共查询到14条相似文献,搜索用时 15 毫秒
1.
Rate and equilibrium measurements of ryanodine binding to terminal cysternae fractions of heavy sarcoplasmic reticulum vesicles
demonstrate that its activation by high concentrations of monovalent salts is based on neither elevated osmolarity nor ionic
strength. The effect of the ions specifically depends on their chemical nature following the Hofmeister ion series for cations
(Li+ < NH+
4 < K−∼ Cs+≤ Na+) and anions (gluconate− < Cl− < NO3
−∼ ClO4
−∼ SCN−) respectively, indicating that both are involved in the formation of the salt-protein complex that can react with ryanodine.
Activation by rising salt concentrations exhibits saturation kinetics with different dissociation constants (25–11 m) and different degrees of cooperativity (n= 1.5–4.0) for the respective salts. Maximal second order binding rates between 40,000 and 80,000 (m
−1· sec−1) were obtained for chlorides and nitrates of 1a group alkali ions with the exception of lithium supporting only rates of
maximally 10,000 (M−1· sec−1). The nitrogen bases, NH+
4 and Tris+, in combination with chloride or nitrate, behave divergently. High maximal binding rates were achieved only with NH4NO3. The dissociation constants for the ryanodine–protein complexes obtained by measurements at equilibrium proved to depend
differently on salt concentration, yet, converging to 1–3 nm for the applied salts at saturating concentrations. The salts do not affect dissociation of the ryanodine protein complex
proving that the effect of salts on the protein's affinity for ryanodine is determined by their effect on the on-rate of ryanodine
binding. ATP and its analogues modify salt action resulting in elevated maximal binding rates and reduction or abolition of
binding cooperativity. Linear relations have been obtained by comparing the rates of ryanodine binding at different salt concentrations
with the rates or the initial amplitudes (15 sec) of salt induced calcium release from actively loaded heavy vesicles indicating
that the various salts promote specifically and concentration dependently channel opening and its reaction with ryanodine.
Received: 9 February 1998/Revised: 24 April 1998 相似文献
2.
Green D Pace SM Hurne AM Waring P Hart JD Dulhunty AF 《The Journal of membrane biology》2000,175(3):223-233
Interactions between the reactive disulfide fungal metabolite, gliotoxin (GTX), and rabbit skeletal ryanodine receptor (RyR)
calcium release channels have been examined. RyRs in terminal cisternae vesicles formed a covalent complex with 100 μm
35S-GTX, which was reversed by 1 mm dithiothreitol (DTT) or 1 mm glutathione. GTX (80–240 μm), added to either cytoplasmic (cis) or luminal (trans) solutions, increased the rate of Ca2+ release from SR vesicles and the frequency of opening of single RyR channels in lipid bilayers. Channel activation was reversed
upon addition of 2 mm DTT to the cis solution, showing that the activation was due to an oxidation reaction (2 mm DTT added to the cis solution in the absence of GTX did not affect RyR activity). Furthermore, RyRs were not activated by trans GTX if the cis chamber contained DTT, suggesting that GTX oxidized a site in or near the membrane. In contrast to cis DTT, 2 mm DTT in the trans solution increased RyR activity when added either alone or with 200 μm
trans GTX. The results suggest that (i) GTX increases RyR channel activity by oxidizing cysteine residues that are close to the
membrane and located on RyR, or associated proteins, and (ii) a disulfide bridge or nitrosothiol, accessible only from the
luminal solution, normally suppresses RyR channel activity. Some of the actions of GTX in altering Ca2+ homeostatsis might depend on its modification of RyR calcium channels.
Received: 12 November 1999/Revised: 14 March 2000 相似文献
3.
Single channel and [3H]ryanodine binding measurements were performed to test for a direct functional interaction between 2,3-butanedione 2-monoxime
(BDM) and the skeletal and cardiac muscle sarcoplasmic reticulum Ca2+ release channels (ryanodine receptors). Single channel measurements were carried out in symmetric 0.25 m KCl media using the planar lipid bilayer method. BDM (1–10 mm) activated suboptimally Ca2+-activated (0.5–1 μm free Ca2+) single, purified and native cardiac and skeletal release channels in a concentration-dependent manner by increasing the
number of channel events without a change of single channel conductances. BDM activated the two channel isoforms when added
to either side of the bilayer. At a maximally activating cytosolic Ca2+ concentration of 20 μm, BDM was without effect on the cardiac channel, whereas it inhibited skeletal channel activities with IC50≈ 2.5 mm. In agreement with single channel measurements, high-affinity [3H]ryanodine binding to the two channel isoforms was increased in a concentration-dependent manner at ≤1 μm Ca2+. BDM was without a noticeable effect at low (≤0.01 μm) Ca2+ concentrations. At 20 μm Ca2+, BDM inhibited the skeletal but not cardiac channel. These results suggest that BDM regulates the Ca2+ release channels from the sarcoplasmic reticulum of skeletal and cardiac muscle in a concentration, Ca2+ and tissue-dependent manner.
