Recent papers of Zhong-Fang LAI

Recent papers in 2000

(This is a copy from Journal of Immunology 165(1):83-90; 2000)


An Amiloride-Sensitive and Voltage-Dependent Na+ Channel in an HLA-DR-Restricted Human T Cell Clone1
Zhong-Fang Lai,2,3* Yu-Zhen Chen,3˘÷ Yasuharu Nishimura,˘÷ and Katsuhide Nishi* *Department of Pharmacology, Kumamoto University School of Medicine, and ˘÷Division of Immunogenetics, Department of Neuroscience and Immunology, Kumamoto University Graduate School of Medical Sciences, Kumamoto, Japan
The Journal of Immunology, 2000, 165: 83-90.

We investigated changes in voltage-gated Na+ currents and effects of extracellular Na+ on proliferation in HLA-DR-restricted human CD4+ T cells after stimulation with a non-self antigenic peptide, M12p54?68. In the absence of antigenic peptide, neither single (n = 80) nor APC-contacted (n = 71) T cells showed voltage-gated inward currents recording with whole-cell patch-clamp techniques, even with Ca2+ and Na+ ions present in the perfusion solution. However, with the same recording conditions, 31% (26 of 84) of APC-contacted T cells stimulated with the antigenic peptide showed voltage-dependent inward currents that were elicited from -60 mV. The inward currents were not inhibited in extracellular Ca2+-free conditions or in the presence of 1 mM NiCl2. However, they were completely inhibited in extracellular Na+-free conditions, which were made by replacing Na+ with iso-osmotic N-methyl-D-glucamine or choline. The Na+ currents were insensitive to tetrodotoxin, a classical blocker of Na+ channels, but were dose-dependently inhibited by amiloride, a potassium-sparing pyrazine diuretic. Furthermore, the Ag-specific proliferative response of T cells was completely inhibited in Na+-free Tyrode's solution and was suppressed by amiloride in a dose-dependent manner. Our findings suggest that activation of amiloride-sensitive and voltage-gated Na+ channels would be an important step to allow an adequate influx of Na+ and maintain a sustained high Ca2+ level during T cell activation.

Recent papers in 1998

(This is a copy from Medline 1998)
Unique Identifier
99077145
Authors
Chen YZ. Lai ZF. Nishi K. Nishimura Y.
Institution
Department of Neuroscience and Immunology, Kumamoto University Graduate
School of Medical Sciences, Japan.
Title
Modulation of calcium responses by altered peptide ligands in a human T cell
clone.
Source
European Journal of Immunology. 28(12):3929-39, 1998 Dec.
Abstract
To determine whether altered peptide ligands (APL) affect calcium signaling events, we investigated changes in intracellular calcium concentration ([Ca2+]i) in human T cell clone stimulated with either the fully agonistic peptide M12p54-68, the partially agonistic analogue E63V or the simple antagonistic analogue E58M. Both E63V and E58M stimulated a Ca2+ response in approximately 40% of T cells, whereas M12p54-68 did so in approximately 70% of T cells. The most predominant pattern of a Ca2+ increase induced by M12p54-68 was a small sinusoidal peak followed by a sustained high response. The most frequent pattern of calcium response induced by E63V was a continuous high response without a preceding sinusoidal peak, whereas that induced by E58M was large with frequent oscillations. Genistein, an inhibitor of the protein tyrosine kinases (PTK), markedly inhibited the wild-type peptide-induced increase in [Ca2+]i, whereas it marginally inhibited the response induced by E63V or E58M. In contrast, GF109203X, a protein kinase C (PKC)-specific inhibitor, markedly inhibited the E63V- or E58M-induced Ca2+ response, whereas it marginally affected the wild peptide-induced Ca2+ response. Furthermore, in nominal Ca2+-free medium, the E58M-induced Ca2+ response was almost completely blocked, while the M12p54-68- or E63V-induced responses were only partially inhibited. Our results suggest that the Ca2+ response induced by the fully agonistic peptide depends on activation of the genistein-sensitive signaling pathway, including PTK, whereas the Ca2+ response to a simple antagonistic APL completely depends on extracellular Ca2+ and activation of the GF109203X-sensitive signaling pathway, including PKC. These differences in the CA2+i response in recognition of different APL may parallel the unique T cell activation patterns induced by APL in human T cells.

