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Y Kurachi

Publications and source records attributed to Y Kurachi.

At least 91 records · Page 5Linked to original sources
PubMed1996

Actin microfilament disrupters enhance K(ATP) channel opening in patches from guinea-pig cardiomyocytes.

1. To determine whether actin filament networks are associated with the regulation of ATP-sensitive K+ (K(ATP)) channel activity, single channel currents were measured in the inside-out configuration, and cytoskeletal disrupters applied to the internal side of patches excised from guinea-pig ventricular myocytes. 2. Treatment of patches with DNase I (10-200 micrograms ml(-1)), which forms complexes with G-actin and prevents actin filament formation, antagonized the ATP-induced inhibition of K(ATP) channels. 3. In the absence of ATP, DNase I did not increase K(ATP) channel activity. 4. When denatured by boiling or co-incubated with purified actin subunits (200 mu g ml(-1)), DNase I(100 mu g ml(-1)) did not antagonize the ATP-induced inhibition of K(ATP) channels. 5. The DNase I-induced decrease in the sensitivity of K(ATP) channels towards ATP-induced inhibition was partially restored by addition of purified actin subunits (200 micrograms ml(-1)). 6. Cytochalasin B (10 microM), another actin filament disrupter, but neither taxol nor nocodazole (30-100 microM), two antimicrotubule agents, enhanced K(ATP) channel activity in the presence of ATP. 7. Hence, actin filament disrupters can attenuate the ATP-dependent inhibitory gating of K(ATP) channels. This suggests that subsarcolemmal actin filament networks may be associated with the regulation of cardiac K(ATP) channels.

Actin Cytoskeleton↗
PubMed1996

G protein-gated K+ channel (GIRK1) protein is expressed presynaptically in the paraventricular nucleus of the hypothalamus.

We prepared a specific antibody against GIRK1, the major subunit of the G protein-gated K+ (KG) channel. Immunohistochemical study revealed that GIRK1 immunoreactivity was detected in the presynaptic, but not in the postsynaptic, region in the paraventricular nucleus of the rat hypothalamus (PVN). Therefore, activation of the KG channel may underlie presynaptic inhibition of neurotransmitter release by various agonists, such as dopamine, noradrenaline, opioids, and histamine, in PVN.

Adenosine↗
PubMed1996

Differential distribution of classical inwardly rectifying potassium channel mRNAs in the brain: comparison of IRK2 with IRK1 and IRK3.

Distribution of IRK2 inwardly rectifying potassium channel mRNA in the mouse brain was studied using in situ hybridization histochemistry and compared with those of other classical inwardly rectifying potassium channel (IRK1 and IRK3) mRNAs. All these IRK channel mRNAs were detected in neurons, but not in glial cells. Their distribution patterns in the brain were, however, quite divergent: IRK2 mRNA was detected extremely high in granule cells of cerebellum, relatively high in motor trigeminal nucleus and moderate in olfactory bulb, piriform cortex, cerebral cortex, CA1 through CA3 regions of hippocampus, dentate gyrus and pontine nucleus. On the other hand, IRK1 mRNA was expressed throughout whole brain but in particular subsets of neurons, and IRK3 mRNA was in forebrain. Expression of these three IRK mRNAs overlapped in hippocampus, olfactory bulb, and cerebral cortex. This differential distribution of IRK mRNAs suggests that each of these channels has its specific function in regulation of the excitability of brain neurons.

Animals↗
PubMed1996

A novel ubiquitously distributed isoform of GIRK2 (GIRK2B) enhances GIRK1 expression of the G-protein-gated K+ current in Xenopus oocytes.

