scholarly journals Hydrogen Sulfide Regulates the [Ca2+]iLevel in the Primary Medullary Neurons

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Xiaoni Liu ◽  
Nana Zhang ◽  
Yingjiong Ding ◽  
Dongqing Cao ◽  
Ying Huang ◽  
...  
Keyword(s):  
Hydrogen Sulfide ◽  
Intracellular Calcium ◽  
Calcium Influx ◽  
Cultured Neurons ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Reversal Effect ◽  
Intracellular Ca ◽  
Medullary Neurons

In the present study, we attempted to elucidate mechanisms for the regulation of intracellular calcium levels by H2S in primary rat medullary neurons. Our results showed that NaHS significantly increased the level ofCa2+iin rat medullary neurons in a concentration-dependent manner. L-Cysteine and SAM significantly raised the level ofCa2+iin the medullary neurons while HA and/or AOAA produced a reversal effect. In addition, L-cysteine and SAM significantly increased but HA and/or AOAA decreased the production of H2S in the cultured neurons. TheCa2+ielevation induced by H2S was significantly diminished by EGTA-Ca2+-free solutions, and this elevation was also reduced by nifedipine or nimodipine and mibefradil, suggesting the role of L-type and/or T-type Ca2+channels. Moreover, the effect of H2S onCa2+ilevel in neurons was significantly attenuated by BAPTA-AM and thapsigargin, suggesting the source of Ca2+. Therefore, we concluded that both exogenous and endogenous H2S elevatesCa2+ilevel in primarily cultured rat medullary neurons via both increasing calcium influx and mobilizing intracellular Ca2+stores from ER.

The Role Of Calcium In Platelet Microtubule Assembly- Disassembly

1981 ◽  
Author(s):  
M Kikuchi ◽  
Y Ikeda ◽  
M Handa ◽  
S Matsuda ◽  
H Muraki ◽  
...  
Keyword(s):  
Intracellular Calcium ◽  
Dynamic Equilibrium ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Microtubule Assembly ◽  
Base Line ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Transient Decrease

Microtubules exist in a dynamic equilibrium between polymerized and depolymerized forms in human platelets, playing a major role to maintain the discoid shape of platelets. It has been previously shown that the interaction of aggregating agents with platelets leads to a rapid but transient disassembly of microtubules. ( Steiner and Ikeda, J.Clin. Invest. 63:443,1979 ) In this paper, the role of calcium in the equilibrium between assembled and disassembled microtubules was investigated. The respective pools of soluble and polymerized tubulin were “frozen” by addition of a glycerol-dimethyl sulfoxide-containing medium to platelet rich plasma, preincubated with 2 µM A23187 for various time intervals. The two pools of tubulin were estimated by measuring the colchicine binding activities of total and polymerized tubulin according to the method of Wilson.Resting platelets were found to contain 56.2 ± 2.7 µg tubulin per 109 platelets, of which 56.7 % was in polymerized form. Addition of A23187 to platelet rich plasma produced a transient decrease in the pool of polymerized tubulin within 30 sec., followed by a return to base-line values within 2 min.. TMB-8, a known intracellular calcium antagonist, abolished this transient decrease in polymerized tubulin induced by A23187 in a concentration dependent manner, while indomethacin or acetylsalycylic acid did not.These findings may indicate the important role of intracellular calcium in microtubule assembly-disassembly.

Modification of intracellular calcium concentration in cardiomyocytes by inhibition of sarcolemmal Na+/H+ exchanger

2006 ◽  
Vol 291 (6) ◽  
pp. H2790-H2800 ◽  
Author(s):  
Harjot K. Saini ◽  
Naranjan S. Dhalla
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Intracellular Calcium ◽  
Calcium Concentration ◽  
Extracellular Ph ◽  
The Other ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Rat Cardiomyocytes ◽  
Intracellular Ca ◽  
Isolated Rat