Received: 31 December 1998/Revised: 9 March 1999 相似文献
4.
N. Hadad H.E. Meyer M. Varsanyi S. Fleischer V. Shoshan-Barmatz 《The Journal of membrane biology》1999,170(1):39-49
Cardiac sarcoplasmic reticulum (SR) contains an endogenous phosphorylation system that under specific conditions phosphorylates
two proteins with apparent molecular masses of 150 and 130 kDa. The conditions for their phosphorylation are as for the skeletal
muscle sarcalumenin and the histidine-rich Ca2+ binding protein (HCP) with respect to: (i) Ca2+ and high concentrations of NaF are required; (ii) phosphorylation is obtained with no added Mg2+ and shows a similar time course and ATP concentration dependence; (iii) inhibition by similar concentrations of La3+; (iv) phosphorylation is obtained with [γ-32P]GTP; (v) ryanodine binding is inhibited parallel to the phosphorylation of the two proteins. The endogenous kinase is identified
as casein kinase II (CK II) based on its ability to use GTP as effectively as ATP, and its inhibition by La3+. The association of CK II with the cardiac SR, even after EGTA extraction at alkaline pH, is demonstrated using antibodies
against CK II. The cardiac 130 kDa protein is identified as sarcalumenin based on its partial amino acid sequence and its
blue staining with Stains-All. Cardiac sarcalumenin is different from the skeletal muscle protein based on electrophoretic
mobilities, immunological analysis, peptide and phosphopeptide maps, as well as amino acid sequencing. Preincubation of SR
with NaF and ATP, but not with NaF and AMP-PNP caused strong inhibition of ryanodine binding. This is due to decrease in Ca2+- and ryanodine-binding affinities of the ryanodine receptor (RyR) by about 6.6 and 18-fold, respectively.
These results suggest that cardiac sarcalumenin is an isoform of the skeletal muscle protein. An endogenous CK II can phosphorylate
sarcalumenin, and in parallel to its phosphorylation the properties of the ryanodine receptor are modified.
Received: 15 December 1998/Revised: 25 March 1999 相似文献
5.
The modulation of the calmodulin-induced inhibition of the calcium release channel (ryanodine receptor) by two sulfhydryl
oxidizing compounds, 4-(chloromercuri)phenyl–sulfonic acid (4-CMPS) and 4,4′-dithiodipyridine (4,4′-DTDP) was determined by
single channel current recordings with the purified and reconstituted calcium release channel from rabbit skeletal muscle
sarcoplasmic reticulum (HSR) and [3H]ryanodine binding to HSR vesicles. 0.1 μm CaM reduced the open probability (P
o
) of the calcium release channel at maximally activating calcium concentrations (50–100 μm) from 0.502 ± 0.02 to 0.137 ± 0.022 (n= 28), with no effect on unitary conductance. 4-CMPS (10–40 μm) and 4,4′-DTDP (0.1–0.3 mm) induced a concentration dependent increase in P
o (> 0.9) and caused the appearance of longer open states. CaM shifted the activation of the calcium release channel by 4-CMPS
or 4,4′-DTDP to higher concentrations in single channel recordings and [3H]ryanodine binding. 40 μm 4-CMPS induced a near maximal (P
o
> 0.9) and 0.3 mm 4,4′-DTDP a submaximal (P
o
= 0.74) channel opening in the presence of CaM, which was reversed by the specific sulfhydryl reducing agent DTT. Neither
4-CMPS nor 4,4′-DTDP affected Ca-[125I]calmodulin binding to HSR. 1 mm MgCl2 reduced P
o
from 0.53 to 0.075 and 20–40 μm 4-CMPS induced a near maximal channel activation (P
o
> 0.9). These results demonstrate that the inhibitory effect of CaM or magnesium in a physiological concentration is diminished
or abolished at high concentrations of 4-CMPS or 4,4′-DTDP through oxidation of activating sulfhydryls on cysteine residues
of the calcium release channel.
Received: 22 July 1999/Revised: 15 November 1999 相似文献
6.
