scholarly journals Calcium Channels and Nifedipine Inhibition of Serotonin-Induced [3H]Thymidine Incorporation in Cultured Cerebral Smooth Muscle Cells

1992 ◽  
Vol 12 (1) ◽  
pp. 139-146 ◽  
Author(s):  
Thomas A. Kent ◽  
Allahyar Jazayeri ◽  
J. Marc Simard
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Thymidine Incorporation ◽  
Membrane Surface ◽  
Muscle Cells ◽  
Thymidine Uptake ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent ◽  
Alternate Mechanism

Cultures of smooth muscle cells were prepared from the basilar artery of adult guinea pigs. Passaged cultures (10–30 passages) that expressed serotonin receptors were studied using [3H]thymidine incorporation. When tested in quiescent medium, serotonin potently stimulated [3H]thymidine incorporation (EC50 of 31 n M) by as much as 400% at 24 h. The number of cells was not significantly increased at 24 or 48 h. At concentrations of 10−8–10−5 M 5-HT, [3H]thymidine uptake was reduced 40–50% by the dihydropyridine Ca2+ channel blocker, nifedipine (1 μ M). To demonstrate a possible mechanism for the sensitivity to nifedipine, Ca2+ currents were measured using the whole cell patch clamp technique. The cells expressed dihydropyridine-sensitive L-type Ca2+ channels, but not other subtypes of Ca2+ channels, as indicated by the kinetic and voltage-dependent characteristics of the current and by the stimulatory effect of Bay K 8644. The magnitude of the Ca2+ currents was related exponentially to the membrane surface area, measured as cell capacitance. These data support the association of dihydropyridine-sensitive Ca2+ channels with mitogenesis in vascular smooth muscle, and suggest an alternate mechanism of action for the beneficial effect of dihydropyridines in prophylaxis of cerebral vasospasm.

Dual effect of phosphatase inhibitors on calcium channels in intestinal smooth muscle cells

1993 ◽  
Vol 264 (2) ◽  
pp. C296-C301 ◽  
Author(s):  
K. Obara ◽  
H. Yabu
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Phosphatase Inhibitors ◽  
Inward Current ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent ◽  
High Concentrations

The effects of okadaic acid (OA) and calyculin A (CL-A), potent inhibitors of protein phosphatases type 1 (PP1) and type 2A (PP2A), on inward current carried by Ba2+ through voltage-dependent Ca2+ channel in guinea pig teniae coli smooth muscle cells were investigated using whole-cell patch-clamp technique. High concentrations of OA (5 x 10(-8)-5 x 10(-6) M) and CL-A (10(-9)-10(-7) M) dose dependently increased the inward current. The concentration producing apparent half-maximum enhancing effect values for OA and CL-A were 1.12 x 10(-7) and 1.78 x 10(-9) M, respectively. CL-A appeared to be approximately 100-fold more potent in increasing the inward current than OA. Lower concentrations of OA (10(-10)-2 x 10(-8) M) and CL-A (10(-11)-10(-9) M) decreased the inward current. The maximum inhibitory effects of OA and CL-A were observed at 10(-8) M OA and 5 x 10(-10) M CL-A, respectively. CL-A is approximately 100 times more effective inhibitor of PP1 than OA, and lower concentrations of OA and CL-A used in the present study inhibit PP2A activity, but they have no or little effect on PP1 activity (Ishihara, H., B. L. Martin, D. L. Brautigan, H. Karaki, H. Ozaki, Y. Kato, N. Fusetani, S. Watabe, K. Hashimoto, D. Uemura and D. J. Hartshorne. Biochem. Biophys. Res. Commun. 159: 871-877, 1989). In the absence of ATP in pipette solution, OA and CL-A did not affect the inward current.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin 6 stimulates growth of vascular smooth muscle cells in a PDGF-dependent manner

1991 ◽  
Vol 260 (5) ◽  
pp. H1713-H1717 ◽  
Author(s):  
U. Ikeda ◽  
M. Ikeda ◽  
T. Oohara ◽  
A. Oguchi ◽  
T. Kamitani ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Interleukin 6 ◽  
Thymidine Incorporation ◽  
Tritiated Thymidine ◽  
Muscle Cells ◽  
Thymidine Uptake ◽  
Dependent Manner