We have isolated a novel variant form of GIRK2, designated GIRK2B, from mouse brain cDNA library. GIRK2B was much shorter than the first type of GIRK2 (GIRK2A), but its amino acid sequence was identical to the corresponding part of GIRK2A except the C-terminal eight amino acid residues. When GIRK2B cRNA was co-injected with GIRK1 and m2-receptor cRNAs to Xenopus oocytes, acetylcholine-induction of the inwardly rectifying K+ current was enhanced dramatically. This suggests that GIRK2B can form a heteromultimeric G-protein-gated K+ channel with GIRK1. The reverse transcription polymerase chain reaction analysis showed that GIRK2B mRNA distributed much more broadly than GIRK1 mRNA. Therefore, GIRK2B might also play other unrecognized roles in various tissues than to form a K+ channel with GIRK1.

Amino Acid Sequence↗
PubMed1996

Endothelin-1 and vasopressin activate Ca(2+)-permeable non-selective cation channels in aortic smooth muscle cells: mechanism of receptor-mediated Ca2+ influx.

The effects of vasopressin and endothelin-1 on cultured aortic smooth muscle cell lines (A7r5) were investigated by measurements of intracellular calcium [Ca2+]i and the patch-clamp techniques. Vasopressin and endothelin-1 (100 nM) evoked an initial peak followed by a smaller sustained rise of [Ca2+]i in the presence of extracellular calcium [Ca2+]o. In the absence of [Ca2+]o, only the initial peak of [Ca2+]i was observed. Therefore, the initial peak of [Ca2+]i was mainly due to calcium release from the storage sites, whereas the later sustained rise of [Ca2+]i was due to the calcium entry from outside. The sustained rise of [Ca2+]i was unaffected by nifedipine (10 microM) significantly, but was completely abolished by La3+ (1 mM). Under current clamp conditions with K(+)-internal solution, vasopressin and endothelin-1 (100 nM) produced hyperpolarization, then followed by depolarization. Under voltage clamp conditions at a holding potential of -40 mV, both vasopressin and endothelin-1 first activated the outward current, then followed by a long-lasting inward current with a high noise level. The first outward current was abolished by charybdotoxin (100 nM), Cs+ in the patch pipette and high EGTA (10 mM) in the pipette, suggesting that it was a Ca(2+)-sensitive K+ current (IK.Ca). The inward current was still elicited with the patch pipette containing Cs(+)-internal solution, and reversed at about 0 mV. The reversal potential was not significantly altered by the replacement of [Cl-]i or [Cl-]o, proposing that the inward current is a cation selective channel (IN.S.). The inward current was also observed even when extracellular cations are Ca2+. La3+ (1 mM), Cd2+ (1 mM) completely abolished the vasopressin-induced (IN.S.), however, nifedipine (10 microM) failed to inhibit it significantly. Single channel activities were recorded in the cell-attached configurations when vasopressin or endothelin-1 was applied to the bathing solution. The unitary conductance of the channels was approximately 20 pS with 140 mM Na+, Cs+, or K+ in the pipette, but was 15 pS with 110 mM Ca2+ in the pipette. Permeabilities sequence calculated from the reversal potentials was Na+ not equal to Cs+ not equal to K+ > Ca+. These results provide evidence that calcium entry and membrane depolarization elicited by vasopressin or endothelin-1 are mediated by a receptor-mediated Ca(2+)-permeable non-selective cation channel in aortic smooth muscle cells.

Aorta, Thoracic↗
PubMed1996

Molecular and functional heterogeneity of inward rectifier potassium channels in brain and heart.

We have cloned cDNAs encoding three background inward rectifier potassium channels from a mouse brain cDNA library. We designated them mouse brain (MB)- IRK1, MB-IRK2, and MB-IRK3, based on their amino acid sequences and the electrophysiological properties of currents expressed in Xenopus oocytes. Xenopus oocytes injected with cRNAs derived from these clones expressed K+ currents that showed classical inward rectifier potassium channel characteristics at whole cell current level and that were blocked by Ba2+ and Ca+ in a concentration- and voltage- dependent manner. The single channel recordings revealed, however, that the unitary conductance of MB-IRKI was approximately 22pS; MB-IRK2, approximately 34pS; and MB-IRK3, approximately 12pS. By Northern blot analysis, MB-IRKI was more predominantly expressed in forebrain than in cerebellum, and vice versa in the case MB-IRK2. MB-IRK3 was expressed specifically in forebrain. On the other hand, MB-IRK1 and MB-IRK2, but not MB-IRK3, were expressed in heart. These results indicate the heterogeneity of the molecular structure and functional role of the two transmembrane region type of background inward rectifier potassium channels in brain and heart.