S. Shevchenko W. Feng M. Varsanyi V. Shoshan-Barmatz 《The Journal of membrane biology》1998,161(1):33-43
A 94 kDa large subunit thiol-protease, as identified by anti-calpain antibodies, has been isolated from skeletal muscle junctional
sarcoplasmic reticulum (SR). This protease cleaves specifically the skeletal muscle ryanodine receptor (RyR)/Ca2+ release channel at one site resulting in the 375 kDa and 150 kDa fragments. The 94 kDa thiol-protease degrades neither other
SR proteins nor the ryanodine receptor of cardiac nor brain membranes. The partially purified 94 kDa protease, like the SR
associated protease, had an optimal pH of about 7.0, was absolutely dependent on the presence of thiol reducing reagents,
and was completely inhibited by HgCl2, leupeptin and the specific calpain I inhibitor. However, while the SR membrane-associated protease requires Ca2+ at a submicromolar concentration, the isolated thiol-protease has lost the Ca2+ requirement.
The 94 kDa thiol-protease had no effect on ryanodine binding but modified the channel activity of RyR reconstituted into planar
lipid bilayer: in a time-dependent manner, the channel activity decreases and within several minutes the channel is converted
into a subconducting state. The protease-modified channel activity is still Ca2+-dependent and ryanodine sensitive.
This 94 kDa thiol-protease cross react with anti-calpain antibodies thus, may represent the novel large subunit of the skeletal
muscle specific calpain p94.
Received: 10 December 1996/Revised: 11 August 1997 相似文献
7.
Modification of the Conductance and Gating Properties of Ryanodine Receptors by Suramin 总被引:3,自引:0,他引:3
Suramin, a polysulfonated napthylurea, increases the open probability and the single-channel conductance of rabbit skeletal
and sheep cardiac ryanodine receptor channels. The main mechanism for the increase in P
o
is an increase in the duration of open lifetimes. The effects on conduction and gating are completely reversible and involve
an interaction with the cytosolic side of the channel. 10 mm dithiothreitol had no effect on the suramin-induced increase in conductance or P
o
. Therefore oxidation of sulfhydryl groups on the channels does not appear to be involved. Suramin has been used as an antagonist
of ATP at P2 purinoceptors, however, we find that suramin does not antagonize the effect of ATP at skeletal or cardiac ryanodine receptor
channels. The unusual gating kinetics induced by suramin suggest that it does not interact with the adenine nucleotide binding
site on the ryanodine receptor but rather binds at a novel site(s). The suramin-induced changes to channel gating and conduction
do not prevent the characteristic modification of single-channel properties by micromolar ryanodine.
Received: 19 March 1996/Revised: 5 June 1996 相似文献
8.
The mechanisms underlying skeletal muscle functional impairment and structural changes with advanced age are only partially
understood. In the present study, we support and expand our theory about alterations in sarcolemmal excitation-sarcoplasmic
reticulum Ca2+ release-contraction uncoupling as a primary skeletal muscle alteration and major determinant of weakness and fatigue in mammalian
species including humans. To test the hypothesis that the number of RYR1 (ryanodine receptor) uncoupled to DHPR (dihydropyridine
receptor) increases with age, we performed high-affinity ligand binding studies in soleus, extensor digitorum longus (EDL) and in a pool of several skeletal muscles consisting of a mixture of fast- and slow-twitch muscle fibers in middle-aged
(14-month) and old (28-months) Fisher 344 Brown Norway F1 hybrids rats. The number of DHPR, RYR1, the coupling between both
receptors expressed as the DHPR/RYR1 maximum binding capacity, and their dissociation constant for high-affinity ligands were
measured. The DHPR/RYR1 ratio was significantly reduced in the three groups of muscles (pool: 1.03 ± 0.15 and 0.80 ± 0.11,
soleus: 0.44 ± 0.12 and 0.26 ± 0.10, and EDL: 0.95 ± 0.14 and 0.68 ± 0.10, for middle-aged and old muscles, respectively). These
data support the concept that DHPR-RYR1 uncoupling results in alterations in the voltage-gated sarcoplasmic reticulum Ca2+ release mechanism, decreases in myoplasmic Ca2+ elevation in response to sarcolemmal depolarization, reduced Ca2+ supply to contractile proteins and reduced contraction force with aging.
Received: 26 August 1996/Revised: 30 December 1996 相似文献
9.
