The positive inotropic dihydropyridine BAY K 8644 does not affect calium sensitivity or calcium release of skinned cardiac fibres

1985 ◽  
Vol 328 (4) ◽  
pp. 378-381 ◽  
Author(s):  
G�nter Thomas ◽  
Rainer Gro� ◽  
G. Pfitzer ◽  
J. C. R�egg
Keyword(s):  
Calcium Release ◽  
Bay K 8644 ◽  
Skinned Cardiac Fibres

scholarly journals Contractions of dysgenic skeletal muscle triggered by a potentiated, endogenous calcium current.

1991 ◽  
Vol 97 (4) ◽  
pp. 687-696 ◽  
Author(s):  
B A Adams ◽  
K G Beam
Keyword(s):  
Skeletal Muscle ◽  
Electrical Stimulation ◽  
Sarcoplasmic Reticulum ◽  
Calcium Current ◽  
Calcium Release ◽  
Voltage Sensor ◽  
Bay K 8644 ◽  
Direct Electrical Stimulation ◽  
External Calcium ◽  
Muscular Dysgenesis

The dihydropyridine (DHP) receptor of normal skeletal muscle is hypothesized to function as the voltage sensor for excitation-contraction (E-C) coupling, and also as the calcium channel underlying a slowly activating, DHP-sensitive current (termed ICa-s). Skeletal muscle from mice with muscular dysgenesis lacks both E-C coupling and ICa-s. However, dysgenic skeletal muscle does express a small DHP-sensitive calcium current (termed ICa-dvs) which is kinetically and pharmacologically distinct from ICa-s. We have examined the ability of ICa-dys, or the DHP receptor underlying it, to couple depolarization and contraction. Under most conditions ICa-dys is small (approximately 1 pA/pF) and dysgenic myotubes do not contract in response to sarcolemmal depolarization. However, in the combined presence of the DHP agonist Bay K 8644 (1 microM) and elevated external calcium (10 mM), ICa-dys is strongly potentiated and some dysgenic myotubes contract in response to direct electrical stimulation. These contractions are blocked by removing external calcium, by adding 0.5 mM cadmium to the bath, or by replacing Bay K 8644 with the DHP antagonist (+)-PN 200-110. Only myotubes having a density of ICa-dys greater than approximately 4 pA/pF produce detectible contractions, and the strength of contraction is positively correlated with the density of ICa-dys. Thus, unlike the contractions of normal myotubes, the contractions of dysgenic myotubes require calcium entry. These results demonstrate that the DHP receptor underlying ICa-dys is unable to function as a "voltage sensor" that directly couples membrane depolarization to calcium release from the sarcoplasmic reticulum.

scholarly journals Unitary behavior of skeletal, cardiac, and chimeric L-type Ca2+ channels expressed in dysgenic myotubes.

1996 ◽  
Vol 107 (6) ◽  
pp. 731-742 ◽  
Author(s):  
R T Dirksen ◽  
K G Beam
Keyword(s):  
Calcium Release ◽  
Dihydropyridine Receptor ◽  
Bay K 8644 ◽  
Dihydropyridine Receptors ◽  
Internal Repeat ◽  
Open Time ◽  
Activation Rate ◽  
Voltage Dependent ◽  
The Difference ◽  
Intracellular Loops