Although the Na+/H+ exchanger (NHE) is considered to be involved in regulation of intracellular Ca2+ concentration ([Ca2+]i) through the Na+/Ca2+ exchanger, the exact mechanisms of its participation in Ca2+ handling by cardiomyocytes are not fully understood. Isolated rat cardiomyocytes were treated with or without agents that are known to modify Ca2+ movements in cardiomyocytes and exposed to an NHE inhibitor, 5-( N-methyl- N-isobutyl)amiloride (MIA). [Ca2+]i in cardiomyocytes was measured spectrofluorometrically with fura 2-AM in the absence or presence of KCl, a depolarizing agent. MIA increased basal [Ca2+]i and augmented the KCl-induced increase in [Ca2+]i in a concentration-dependent manner. The MIA-induced increase in basal [Ca2+]i was unaffected by extracellular Ca2+, antagonists of the sarcolemmal (SL) L-type Ca2+ channel, and inhibitors of the SL Na+/Ca2+ exchanger, SL Ca2+ pump ATPase and mitochondrial Ca2+ uptake. However, the MIA-induced increase in basal [Ca2+]i was attenuated by inhibitors of SL Na+-K+-ATPase and sarcoplasmic reticulum (SR) Ca2+ transport. On the other hand, the MIA-mediated augmentation of the KCl response was dependent on extracellular Ca2+ concentration and attenuated by agents that inhibit SL L-type Ca2+ channels, the SL Na+/Ca2+ exchanger, SL Na+-K+-ATPase, and SR Ca2+ release channels and the SR Ca2+ pump. However, the effect of MIA on the KCl-induced increase in [Ca2+]i remained unaffected by treatment with inhibitors of SL Ca2+ pump ATPase and mitochondrial Ca2+ uptake. MIA and a decrease in extracellular pH lowered intracellular pH and increased basal [Ca2+]i, whereas a decrease in extracellular pH, in contrast to MIA, depressed the KCl-induced increase in [Ca2+]i in cardiomyocytes. These results suggest that NHE may be involved in regulation of [Ca2+]i and that MIA-induced increases in basal [Ca2+]i, as well as augmentation of the KCl-induced increase in [Ca2+]i, in cardiomyocytes are regulated differentially.

Ca dependence of Na influx during treatment of rabbit aorta with NE and high K solutions

1988 ◽  
Vol 254 (1) ◽  
pp. C75-C83 ◽  
Author(s):  
P. I. Aaronson ◽  
A. W. Jones
Keyword(s):  
Rabbit Aorta ◽  
Dependent Manner ◽  
Free Solution ◽  
Concentration Dependent Manner ◽  
High K ◽  
Ca Antagonist ◽  
Intracellular Ca ◽  
Isolated Rabbit Aorta ◽  
Ca Agonist

Cellular influx of 24Na was measured in isolated rabbit aorta during stimulation with 10 microM norepinephrine (NE) or depolarization with 80 mM K solution, using a pulse-labeling, cold-wash technique. NE caused a two- to threefold increase in Na influx; a smaller but significant increase was also observed in depolarized tissues. Basal and NE-induced fluxes at 1 min were significantly increased by a 20-min preincubation in a Ca-free solution containing 2 mM EGTA; elevation of [Mg] in this solution reduced these effects. The high K-induced influx was prevented by a combination of low Ca (30 microns) and elevated Mg (10 mM). The Ca agonist, BAY-K 8644, increased 24Na influx. The Ca antagonist, diltiazem, inhibited the depolarization-stimulated 24Na influx in a concentration-dependent manner, but was less effective in blocking the response to NE. Extension of the preincubation in NE plus Ca-free medium from 30 s to 15 min decreased the influx response and contraction. After exposure to NE in Ca-free solution, 24Na influx remained elevated 10 min after washing out NE in the continued absence of Ca. A second exposure to NE at that time did not increase influx. We propose that a component of 24Na influx during excitation depends directly on a rise in intracellular [Ca]. The role of an indirect effect of [Ca] on metabolic H+ production with subsequent stimulation of the Na+-H+ exchange may also be a factor.

Regulation of intracellular calcium in N1E-115 neuroblastoma cells: the role of Na+/Ca2+exchange

2002 ◽  
Vol 282 (5) ◽  
pp. C1000-C1008 ◽  
Author(s):  
Kara L. Kopper ◽  
Joseph S. Adorante
Keyword(s):  
Membrane Potential ◽  
Intracellular Calcium ◽  
Normal Cell ◽  
Buffer Capacity ◽  
Neuroblastoma Cells ◽  
Fold Increase ◽  
Scorpion Venom ◽  
Reverse Mode ◽  
Intracellular Ca