J.I. Kourie 《The Journal of membrane biology》1999,172(1):25-36
Data obtained with the lipid bilayer technique indicate that cis (cytoplasmic) concentration of 4.4–22 mm hydrogen peroxide (H2O2), is a water-soluble oxidant. [H2O2]
cis
(n= 26) reversibly inhibits the multisubconductance SCl channel of the sarcoplasmic reticulum vesicles from rabbit skeletal
muscle. At −40 mV, the mean values of the current amplitude (I) and the probability of the SCl channel being open (P
o
) were reduced significantly (n= 8) from −6.14 ± 0.42 pA and 0.69 ± 0.06 (for all conductance levels) in control 0.0 mm [H2O2]
cis
to −1.10 ± 0.51 pA and 0.13 ± 0.04 (for the intermediate subconductance states) in 8.8 mm [H2O2]
cis
, respectively. The [H2O2]
cis
-induced decrease in P
o
is mainly due to a decrease in the mean open time T
o
. The mechanism of [H2O2]
cis
effects on the multiconductance SCl channel is characterized by a mode shift in the channel state from the main conductance
state to the low subconductance states. The estimated concentration of the [H2O2]
cis
for the half inhibitory constant, K
i
, was 11.78 mm, higher than the estimated 8.0 and 8.1 mm for the parameters P
o
and T
o
, respectively, indicating that the conductance of the SCl channel is less sensitive than the gating kinetics of the channel.
After a lag period of between 30 to 60 sec, the lipophilic SH-oxidizing agent 4,4′-dithiodipyridine (4,4′-DTDP) added to the
cis side at 1.0 mm removed the inhibitory effects of 8.8 mm [H2O2]
cis
. The 4,4′-DTDP-enhanced SCl channel activity was blocked after the addition of 0.5 mm ATP to the cis side of the channel. The addition of 1.0 mm 4,4′-DTDP to the cis or trans solutions facing an SCl channel already subjected to 0.5 mm [ATP]
cis
or [ATP]
trans
failed to activate the ATP-inhibited SCl channel. These findings suggest that 4,4′-DTDP is not preventing the binding of
ATP to its binding site on the channel protein. The interaction of H2O2 with the SCl channel proteins is consistent with a thiol-disulfide redox state model for regulating ion transport, where
SH groups can directly modify the function of the channel and/or the availability of regulatory sites on the channel proteins.
The H2O2 effects on the Ca2+ countercurrent through the SCl channel are also consistent with H2O2-modification of the mechanisms involved in the Ca2+ regulation, which underlies excitation-contraction coupling in skeletal muscle.
Received: 27 April 1999/Revised: 1 July 1999 相似文献
10.
M.C. García Z. Hernández-Gallegos J. Escamilla J.A. Sánchez 《The Journal of membrane biology》2001,184(2):121-129
Calciseptine is a natural peptide consisting of 60 amino acids with four disulfide bonds. The peptide is a natural L-type
Ca2+-channel blocker in heart and other systems, but its actions in skeletal muscle have not been previously described. The aim
of this study is to characterize the effects of calciseptine on L-type Ca2+ channels of skeletal muscle and on contraction. Whole-cell, patch-clamp experiments were performed to record Ca2+ currents (I
Ca) from mouse myotubes, whereas Vaseline-gap voltage-clamp experiments were carried out to record I
Ca from frog skeletal muscle fibers. We found that calciseptine acts as a channel agonist in skeletal muscle, increasing peak
I
Ca by 37% and 49% in these two preparations. Likewise, the peptide increased intramembrane charge movement, though it had little
effect on contraction. The molecular analysis of the peptide indicated the presence of a local, electrostatic potential that
resembles that of the 1,4-dihydropyridine agonist Bay K 8644. These observations suggest that calciseptine shares the properties
of 1,4-dihydropyridine derivatives in modulating the permeation of divalent cations through L-type channels.
Received: 18 December 2000/Revised: 16 July 2001 相似文献
11.
García MC Farías JM Escamilla J Sánchez-Armass S Sánchez JA 《The Journal of membrane biology》1999,168(2):141-148
The effects of a long-term blockade of L-type Ca2+ channels on membrane currents and on the number of dihydropyridine binding sites were investigated in skeletal muscle fibers.
Ca2+ currents (I
Ca) and intramembrane charge movement were monitored using a voltage-clamp technique. The peak amplitude of I
Ca increased by more than 40% in fibers that were previously incubated for 24 hr in solutions containing the organic Ca2+ channel blocker nifedipine or in Ca2+-free conditions. A similar incubation period with Cd2+, an inorganic blocker, produced a moderate increase of 20% in peak I
Ca. The maximum mobilized charge (Q
max) increased by 50% in fibers preincubated in Ca2+-free solutions or in the presence of Cd2+.
Microsomal preparations from frog skeletal muscle were isolated by differential centrifugation. Preincubation with Cd2+ prior to the isolation of the microsomal fraction doubled the number of 3H-PN200-110 binding sites and produced a similar increase in the values of the dissociation constant. The increase in the
number of binding sites is consistent with the increase in the peak amplitude of I
Ca as well as with the increase in Q
max.