Skeletal and cardiac dihydropyridine receptors function both as voltage-dependent L-type calcium channels (L-channels) and as critical proteins that trigger calcium release from the sarcoplasmic reticulum in muscle. In spite of these similarities, skeletal L-channels exhibit a markedly slower activation rate than cardiac L-channels. We investigated the mechanisms underlying this difference by comparing the unitary behavior of L-channels in cell-attached patches of dysgenic myotubes expressing skeletal, cardiac, or chimeric dihydropyridine receptors. Our results demonstrate that ensemble averages activate rapidly for the purely cardiac dihydropyridine receptor and approximately five times more slowly for L-channels attributable to the purely skeletal dihydropyridine receptor or a chimeric dihydropyridine receptor in which only the first internal repeat and all of the putative intracellular loops are of skeletal origin. All of the constructs studied similarly exhibit a brief (2-ms) and a long (> or = 15-ms) open time in the presence of Bay K 8644, neither of which depend significantly on voltage. In the absence of Bay K 8644, the fraction of total open events is markedly shifted to the briefer open time without altering the rate of ensemble activation. Closed time analysis of L-channels with cardiac-like, rapid activation (recorded in the presence of dihydropyridine agonist) reveals both a brief (approximately 1-ms) closed time and a second, voltage-dependent, long-lasting closed time. The time until first opening after depolarization is three to six times faster for rapidly activating L-channels than for slowly activating L-channels and depends strongly on voltage for both types of channels. The results suggest that a voltage-dependent, closed-closed transition that is fast in cardiac L-channels and slow in skeletal L-channels can account for the difference in activation rate between these two channels.

Membrane Changes in Polymorphonuclear Leukocytes During Ionophore (A23187) - Induced Lysosomal Release

1977 ◽  
Vol 35 ◽  
pp. 482-483
Author(s):  
P.L. Moore ◽  
P.L. Sannes ◽  
H.L. Bank ◽  
S.S. Spicer
Keyword(s):  
Structural Changes ◽  
Calcium Release ◽  
Clear Understanding ◽  
Ionophore A23187 ◽  
Enzyme Release ◽  
Extracellular Medium ◽  
Pmn Leukocytes ◽  
Intracellular Ion Concentrations ◽  
Membrane Changes ◽  
Pmn Leukocyte

It is thought that calcium and/or magnesium may play important roles in polymorphonuclear (PMN) leukocyte functions such as chemotaxis, adhesion and phagocytosis. Yet, a clear understanding of the biological roles of these ions has awaited the development of techniques which permit a selective alteration of intracellular ion concentrations. Recently, treatment of cells with the ionophore A23187 has been used to alter intracellular divalent cation concentrations. This ionophore is a lipid soluble antibiotic produced by Streptomyces chartreusensis that complexes with both calcium and magnesium (3) and is believed to carry these ions across biological membranes (4). Biochemical investigations of human PMN leukocytes demonstrate that cells treated with A23187 and extracellular calcium release their lysosomal enzymes into the extracellular medium without rupturing and releasing their soluble cytoplasmic enzymes (5,6). The aim of the present study and and a companion report (7) was to investigate the structural changes that occur in leukocytes during ionophore-induced lysosomal enzyme release.

Inhibition of dihydropyridine-sensitive calcium entry in hypoxic relaxation of airway smooth muscle

1995 ◽  
Vol 268 (2) ◽  
pp. L201-L206 ◽  
Author(s):  
C. Vannier ◽  
T. L. Croxton ◽  
L. S. Farley ◽  
C. A. Hirshman
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Muscle Tone ◽  
Bay K 8644 ◽  
O2 Concentration ◽  
Voltage Dependent ◽  
Progressive Hypoxia ◽  
Carbamyl Choline

Hypoxia dilates airways in vivo and reduces active tension of airway smooth muscle in vitro. To determine whether hypoxia impairs Ca2+ entry through voltage-dependent channels (VDC), we tested the ability of dihydropyridines to modulate hypoxia-induced relaxation of KCl- and carbamyl choline (carbachol)-contracted porcine bronchi. Carbachol- or KCl-contracted bronchial rings were exposed to progressive hypoxia in the presence or absence of 1 microM BAY K 8644 (an L-type-channel agonist). In separate experiments, rings were contracted with carbachol or KCl, treated with nifedipine (a VDC antagonist), and finally exposed to hypoxia. BAY K 8644 prevented hypoxia-induced relaxation in KCl-contracted bronchi. Nifedipine (10(-5) M) totally relaxed KCl- contracted bronchi. Carbachol-contracted bronchi were only partially relaxed by nifedipine but were completely relaxed when the O2 concentration of the gas was reduced from 95 to 0%. These data indicate that hypoxia can reduce airway smooth muscle tone by limiting entry of Ca2+ through a dihydropyridine-sensitive pathway, but that other mechanisms also contribute to hypoxia-induced relaxation of carbachol-contracted bronchi.