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Citation 2
Unique Identifier
99032718
Authors
Lai ZF. Nishi K.
Institution
Department of Pharmacology, Kumamoto University School of Medicine, Kumamoto
860-0811, Japan.
Title
Intracellular chloride activity increases in guinea pig ventricular muscle
during simulated ischemia.
Source
American Journal of Physiology. 275(5 Pt 2):H1613-9, 1998 Nov.
Abstract
We investigated the effects of simulated ischemia on intracellular Cl- activity ([Cl-]i) in isolated guinea pig ventricular papillary muscles using ion-selective microelectrode techniques. Simulated ischemia in ventricular muscles was produced by stopping the flow of superfusion and immersing preparations in mineral oil as previously described [B. Vanheel, L. Leybaert, A. De Hemptinne, and I. Leusen. Am. J. Physiol. 257 (Cell Physiol. 26): C365-C379, 1989; Z. F. Lai, J. Liu, and K. Nishi. Jpn. J. Pharmacol. 72: 161-174, 1996]. When preparations were exposed to paraffin oil for 15 min, [Cl-]i markedly increased and the peak magnitude of [Cl-]i reached 55.3 +/- 2.5 mM from 18.7 +/- 3.5 mM, whereas membrane potentials (Vm) depolarized from -82.5 +/- 1.1 to -54.7 +/- 2.4 mV (n = 6 muscles from 6 animals). SITS (0.5 mM), a known blocker of the Cl-/HCO-3 exchanger, suppressed the ischemia-induced depolarization of Vm and delayed the onset of the ischemia-induced increase in [Cl-]i but did not suppress the magnitude of the increase of [Cl-]i. Under Cl--free conditions created by replacing Cl- with equimolar gluconate, the increase in [Cl-]i during ischemia was transient and suppressed by >60% compared with that in normal-Cl- conditions (peak value was 20. 3 +/- 1.7 mM, n = 6 muscles from 6 animals). The present results provide direct evidence that [Cl-]i in ventricular muscle increases in ischemic conditions in quiescent guinea pig ventricular muscle, suggesting that activation of the Cl-/HCO-3 exchanger by ischemia would partially contribute to the elevation of [Cl-]i during the initial stage of ischemia.

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Citation 3
Unique Identifier
98213527
Authors
Liu J. Lai ZF. Wang XD. Tokutomi N. Nishi K.
Institution
Department of Pharmacology, Kumamoto University School of Medicine, Honjo,
Japan.
Title
Inhibition of sodium current by chloride channel blocker
4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) in guinea pig
cardiac ventricular cells.
Source
Journal of Cardiovascular Pharmacology. 31(4):558-67, 1998 Apr.
Abstract
The effects of 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), a potent anion transport blocker, on transmembrane action potentials (APs) and the sodium current (I[Na]) of guinea pig ventricular myocytes were examined by using conventional microelectrode and whole-cell patch-clamp recording techniques. In papillary muscle preparations, DIDS (> or =0.1 mM) suppressed the maximal upstroke velocity (.v[max]) of the AP without significant changes in other AP parameters. Extracellular application of DIDS on single cardiomyocytes isolated from the guinea pig ventricle markedly reduced the peak amplitude of the tetrodotoxin (TTX)-sensitive and voltage-activated sodium current. The concentration-dependent block of DIDS could be expressed by the Hill equation with a Hill coefficient of 0.97 and a dissociation constant of 0.15 mM at a holding potential of (VH) -120 mV. DIDS (0.1 mM) shifted the steady-state inactivation curve for I(Na) toward more negative potentials by 6.0 +/- 0.5 mV and the activation curve to more positive potentials by 5.0 +/- 1.0 mV, although the slope factors were unaffected. With repetitive depolarizing pulses from -120 mV, DIDS produced a use-dependent block on the I(Na). Recovery of I(Na) from inactivation was slowed (time constant = 245 ms, compared with 10 ms of control) in the presence of 0.1 mM DIDS. In the two-pulse experiments, DIDS produced two distinct phases of development of I(Na) block, the rapid phase (tau = 5 ms) caused by an open channel block, and the slower phase (tau = 382 ms) induced by an inactivated channel block. These results suggest that the Cl- transport blocker DIDS has a direct inhibitory effect on the cardiac sodium channel. DIDS-induced use dependence of I(Na) block may result from the interaction of the drug with sodium channels in both the open and inactivated channel states.

Recent papers in 1996

(This is a copy from current contents 1996)