Amino Acid Sequence↗
PubMed1996

A functional model for G protein activation of the muscarinic K+ channel in guinea pig atrial myocytes. Spectral analysis of the effect of GTP on single-channel kinetics.

To elucidate the functional interaction between the active G protein subunit (GK*) and the cardiac muscarinic K+ (KACh) channel, the effect of intracellular GTP on the channel current fluctuation in the presence of 0.5 microM extracellular acetylcholine was examined in inside-out patches from guinea pig atrial myocytes using spectral analysis technique. The power density spectra of current fluctuations induced at various concentrations of GTP ([GTP]) were well fitted by the sum of two Lorentzian functions. Because the channel has one open state, the open-close transitions of the channel gate represented by the spectra could be described as C2<-->C1<-->O. As [GTP] was raised, the channel activity increased in a positive cooperative manner. The powers of the two Lorentzian components concomitantly increased, while the corner frequencies and the ratio of the powers at 0 Hz remained almost constant. This indicates that G protein activation did not affect the gating of each channel but mainly increased the number of functionally active channels in the patch to enhance the channel activity. Regulation of the number of functionally active channels could be described by a slow transition of the channel states, U (unavailable)<-->A (available), which is independent of the gating. The equilibrium of this slow transition was shifted by GTP from U to A. Monod-Wyman-Changeux's allosteric model for the channel state transition(U<-->A) could well describe the positive cooperative increase in the channel availability by GTP, assuming that, in the presence of saturating concentrations of ACh, [GK*] linearly increased as [GTP] was raised in our experimental range. The model indicates that the cardiac KACh channel could be described as a multimer composed of four or more functionally identical subunits, to each of which one GK* binds.

Animals↗
PubMed1996

[Molecular structure and expression of ion channels in the heart].

Ion channels are essential for the generation and maintenance of the cardiac action potential. The successful cloning of ion channels has shed considerable light into their structure and function. The combination of molecular biological and electrophysiological studies has paved the ground to develop general concepts for understanding properties of these channels, such as gating mechanism, kinetics of activation and inactivation, ion selectivity and pharmacological properties. Recently, several cDNAs encoding cardiac inwardly rectifying K+ channels, including classical inwardly rectifying, G-protein-gated, and ATP-sensitive K+ channels, have been isolated. Identification of these molecular mechanisms may point to novel approaches to the treatment of cardiac arrhythmias. This review focuses on the molecular properties and functional roles of cardiac ion channels.

Adenosine Triphosphate↗
PubMed1996

[Molecular and biophysical aspects of potassium channels].

K+ channels of known sequence fall into two distantly related families, voltage-dependent channels and inwardly rectifying channels. Several types of inwardly rectifying K+ channels, containing two putative transmembrane domains in each subunit, have recently been cloned and can be classified into four groups: 1) IRK family; classical inward rectifying, 2) GIRK family; G-protein-activated, 3) KATP family; ATP-inhibited, and 4) KAB family; ATP-dependent. Recent studies have revealed that atrial and neuronal G-protein-gated inwardly rectifying K+ channels function as heteromultimers of two distinct GIRKs, and that pancreatic beta cell KATP channels are a complex composed of at least two subunits, a member of the ATP-inhibited inwardly rectifying K+ channel family (BIR) and the sulfonylurea receptor.

Adenosine Triphosphate↗
PubMed1995

17beta-Estradiol inhibits the voltage-dependent L-type Ca2+ currents in aortic smooth muscle cells.