We have investigated the effect of interleukin 6 (IL-6) on the growth of vascular smooth muscle cells (VSMC) isolated from rat aortas. Murine recombinant IL-6 significantly increased the number of VSMC and stimulated tritiated thymidine incorporation into VSMC in a dose-dependent manner. The IL-6-induced thymidine incorporation into VSMC was totally inhibited by the Ca2+ channel blocker verapamil; however, IL-6 showed no effects on the intracellular Ca2+ level ([Ca2+]i) in VSMC. Antibody against platelet-derived growth factor (PDGF) also totally inhibited the IL-6-induced thymidine uptake. PDGF caused a significant increase in the [Ca2+]i, which was totally inhibited by verapamil. IL-6 mRNA was not detected in unstimulated “quiescent” VSMC, but its expression was stimulated by exposure of VSMC to 10% fetal bovine serum. Immunohistochemical study using anti-PDGF antibody showed that IL-6 stimulated PDGF production in VSMC. These results support the premise that IL-6 is released by VSMC in an autocrine manner and promotes the growth of VSMC via induction of endogenous PDGF production.

Voltage-dependent calcium channel current in isolated gallbladder smooth muscle cells of guinea pig

1993 ◽  
Vol 264 (6) ◽  
pp. G1066-G1076 ◽  
Author(s):  
T. Shimada
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Time Course ◽  
Reversal Potential ◽  
High Sensitivity ◽  
Muscle Cells ◽  
Patch Clamp Technique ◽  
Decay Component ◽  
Voltage Dependent

The voltage-dependent Ca2+ current was studied in enzymatically dispersed guinea pig gallbladder smooth muscle cells using the whole cell patch-clamp technique. Depolarizing voltage (V) steps induced an inward current (I) that was carried by Ca2+. The threshold potential was -40 to -30 mV, the maximal current was observed at +10 to +20 mV, and the reversal potential was around +80 mV. I-V curves obtained with holding potentials of -80 and -40 mV were not significantly different. This current had a high sensitivity to dihydropyridine drugs, and the Ba2+ or Sr2+ current was larger than the Ca2+ current. Activation was accelerated by increasing the membrane potential. In general, the time course of decay was well fitted by the sum of two exponentials, but consideration of a third (ultra-slow) decay component was also necessary when the current generated by a 2-s command pulse was analyzed. Superimposition of activation and inactivation curves showed the presence of a significant window current. Carbachol suppressed the Ca2+ current only when the pipette contained a low concentration of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. These results show that the L-type Ca2+ current is dominant in gallbladder smooth muscle cells and may contribute to excitation-contraction coupling.

Fatty acid modifies Ca 2+ -dependent potassium channel activity in smooth muscle cells from the human aorta

1989 ◽  
Vol 237 (1288) ◽  
pp. 259-266 ◽  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Single Channel ◽  
Membrane Surface ◽  
Muscle Cells ◽  
Human Aorta ◽  
Channel Activity ◽  
Patch Clamp Technique ◽  
Membrane Fluidization ◽  
Potassium Channel Activity

By using the patch–clamp technique the effect of 2-decenoic acid (DA) on Ca 2+ -activated potassium (K + ) channels in the membrane of smooth muscle cells from the human aorta was studied. In the presence of 0.5 μM Ca 2+ and 2 mM Mg 2+ on the cytoplasmic side of the membrane, a more than tenfold elevation in the probability of the channels being open ( p o ) was observed under the effect of DA. With divalent cation concentrations of less than 1 nM DA caused a more than twofold elevation in p o . In the DA-treated membranes Mg 2+ ions, which normally fail to activate the channels, brought about a nearly threefold increase in the channel activity when applied to the inner membrane surface. Channel sensitivity to the activating effect of cytoplasmic Ca 2+ ions did not increase with the application of DA. Single-channel conductance was unchanged by DA exposure. We suggest that DA alters the Ca 2+ -binding mechanism of the channel, increasing its sensitivity to Mg 2+ ions, presumably owing to membrane fluidization.