Received: 31 August 1998/Revised: 7 December 1998 相似文献
12.
R. Barrett-Jolley A. Comtois N.W. Davies P.R. Stanfield N.B. Standen 《The Journal of membrane biology》1996,152(2):111-116
We investigated the action of adenosine and GTP on KATP channels, using inside-out patch clamp recordings from dissociated single fibers of rat flexor digitorum brevis (FDB) skeletal
muscle. In excised patches, KATP channels could be activated by a combination of an extracellular adenosine agonist and intracellular Mg2+-ATP and GTP or GTP-γ-S. The activation required hydrolyzable ATP and could be partially reversed with Mg2+, suggesting that it may involve a G-protein dependent phosphorylation of KATP channels. We found that KATP channels of the rat FDB could not be activated by Mg2+-ATP alone or by Mg2+-ATP in the presence of extracellular adenosine. Patches whose channel activity had been `rundown' by Ca2+ could not be recovered by adenosine, GTP or Mg2+-ATP. KATP channels activated by adenosine receptor agonists had a similar ATP sensitivity to those under control conditions; but adenosine
appears to be able to switch these KATP channels from an inactive to an active mode.
Received: 29 December 1995/Revised: 22 March 1996 相似文献
13.
K-Cl cotransport (COT) is the coupled movement of K and Cl, present in most cells, associated with regulatory volume decrease,
susceptible to oxidation and functionally overexpressed in sickle cell anemia. The aim of this study was to characterize the
effect of the oxidant nitrite (NO2
−) on K-Cl COT. NO2
− is a stable metabolic end product of the short-lived highly reactive free radical nitric oxide (NO), an oxidant and modulator
of ion channels, and a vasodilator. In some systems, the response to NO2
− is identical to that of NO. We hypothesized that NO2
− activates K-Cl COT. Low potassium (LK) sheep red blood cells (SRBCs) were used as a model. The effect of various concentrations
(10−6 to 10−1
m) of NaNO2 was studied on K efflux in hypotonic Cl and NO3 media, Cl-dependent K efflux (K-Cl COT), glutathione (GSH), and methemoglobin (MetHb) formation. In support of our hypothesis,
K efflux and K-Cl COT were stimulated by increasing concentrations of NaNO2. Stimulation of K efflux was dependent upon external Cl and exhibited a lag phase, consistent with activation of K-Cl COT
through a regulatory mechanism. Exposure of LK SRBCs to NaNO2 decreased GSH, an effect characteristic of a thiol-oxidizing agent, and induced MetHb formation. K-Cl COT activity was positively
correlated with Methb formation. N-ethyl-maleimide (NEM), a potent activator of K-Cl COT, was used to assess the mechanism
of NO2
− action. The results suggest that NEM and NO2
− utilize at least one common pathway for K-Cl COT activation. Since NaNO2 is also a well known vasodilator, the present findings suggest a role of K-Cl COT in vasodilation.
Received: 15 January 1998/Revised: 3 September 1998 相似文献
14.
V. Casavola L. Guerra S.J. Reshkin K.A. Jacobson F. Verrey H. Murer 《The Journal of membrane biology》1996,151(3):237-245
The effect of adenosine regulation on sodium and chloride transport was examined in cultured A6 renal epithelial cells. Adenosine and its analogue N6-cyclopentyladenosine (CPA) had different effects on short-circuit current (I
sc) depending on the side of addition. Basolateral CPA addition induced an approximately threefold increase of the I
sc that reached a maximum effect 20 min after addition and was completely inhibited by preincubation with either an A2 selective antagonist, CSC, or the sodium channel blocker, amiloride. Apical CPA addition induced a biphasic I
sc response characterized by a rapid fourfold transient increase over its baseline followed by a decline and a plateau phase
that were amiloride insensitive. The A1 adenosine antagonist, CPX, completely prevented this response. This I
sc response to apical CPA was also strongly reduced in Cl−-free media and was significantly inhibited either by basolateral bumetanide or apical DPC preincubation. Only basolateral
CPA addition was able to induce an increase in cAMP level. CPA, added to cells in suspension, caused a rapid rise in [Ca2+]
i
that was antagonized by CPX, not affected by CSC and prevented by thapsigargin preincubation. These data suggest that basolateral
CPA regulates active sodium transport via A2 adenosine receptors stimulating adenylate cyclase while apical CPA regulates Cl− secretion via A1 receptor-mediated changes in [Ca2+]
i
. 相似文献