1. Unique Identifier
98213527
Authors
Liu J Lai ZF,Wang XD, Tokutomi N, Nishi K.
Institution
Department of Pharmacology, Kumamoto University School of Medicine, Honjo,
Japan.
Title
Inhibition of sodium current by chloride channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) in guinea pig cardiac ventricular cells.
Source
Journal of Cardiovascular Pharmacology. 31(4):558-67, 1998 Apr.
Abstract
The effects of 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), a potent anion transport blocker, on transmembrane action potentials (APs) and the sodium current (I[Na]) of guinea pig ventricular myocytes were examined by using conventional microelectrode and whole-cell patch-clamp recording techniques. In papillary muscle preparations, DIDS (> or =0.1 mM) suppressed the maximal upstroke velocity (.v[max]) of the AP without significant changes in other AP parameters. Extracellular application of DIDS on single cardiomyocytes isolated from the guinea pig ventricle markedly reduced the peak amplitude of the tetrodotoxin (TTX)-sensitive and voltage-activated sodium current. The concentration-dependent block of DIDS could be expressed by the Hill equation with a Hill coefficient of 0.97 and a dissociation constant of 0.15 mM at a holding potential of (VH) -120 mV. DIDS (0.1 mM) shifted the steady-state inactivation curve for I(Na) toward more negative potentials by 6.0 +/- 0.5 mV and the activation curve to more positive potentials by 5.0 +/- 1.0 mV, although the slope factors were unaffected. With repetitive depolarizing pulses from -120 mV, DIDS produced a use-dependent block on the I(Na). Recovery of I(Na) from inactivation was slowed (time constant = 245 ms, compared with 10 ms of control) in the presence of 0.1 mM DIDS: In the two-pulse experiments, DIDS produced two distinct phases of development of I(Na) block, the rapid phase (tau = 5 ms) caused by an open channel block, and the slower phase (tau = 382 ms) induced by an inactivated channel block. These results suggest that the Cl- transport blocker DIDS has a direct inhibitory effect on the cardiac sodium channel. DIDS-induced use dependence of I(Na) block may result from the interaction of the drug with sodium channels in both the open and inactivated channel states.
2
UI - VT747
AU - Sato D
AU - Lai ZF
AU - Tokutomi N
AU - Tokutomi Y
AU - Maeda H
AU - Nishikawa S
AU - Nishikawa SI
AU - Ogawa M
AU - Nishi K
TI - Impairment of Kit-dependent development of interstitial cells alters contractile responses of murine intestinal tract.
LA - English
RF - Article
AD - K Nishi, Kumamoto Univ, Sch Med, Inst Mol Embryol & Genet, Dept Pharmacol, 2-2-1 Honjo, Kumamoto 860, Japan.

AB - We examined developmental changes in responses of the isolated segment of the ileum of BALB/c mice treated with a monoclonal antibody (ACK2) to the receptor tyrosine kinase (Kit) for 4 days postnatally to pharmacological agents in vitro. Rhythmic contraction and relaxation of the isolated ileum started to appear on day 4 postpartum, and the sensitivity to acetylcholine (ACh) decreased gradually after birth. Treatment with ACK2 induced augmentation of contractile responses and receptor sensitivity of the longitudinal muscle of the ileum to ACh, bradykinin, and prostaglandin F-2 alpha. ACh induced larger depolarization in smooth muscle cells of the ileum in the ACK2-treated mice than in the control. Circular muscle responses to these substances, as measured by changes in intraluminal pressure, were not altered by ACK2 treatment. Results suggest that interstitial cells play an important role not only in the development of the pacemaking system of the small intestine but also in the functional development of the contractile properties of the intestinal smooth muscle.
SO - Amer J Physiol-Gastrointest L 1996 NOV;34(5):G762-G771

3
UI - VN974
AU - Lai ZF
AU - Liu J
AU - Nishi K
TI - Effects of stilbene derivatives SITS and DIDS on development of intracellular acidosis during ischemia in isolated guinea pig ventricular papillary muscle in vitro.
LA - English
RF - Article
AD - K Nishi, Kumamoto Univ, Sch Med, Dept Pharmacol, 2-2-1 Honjo, Kumamoto 860, Japan.
AB - Using ion-selective microelectrode techniques, we investigated the effects of 4-acetamido-4'- isothiocyanatostilbene-2,2'-disulfonic acid (SITS) and 4, 4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), which are known as Cl--HCO3- exchange blockers, on action potentials and intracellular pH (pH(i)) in guinea pig ventricular papillary muscles subjected to simulated ischemia. Simulated ischemia was produced by stopping the flow of superfusing solution and then covering the preparations with mineral oil. Simulated ischemia induced a progressive decrease in the maximum upstroke rate and resting membrane potentials, shortened action potential duration, and resulted in cessation of action potentials within 10-12 min after the onset of simulated ischemia. The pH(i)-measurements revealed progressive intracellular acidosis during the period of simulated ischemia. SITS (0.5 mM) or DIDS (0.1 mM) delayed the onset of ischemia-induced deterioration of action potentials and prolonged the time to cessation of action potentials. SITS or DIDS (0.1-0.5 mM) induced an increase in pH(i) in HCO3--buffered solution and suppressed the development of intracellular acidosis during ischemia. Under the external Cl--free condition, the time to cessation of action potentials caused by ischemia was significantly delayed, and the development of intracellular acidosis during ischemia was attenuated. The present results indicate that activation of the Cl--HCO3- exchange system would be involved, in part, in the development of intracellular acidosis during cardiac ischemia.
SO - Jpn J Pharmacol 1996 OCT;72(2):161-174


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