scholarly journals A50 MODULATION OF GOBLET CELL ACTIVITY DURING GIARDIA DUODENALIS INFECTION: A ROLE FOR PAR2

2021 ◽  
Vol 4 (Supplement_1) ◽  
pp. 6-7
Author(s):  
E Fekete ◽  
C B Amat ◽  
T Allain ◽  
M Hollenberg ◽  
K Mihara ◽  
...  
Keyword(s):  
Gene Expression ◽  
Goblet Cell ◽  
Cysteine Protease ◽  
Protease Activity ◽  
Calcium Release ◽  
Cell Activity ◽  
Goblet Cells ◽  
Giardia Infection ◽  
Mucus Production ◽  
Mucin Gene

Abstract Background Giardia duodenalis has been shown to alter the structure of the intestinal mucus layers during infection via obscure mechanisms. We hypothesize that goblet cell activity may be disrupted in part due to proteolytic activation of protease-activated receptor 2 (PAR2) by Giardia proteases, resulting in disruption of mucus production and secretion by intestinal goblet cells. Aims Characterize alterations in goblet cell activity during Giardia infection, focusing on the roles of Giardia protease activity and PAR2. Methods Chinese hamster ovary cells transfected with nano-luciferase tagged PAR2 were incubated with Giardia NF or GSM trophozoites. Cleavage within the activation domain results in release of enzymes into the supernatant. Luminescence in the supernatant was measured as an indication of PAR cleavage by Giardia. LS174T, a human colonic mucus-producing cell line, was infected with Giardia trophozoites (isolates NF, WB, S2, and GSM). Prior to infection, trophozoites were treated with E64, a broad-spectrum cysteine protease inhibitor, and LS174T were treated with a PAR2 antagonist, a calcium chelator, or an ERK1/2 inhibitor. Quantitative PCR (qPCR) was performed for the MUC2 mucin gene. Wild-type (WT) and PAR2 knockout (KO) mice were infected with Giardia. Colonic mucus was stained using fluorescein-coupled wheat-germ agglutinin (WGA), and qPCR was performed for Muc2 and Muc5ac. Results Giardia trophozoites cleaved PAR2 within the N-terminal activation domain in a cysteine protease-dependent manner. Cleavage was isolate dependent, with isolates that show higher protease activity cleaving at a higher rate. High protease activity Giardia isolates increased MUC2 gene expression in LS714T. This increase was attenuated by inhibition of Giardia cysteine protease activity, and by antagonism of PAR2, inhibition of calcium release, or inhibition of ERK1/2 activity in LS174T cells. Both Muc2 and Muc5ac expression were upregulated in the colons of WT mice in response to Giardia infection, while in the jejunum Muc2 expression decreased and Muc5ac expression increased. In KO, no changes in gene expression were seen in the colon in response to Giardia infection, while in the jejunum, Muc2 expression was unchanged and Muc5ac expression decreased. Both WT infected and KO noninfected mice showed thinning of the colonic mucus layer compared to WT controls. There was some recovery in thickness in KO infected mice. Conclusions PAR2 plays a significant role in the regulation of mucin gene expression in mice and in a human colonic cell line. Results suggest that Giardia cysteine proteases cleave and activate PAR2, leading to calcium release and activation of the MAPK pathway in goblet cells, ultimately leading to altered mucin gene expression. Findings identify a novel regulatory pathway for mucus production by intestinal goblet cells. Funding Agencies CAG, CCC

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