scholarly journals Arecoline Hydrobromide Enhances Jejunum Smooth Muscle Contractility via Voltage-Dependent Potassium Channels in W/Wv Mice

2021 ◽  
pp. 437-446
Author(s):  
Q CHEN ◽  
Z JIANG ◽  
J ZHANG ◽  
L CAO ◽  
Z CHEN
Keyword(s):  
Smooth Muscle ◽  
Potassium Channels ◽  
Smooth Muscle Cells ◽  
Isometric Force ◽  
Muscle Cells ◽  
Resting Membrane Potential ◽  
Dependent Manner ◽  
Excitatory Effect ◽  
Patch Clamp Technique ◽  
Voltage Dependent

Gastrointestinal motility was disturbed in W/Wv, which were lacking of interstitial cells of Cajal (ICC). In this study, we have investigated the role of arecoline hydrobromide (AH) on smooth muscle motility in the jejunum of W/Wv and wild-type (WT) mice. The jejunum tension was recorded by an isometric force transducer. Intracellular recording was used to identify whether AH affects slow wave and resting membrane potential (RMP) in vitro. The whole-cell patch clamp technique was used to explore the effects of AH on voltage-dependent potassium channels for jejunum smooth muscle cells. AH enhanced W/Wv and WT jejunum contractility in a dose-dependent manner. Atropine and nicardipine completely blocked the excitatory effect of AH in both W/Wv and WT. TEA did not reduce the effect of AH in WT, but was sufficient to block the excitatory effect of AH in W/Wv. AH significantly depolarized the RMP of jejunum cells in W/Wv and WT. After pretreatment with TEA, the RMP of jejunum cells indicated depolarization in W/Wv and WT, but subsequently perfused AH had no additional effect on RMP. AH inhibited the voltage-dependent K+ currents of acutely isolated mouse jejunum smooth muscle cells. Our study demonstrate that AH enhances the contraction activity of jejunum smooth muscle, an effect which is mediated by voltage-dependent potassium channels that acts to enhance the excitability of jejunum smooth muscle cells in mice.

Multiple transcripts of Ca2+ channel α1-subunits and a novel spliced variant of the α1C-subunit in rat ductus arteriosus

2006 ◽  
Vol 290 (4) ◽  
pp. H1660-H1670 ◽  
Author(s):  
Utako Yokoyama ◽  
Susumu Minamisawa ◽  
Satomi Adachi-Akahane ◽  
Toru Akaike ◽  
Isao Naguro ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Channel Blocker ◽  
Thymidine Incorporation ◽  
Ductus Arteriosus ◽  
Muscle Cells ◽  
Wistar Rat ◽  
Multiple Transcripts ◽  
Voltage Dependent ◽  
Spliced Variant

Voltage-dependent Ca2+ channels (VDCCs), which consist of multiple subtypes, regulate vascular tone in developing arterial smooth muscle, including the ductus arteriosus (DA). First, we examined the expression of VDCC subunits in the Wistar rat DA during development. Among α1-subunits, α1C and α1G were the most predominant isoforms. Maternal administration of vitamin A significantly increased α1C- and α1G-transcripts. Second, we examined the effect of VDCC subunits on proliferation of DA smooth muscle cells. We found that 1 μM nitrendipine (an L-type Ca2+ channel blocker) and kurtoxin (a T-type Ca2+ channel blocker) significantly decreased [3H]thymidine incorporation and that 3 μM efonidipine (an L- and T-type Ca2+ channel blocker) further decreased [3H]thymidine incorporation, suggesting that L- and T-type Ca2+ channels are involved in smooth muscle cell proliferation in the DA. Third, we found that a novel alternatively spliced variant of the α1C-isoform was highly expressed in the neointimal cushion of the DA, where proliferating and migrating smooth muscle cells are abundant. The basic channel properties of the spliced variant did not differ from those of the conventional α1C-subunit. We conclude that multiple VDCC subunits were identified in the DA, and, in particular, α1C- and α1G-subunits were predominant in the DA. A novel spliced variant of the α1C-subunit gene may play a distinct role in neointimal cushion formation in the DA.