In fura 2-loaded N1E-115 cells, regulation of intracellular Ca2+ concentration ([Ca2+]i) following a Ca2+ load induced by 1 μM thapsigargin and 10 μM carbonylcyanide p-trifluoromethyoxyphenylhydrazone (FCCP) was Na+ dependent and inhibited by 5 mM Ni2+. In cells with normal intracellular Na+ concentration ([Na+]i), removal of bath Na+, which should result in reversal of Na+/Ca2+exchange, did not increase [Ca2+]i unless cell Ca2+ buffer capacity was reduced. When N1E-115 cells were Na+ loaded using 100 μM veratridine and 4 μg/ml scorpion venom, the rate of the reverse mode of the Na+/Ca2+ exchanger was apparently enhanced, since an ∼4- to 6-fold increase in [Ca2+]ioccurred despite normal cell Ca2+ buffering. In SBFI-loaded cells, we were able to demonstrate forward operation of the Na+/Ca2+ exchanger (net efflux of Ca2+) by observing increases (∼ 6 mM) in [Na+]i. These Ni2+ (5 mM)-inhibited increases in [Na+]i could only be observed when a continuous ionomycin-induced influx of Ca2+ occurred. The voltage-sensitive dye bis-(1,3-diethylthiobarbituric acid) trimethine oxonol was used to measure changes in membrane potential. Ionomycin (1 μM) depolarized N1E-115 cells (∼25 mV). This depolarization was Na+dependent and blocked by 5 mM Ni2+ and 250–500 μM benzamil. These data provide evidence for the presence of an electrogenic Na+/Ca2+ exchanger that is capable of regulating [Ca2+]i after release of Ca2+ from cell stores.

Intracellular calcium levels in the Plasmodium berghei ookinete

Parasitology ◽  
2008 ◽  
Vol 135 (12) ◽  
pp. 1355-1362 ◽  
Author(s):  
I. SIDÉN-KIAMOS ◽  
C. LOUIS
Keyword(s):  
Intracellular Calcium ◽  
Plasmodium Berghei ◽  
Calcium Concentration ◽  
Insect Cells ◽  
Ionophore A23187 ◽  
Rodent Malaria Parasite ◽  
Calcium Indicators ◽  
Intracellular Ca

SUMMARYOokinetes are the motile and invasive stages of Plasmodium parasites in the mosquito host. Here we explore the role of intracellular Ca2+ in ookinete survival and motility as well as in the formation of oocysts in vitro in the rodent malaria parasite Plasmodium berghei. Treatment with the Ca2+ ionophore A23187 induced death of the parasite, an effect that could be prevented if the ookinetes were co-incubated with insect cells before incubation with the ionophore. Treatment with the intracellular calcium chelator BAPTA/AM resulted in increased formation of oocysts in vitro. Calcium imaging in the ookinete using fluorescent calcium indicators revealed that the purified ookinetes have an intracellular calcium concentration in the range of 100 nm. Intracellular calcium levels decreased substantially when the ookinetes were incubated with insect cells and their motility was concomitantly increased. Our results suggest a pleiotropic role for intracellular calcium in the ookinete.

The P-element-induced silencing effect of KP transposons is dose dependent in Drosophila melanogaster

Genome ◽  
2011 ◽  
Vol 54 (9) ◽  
pp. 752-762 ◽  
Author(s):  
Alireza Sameny ◽  
John Locke
Keyword(s):  
Drosophila Melanogaster ◽  
Critical Role ◽  
Mutagenic Effect ◽  
P Element ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
P Elements ◽  
Single Well ◽  
Dose Dependent

Transposable elements are found in the genomes of all eukaryotes and play a critical role in altering gene expression and genome organization. In Drosophila melanogaster, transposable P elements are responsible for the phenomenon of hybrid dysgenesis. KP elements, a deletion-derivative of the complete P element, can suppress this mutagenic effect. KP elements can also silence the expression of certain other P-element-mediated transgenes in a process called P-element-dependent silencing (PDS), which is thought to involve the recruitment of heterochromatin proteins. To explore the mechanism of this silencing, we have mobilized KP elements to create a series of strains that contain single, well-defined KP insertions that show PDS. To understand the quantitative role of KP elements in PDS, these single inserts were combined in a series of crosses to obtain genotypes with zero, one, or two KP elements, from which we could examine the effect of KP gene dose. The extent of PDS in these genotypes was shown to be dose dependent in a logarithmic rather than linear fashion. A logarithmic dose dependency is consistent with the KP products interacting with heterochromatic proteins in a concentration-dependent manner such that two molecules are needed to induce gene silencing.

