven by H(+) gradient but does not inhibit the hydrolysis of PP(i) nor the formation of a H(+) gradient. 相似文献
16.
J. Mas-Oliva 《Molecular and cellular biochemistry》1989,89(1):87-93
Summary I have investigated the effect of lead on the erythrocyte ghosts (Ca2+,Mg2+)-ATPase, with special attention to the role of calmodulin in this phenomena. Under regular incubation conditions, lead inhibits the enzyme with an IC50 of 6.0 µM. The presence of exogenously added calmodulin apparently does not change this inhibitory value. DTT added during the incubation period does not affect the inhibitory action of lead. However, when the membranes are preincubated with DTT, an important IC50 displacement is observed, either with or without added calmodulin. Since [125I]calmodulin binding to the membranes is enhanced when lead is used, the possibility of a lead/calmodulin complex that optimally stimulates the enzyme using lead concentrations between 1.0 and 10.0 µM, is suggested. Based on the experimental data, I propose two well defined actions of lead; first, an inhibitory action upon the ATPase above 1.0 µM lead, most probably related to essential sullphydryl groups in the enzyme; and second, a direct action of lead upon calmodulin at lead concentrations below 1.0 µM.A preliminary report has been presented at the 5th European Bioenergetics Conference. Aberystwyth, Wales. 20–26 July 1988. EBEC Reports. vol 5; p294 (1988). 相似文献
17.
The actions of the protonophore CCCP on intracellular Ca2+ regulation and exocytosis in chromaffin cells have been examined. Simultaneous fura-2 imaging and amperometry reveal that exposure to CCCP not only perturbs mitochondrial function but that it also alters vesicular storage of Ca2+ and catecholamines. By disrupting the pH gradient of the secretory vesicle membrane, the protonophore allows both Ca(2+) and catecholamine to leak into the cytosol. Unlike the high cytosolic Ca2+ concentrations resulting from mitochondrial membrane disruption, Ca2+ leakage from secretory vesicles may initiate exocytotic release. In conjunction with previous studies, this work reveals that catalytic and self-sustained vesicular Ca(2+) -induced exocytosis occurs with extended exposure to weak acid or base protonophores. 相似文献
18.
Nishigaki T Wood CD Tatsu Y Yumoto N Furuta T Elias D Shiba K Baba SA Darszon A 《Developmental biology》2004,272(2):376-388
Speract, a sperm-activating peptide (SAP) from sea urchin eggs, increases the intracellular concentration of Ca2+ ([Ca2+]i) and modulates sperm motility. We measured the initial sperm response to speract using its caged analog and observed, for the first time, a small but significant decrease in sperm [Ca2+]i before the increase. Both directions of the [Ca2+]i change were completely blocked in high K+ seawater. Using membrane-permeant caged cyclic nucleotides (cNMP), only cGMP induced the decrease in [Ca2+]i although both cGMP and cAMP increased the [Ca2+]i. The decrease in the [Ca2+]i induced by cGMP was more notable following a second photolytic event, once [Ca2+]i had been elevated by an initial flash. This pattern of [Ca2+]i change was confirmed in individual sperm. These results together with pharmacological evidence suggest that the initial [Ca2+]i decrease is due to a Na+/Ca2+ exchanger activity, stimulated by hyperpolarization mediated by K+ efflux through cGMP-regulated K+ channels. 相似文献
19.
Plasma membrane (PM) Na+, K+-ATPase, plays crucial roles in numerous physiological processes. Cardiac steroids (CS), such as ouabain and bufalin, specifically bind to the Na+, K+-ATPase and affect ionic homeostasis, signal transduction, and endocytosed membrane traffic. CS-like compounds, synthesized in and released from the adrenal gland, are considered a new family of steroid hormones. Previous studies showed that ouabain induces slow Ca2+ oscillations in COS-7 cells by enhancing the interactions between Na+, K+-ATPase, inositol 1,4,5-trisphosphate receptor (IP3R) and Ankyrin B (Ank-B) to form a Ca2+ signaling micro-domain. The activation of this micro-domain, however, is independent of InsP3 generation. Thus, the mechanism underlying the induction of these slow Ca2+ oscillations remained largely unclear. We now show that other CS, such as bufalin, can also induce Ca2+ oscillations. These oscillations depend on extracellular Ca2+ concentrations [Ca2+]out and are inhibited by Ni2+. Furthermore, we found that these slow oscillations are Na+out dependent, abolished by Na+/Ca2+ exchanger1 (NCX1)-specific inhibitors and markedly attenuated by NCX1 siRNA knockdown. Based on these results, a model is presented for the CS-induced slow Ca2+ oscillations in COS-7 cells. 相似文献
20.
Summary Proteolytic digestion of sarcoplasmic reticulum vesicles with trypsin has been used as a structural modification with which to examine the interaction between the ATP hydrolysis site and calcium transport sites of the (Ca2++Mg2+)-ATPase. The kinetics of trypsin fragmentation were examined and the time course of fragment production compared with ATP hydrolytic and calcium uptake activities of the digested vesicles. The initial cleavage (TD 1) of the native ATPase to A and B peptides has no effect on the functional integrity of the enzyme, hydrolytic and transport activities remaining at the levels of the undigested control. Concomitant with the second tryptic cleavage (TD 2) of the A peptide to A1 and A2 fragments, calcium transport is inhibited. Kinetic analysis demonstrates that the rate constant for inhibition of calcium uptake is correlated with the rate constant of a fragment disappearance. Both Ca2+-dependent and total ATPase activities are unaffected by this second cleavage. Passive loading of vesicles with calcium and subsequent efflux measurements show that transport inhibition is not due to increased permeability of the membrane to calcium even at substantial extents of digestion. Steady-state levels of acidstable phosphoenzyme are unaffected by either TD 1 or TD 2, indicating that uncoupling of the hydrolytic and transport functions does not increase the turnover rate of the enzyme and that TD 2 does not change the essential characteristics of the ATP hydrolysis site. Sarcoplasmic reticulum (SR) vesicles were examined for the presence of tightly bound nucleotides and are shown to contain 2.8–3.0 nmol ATP and 2.6–2.7 nmol ADP per mg SR protein. The ADP content of SR remains essentially unchanged with TD 1 cleavage of the ATPase enzyme to A and B peptides, but declines upon TD 2 in parallel with the digestion of the A fragment and the loss of calcium uptake activity of the vesicles. The ATP content is essentially constant throughout the course of trypsin digestion. The results are discussed in terms of current models of the SR calcium pump and the molecular mechanism of energy transduction. 相似文献