Volatile anesthetics inhibit voltage-dependent Ca2+ channels in porcine tracheal smooth muscle cells

1995 ◽  
Vol 268 (2) ◽  
pp. L187-L191 ◽  
Author(s):  
M. Yamakage ◽  
C. A. Hirshman ◽  
T. L. Croxton
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Volatile Anesthetics ◽  
Inhibitory Effect ◽  
Tracheal Smooth Muscle ◽  
Lipid Phase ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp

The relaxation of airway smooth muscle by volatile anesthetics is associated with a decreased concentration of intracellular free Ca2+. We hypothesized that inhibition of the entry of extracellular Ca2+ contributes to the relaxation. We therefore examined the effects of halothane, isoflurane, and sevoflurane on macroscopic voltage-activated Ca2+ currents (ICa) in porcine tracheal smooth muscle cells, using the whole cell patch-clamp technique. All three volatile anesthetics significantly inhibited ICa in a dose-dependent manner with no apparent shift in the voltage dependence of induced ICa. The order of inhibitory potencies for ICa was halothane < isoflurane < sevoflurane. When data were plotted as a function of the estimated anesthetic concentrations in the lipid phase, the potencies for inhibition of ICa by the three anesthetics were indistinguishable. We conclude that volatile anesthetics have an inhibitory effect on ICa of porcine tracheal smooth muscle cells at clinically relevant concentrations and that the inhibitory potencies of volatile anesthetics on ICa are closely related to their lipid-phase solubilities.

IGFBP-3 activates TGF-β receptors and directly inhibits growth in human intestinal smooth muscle cells

2004 ◽  
Vol 287 (4) ◽  
pp. G795-G802 ◽  
Author(s):  
John F. Kuemmerle ◽  
Karnam S. Murthy ◽  
Jennifer G. Bowers
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Nuclear Translocation ◽  
Thymidine Incorporation ◽  
Muscle Cells ◽  
Serine Phosphorylation ◽  
Intestinal Smooth Muscle ◽  
Igf I ◽  
Tgf Β1 ◽  
Igfbp 3

We have shown that human intestinal smooth muscle cells produce IGF-I and IGF binding protein-3 (IGFBP-3). Endogenous IGF-I acts in autocrine fashion to stimulate growth of these cells. IGFBP-3 inhibits the binding of IGF-I to its receptor and thereby inhibits IGF-I-stimulated growth. In several carcinoma cell lines and some normal cells, IGFBP-3 regulates growth independently of IGF-I. Two mechanisms for this effect have been identified: IGFBP-3 can directly activate transforming growth factor-β (TGF-β) receptors or it can undergo direct nuclear translocation. The aim of the present study was to determine whether IGFBP-3 acts independently of IGF-I and to characterize the mechanisms mediating this effect in human intestinal smooth muscle cells. The direct effects of IGFBP-3 were determined in the presence of an IGF-I receptor antagonist to eliminate its IGF-I-dependent effects. Affinity labeling of TGF-β receptors (TGF-βRI, TGF-βRII, and TGF-βRV) with 125I-labeled TGF-β1 showed that IGFBP-3 displaced binding to TGF-βRII and TGF-βRV in a concentration-dependent fashion. IGFBP-3 stimulated TGF-βRII-dependent serine phosphorylation (activation) of both TGF-βRI and of its primary substrate, Smad2(Ser465/467). IGFBP-3 also caused IGF-I-independent inhibition of basal [3H]thymidine incorporation. The effects of IGFBP-3 on Smad2 phosphorylation and on smooth muscle cell proliferation were independent of TGF-β1 and were abolished by transfection of Smad2 siRNA. Immunoneutralization of IGFBP-3 increased basal [3H]thymidine incorporation, implying that endogenous IGFBP-3 inhibits proliferation. We conclude that endogenous IGFBP-3 directly inhibits proliferation of human intestinal smooth muscle cells by activation of TGF-βRI and Smad2, an effect that is independent of its effect on IGF-I-stimulated growth.

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