To elucidate the mechanisms of estrogens-induced relaxation effects on vascular smooth muscle cells, the effects of estrogens and the related hormones were examined in cultured rat thoracic aortic smooth muscle cell lines (A7r5), using the whole-cell voltage clamp technique. The patch pipette was filled with 140 mM CsCl- or KCl-containing internal solution. With CsCl-internal solution, 17beta-estradiol and synthetic estrogens, ethynylestradiol and diethylstilbestrol (0.1-30 mu M) inhibited the Ba2+ inward current (IBa) through the voltage-dependent L-type Ca2+ channel in a concentration-dependent and reversible manner. The potency of the inhibitory effects on IBa was 17beta-estradiol < ethynylestradiol < diethylstilbestrol. 17beta-Estradiol (10 mu M) appeared to reduce the maximal conductance of IBa with only a slight shift of voltage-dependency of inactivation and to affect IBa in a use-independent fashion. On the other hand, testosterone and progesterone (30 mu M) failed to affect IBa. At a holding potential of -40 mV, both vasopressin and endothelin-1 (100 nM) activated a long-lasting inward current. After endothelin-1 (100 nM) activated the current, the additional application of vasopressin (100 nM) could not induce it furthermore, suggesting that each agonist activates the same population of the channels. The reversal potential of the current was about 0 mV and was not significantly altered by replacement of [Cl-]i or [Cl-]0 and the inward current was also observed even when extracellular cations are Ca2+, proposing that it was a Ca2+-permeable non-selective cation channel (IN.S.). La3+ or Cd2+ (1 nM) completely abolished IN.S., however, nifedipine (10 mu M) failed to inhibit it at all. Diethylstilbestrol (1-30 mu M) suppressed the IN.S. evoked by both endothelin-1 and vasopressin in a concentration-dependent manner, while 17beta-estradiol, ethynylestradiol, progesterone and testosterone (30 mu M) failed to inhibit it significantly. In addition, at a holding potential of +0 mV, 17beta-estradiol by itself did not affect the holding currents, and did not inhibit K+ currents evoked by endothelin-1 or vasopressin, possibly due to the Ca2+ release from the storage sites. These results suggest that 17beta-estradiol may play a role in regulating vascular tone, selectively by inhibiting the voltage-dependent L-type Ca2+ current in vascular smooth muscle cells.

Animals↗
PubMed1995

Immunological and physical characterization of the brain G protein-gated muscarinic potassium channel.

The subunit composition of the G protein-gated inwardly rectifying K+ (KG) channel protein extracted from mouse forebrain membranes was examined. A polyclonal antibody (anti-GIRK1C1) was prepared against the carboxyl terminal region of GIRK1, the major subunit of the KG channel. The anti-GIRK1C1 IgG detected a single protein at approximately 65 kDa in SDS-PAGe of brain membranes. This IgG immunoprecipitated a macroprotein complex composed of GIRK1 and several associated proteins whose molecular weights ranging from 50 to 62 kDa on SDS-polyacrylamide gel. Upon size fractionation by both sucrose density gradient centrifugation and gel filtration, the solubilized KG channel proteins migrated into a single peak, which suggests that the component subunits form a single macromolecule. On the basis of the values of size fractionation, the molecular mass of the KG channel macromolecule could be estimated at approximately 231,000. These results suggest that the KG channel is most likely a tetrameric macroprotein composed of GIRK1 and co-immunoprecipitated proteins in the forebrain.

Amino Acid Sequence↗
PubMed1995

Spermine gates inward-rectifying muscarinic but not ATP-sensitive K+ channels in rabbit atrial myocytes. Intracellular substance-mediated mechanism of inward rectification.

The effect of spermine, a low molecular mass aliphatic amine with positive charges, on the strongly inwardly rectifying muscarinic K+ (KACh) channel was examined in rabbit atrial myocytes. In inside-out patch membranes, the single channel current-voltage relationship of KACh channels activated by guanosine 5'-3-O-(thio)triphosphate became linear in the absence of intracellular Mg2+. The open probability (Po) of the channels did not show significant voltage dependence under these conditions. Spermine specifically reduced Po of outwardly flowing KACh channel currents without affecting the unitary current amplitude at depolarized potentials, but had no effect on inward KACh currents under hyperpolarization. This voltage dependence of Po of KACh channels in the presence of spermine resembled that normally observed in the whole cell or open cell-attached configurations. Spermine (300 nM to 3 microM) also restored the relaxation of KACh currents which had been lost in the inside-out configuration. The effect of spermine was concentration-dependent with IC50 of approximately 10 nM at +40 mV. The order of potency of polyamines in reducing Po at +40 mV was spermine > or = spermidine > putrescine > ornithine; arginine had no significant effect. Intracellular Mg2+ antagonized the effect of spermine. Neither the single channel conductance nor Po of the ATP-sensitive K+ channel, a weak inward rectifier, was affected by spermine. Because submillimolar concentrations of spermine and spermidine are available in the cytosol of most cells, these substances may be the unidentified intracellular gating factors for strong inward rectifiers such as KACh and IK1 channels.

Adenosine Triphosphate↗
PubMed1995

Ionic basis of neurokinin-A-induced depolarization in single smooth muscle cells isolated from guinea-pig trachea.

Neurokinin A (NKA) caused single tracheal smooth muscle cells (TSMCs) to contract. The effects of NKA on the electrical activity of guinea-pig TSMCs were examined using the tight-seal whole-cell patch-clamp technique. Under current-clamp conditions at rest, the membrane potential of TSMCs spontaneously oscillated at about -40 mV and NKA rapidly depolarized the membrane potential to nearly 0 mV, which then gradually repolarized to about -20 mV in the presence of NKA. The oscillations in potential disappeared transiently during the rapid phase of depolarization in response to NKA and reappeared during the sustained phase of depolarization. Under voltage-clamp conditions, NKA evoked an inward current which faded quickly. Subsequently, the cell conductance in the presence of NKA at potentials greater than -40 mV decreased gradually. The reversal potential of the NKA-induced inward current was about 0 mV, and shifted with changes in the Cl- equilibrium potential. The Cl- current was not elicited by NKA when using a pipette solution containing 10 mM ethylenebis(oxonitrilo)tetraacetic acid (EGTA). During the sustained phase, K+ currents evoked by depolarizing voltage steps were inhibited by NKA. The present results indicate that NKA causes rapid and sustained depolarization of TSMCs by two distinct mechanisms: (1) initial transient activation of the Ca(2+)-dependent Cl- current, and (2) sustained inhibition of K+ currents.

Animals↗
PubMed1995

Cardiac ATP-sensitive K+ channels: regulation by intracellular nucleotides and K+ channel-opening drugs.

ATP-sensitive K+ (KATP) channels are present at high density in membranes of cardiac cells where they regulate cardiac function during cellular metabolic impairment. KATP channels have been implicated in the shortening of the action potential duration and the cellular loss of K+ that occurs during metabolic inhibition. KATP channels have been associated with the cardioprotective mechanism of ischemia-related preconditioning. Intracellular ATP (ATPi) is the main regulator of KATP channels. ATPi has two functions: 1) to close the channel (ligand function) and 2) in the presence of Mg2+, to maintain the activity of KATP channels (presumably through an enzymatic reaction). KATP channel activity is modulated by intracellular nucleoside diphosphates that antagonize the ATPi-induced inhibition of channel opening or induce KATP channels to open. How nucleotides will affect KATP channels depends on the state of the channel. K+ channel-opening drugs are pharmacological agents that enhance KATP channel activity through different mechanisms and have great potential in the management of cardiovascular conditions. KATP channel activity is also modulated by neurohormones. Adenosine, through the activation of a GTP-binding protein, antagonizes the ATPi-induced channel closure. Understanding the molecular mechanisms that underlie KATP channel regulation should prove essential to further define the function of KATP channels and to elucidate the pharmacological regulation of this channel protein. Since the molecular structure of the KATP channel has now become available, it is anticipated that major progress in the KATP channel field will be achieved.

Adenosine Triphosphate↗
PubMed1995

G protein regulation of cardiac muscarinic potassium channel.

Several ion channels can be regulated by G proteins in a "membrane-delimited" manner. The cardiac muscarinic K+ (KACh) channel, which is responsible for the acetylcholine (ACh) or adenosine-induced deceleration of heart beat and atrioventricular conduction, is the prototype of this type of receptor-dependent regulation of ion channels. Because similar transduction mechanisms are utilized by various membrane receptors, such as somatostatin, 5-hydroxytryptamine-1, alpha 2-adrenergic, mu-and delta-opioid, D2-dopamine, and gamma-aminobutyric acid B receptors, in neuronal, hormone-secreting, renal, or smooth muscle cells, the G protein (GK)-KACh channel system illustrates the principles underlying one of the most important cell signaling mechanisms (B. Hille. Neuron 9: 187-195, 1992). It seems that both alpha- and beta gamma-subunits of GK may be involved in the regulation of the KACh channel of mammalian atrial muscle. A general consensus of opinion has emerged, after some years of controversy, to support the notion that physiological activation of the channel by GK is the responsibility of the beta gamma-subunits. Recent evidence suggests that the KACh channel interacts with the alpha-subunit in the terminating process of activation.

Aged↗
PubMed1995

Mechanisms of adenosine-mediated actions on cellular and clinical cardiac electrophysiology.

OBJECTIVE: To provide insights into the molecular mechanisms of adenosine-mediated cardiac cellular electrophysiology and how information about these mechanisms can be used to facilitate diagnostic and therapeutic approaches to various clinical arrhythmias. DESIGN: A review of (1) adenosine metabolism and receptors in the cardiac system, (2) adenosine-mediated signal transduction pathways in the regulation of cellular electrophysiology in various cardiac cell types, and (3) the clinical usefulness of adenosine in cardiac electrophysiology is presented. RESULTS: The effects of adenosine on cardiac electrophysiologic properties are consequences of complex interactions among the specific cardiac target structures, the density and type of adenosine receptors, and the effector systems. The easy application of adenosine and its short half-life, favorable side-effects profile, and electrophysiologic properties make it an excellent diagnostic and therapeutic tool for the initial assessment of various tachyarrhythmias. CONCLUSION: The direct adenosine-activated KACh (potassium acetylcholine) channel signal transduction system explains the effects of adenosine on the sinus node, atrioventricular node, and atrial myocardium. The indirect adenosine-inhibited adenylate cyclase system accounts for its negative inotropic effects on the catecholamine-entrained contractility in atrial and ventricular myocardium. Because of the recent purification and cloning of adenosine receptors and subunits of G proteins, additional adenosine-mediated electrophysiologic mechanisms can be explored.

Adenosine↗
PubMed1995

Flecainide inhibits the transient outward current in atrial myocytes isolated from the rabbit heart.

We examined the effects of flecainide, a class Ic antiarrhythmic agent, on membrane currents in single rabbit atrial myocytes, using the tight-seal whole cell voltage-clamp technique. Under the current-clamp condition, flecainide (1-100 microM) prolonged the action potential duration at both the early and the late phases of repolarization in a concentration-dependent manner without affecting the resting membrane potential. In the presence of 4-aminopyridine, however, the drug affected the atrial action potential duration differently than it did in the absence of 4-aminopyridine: it shortened the early phase and only slightly lengthened the late phase of the atrial action potential. Under the voltage-clamp condition, flecainide suppressed the 4-amino-pyridine-sensitive, Ca(++)-insensitive transient outward current in a concentration-dependent fashion (the concentration for the half-maximal effect was approximately 17 microM). The drug also slightly inhibited the voltage-dependent L-type Ca++ current and delayed outward K+ current. Flecainide apparently accelerated the inactivation time course of the transient outward current but did not affect the voltage-dependence of its steady-state inactivation. These actions of flecainide on the transient outward current could be described by a voltage-dependent first-order interaction of the drug with the channel.

4-Aminopyridine↗