Bitter stimuli induce Ca2+ signaling and CCK release in enteroendocrine STC-1 cells: role of L-type voltage-sensitive Ca2+ channels

2006 ◽  
Vol 291 (4) ◽  
pp. C726-C739 ◽  
Author(s):  
Monica C. Chen ◽  
S. Vincent Wu ◽  
Joseph R. Reeve ◽  
Enrique Rozengurt
Keyword(s):  
Endocrine Cells ◽  
Bitter Taste ◽  
Dependent Manner ◽  
Gi Tract ◽  
Intracellular Ca ◽  
Α Subunits ◽  
Ca2 Channels ◽  
Cck Release

We previously demonstrated the expression of bitter taste receptors of the type 2 family (T2R) and the α-subunits of the G protein gustducin (Gαgust) in the rodent gastrointestinal (GI) tract and in GI endocrine cells. In this study, we characterized mechanisms of Ca2+ fluxes induced by two distinct T2R ligands: denatonium benzoate (DB) and phenylthiocarbamide (PTC), in mouse enteroendocrine cell line STC-1. Both DB and PTC induced a marked increase in intracellular [Ca2+] ([Ca2+]i) in a dose- and time-dependent manner. Chelating extracellular Ca2+ with EGTA blocked the increase in [Ca2+]i induced by either DB or PTC but, in contrast, did not prevent the effect induced by bombesin. Thapsigargin blocked the transient increase in [Ca2+]i induced by bombesin, but did not attenuate the [Ca2+]i increase elicited by DB or PTC. These results indicate that Ca2+ influx mediates the increase in [Ca2+]i induced by DB and PTC in STC-1 cells. Preincubation with the L-type voltage-sensitive Ca2+ channel (L-type VSCC) blockers nitrendipine or diltiazem for 30 min inhibited the increase in [Ca2+]i elicited by DB or PTC. Furthermore, exposure to the L-type VSCCs opener BAY K 8644 potentiated the increase in [Ca2+]i induced by DB and PTC. Stimulation with DB also induced a marked increase in the release of cholecystokinin from STC-1 cells, an effect also abrogated by prior exposure to EGTA or L-type VSCC blockers. Collectively, our results demonstrate that bitter tastants increase [Ca2+]i and cholecystokinin release through Ca2+ influx mediated by the opening of L-type VSCCs in enteroendocrine STC-1 cells.

Protein kinase C potentiates cAMP-stimulated mouse duodenal mucosal bicarbonate secretion in vitro

2004 ◽  
Vol 286 (5) ◽  
pp. G814-G821 ◽  
Author(s):  
Bi-Guang Tuo ◽  
Jimmy Y. C. Chow ◽  
Kim E. Barrett ◽  
Jon I. Isenberg
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Duodenal Mucosa ◽  
Bicarbonate Secretion ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Ussing Chambers ◽  
Duodenal Bicarbonate Secretion

PKC has been shown to regulate epithelial Cl- secretion in a variety of models. However, the role of PKC in duodenal mucosal bicarbonate secretion is less clear. We aimed to investigate the role of PKC in regulation of duodenal mucosal bicarbonate secretion. Bicarbonate secretion by murine duodenal mucosa was examined in vitro in Ussing chambers using a pH-stat technique. PKC isoform expression and activity were assessed by Western blotting and in vitro kinase assays, respectively. PMA (an activator of PKC) alone had no effect on duodenal bicarbonate secretion or short-circuit current ( Isc). When PMA and dibutyryl-cAMP (db-cAMP) were added simultaneously, PMA failed to alter db-cAMP-stimulated duodenal bicarbonate secretion or Isc ( P > 0.05). However, a 1-h preincubation with PMA potentiated db-cAMP-stimulated duodenal bicarbonate secretion and Isc in a concentration-dependent manner (from 10-8 to 10-5M) ( P < 0.05). PMA preincubation had no effects on carbachol- or heat-stable toxin-stimulated bicarbonate secretion. Western blot analysis revealed that PKCα, -γ, -ϵ, -θ, -μ, and -ι/λ were expressed in murine duodenal mucosa. Ro 31–8220 (an inhibitor active against PKCϵ, -α, -β, and -γ), but not Gö 6983 (an inhibitor active against PKCα, -γ, -β, and -δ), reversed the potentiating effect of PMA on db-cAMP-stimulated bicarbonate secretion. PMA also time- and concentration-dependently increased the activity of PKCϵ, an effect that was prevented by Ro 31–8220 but not Gö 6983. These results demonstrate that activation of PKC potentiates cAMP-stimulated duodenal bicarbonate secretion, whereas it does not modify basal secretion. The effect of PKC on cAMP-stimulated bicarbonate secretion is mediated by the PKCϵ isoform.

Changes in [Ca2+]i induced by several glucose transport-enhancing stimuli in rat epitrochlearis muscle

2003 ◽  
Vol 94 (5) ◽  
pp. 1813-1820 ◽  
Author(s):  
Shin Terada ◽  
Isao Muraoka ◽  
Izumi Tabata
Keyword(s):  
Glucose Transport ◽  
Quantitative Information ◽  
Dependent Manner ◽  
Sprague Dawley ◽  
Concentration Dependent Manner ◽  
Fura 2 ◽  
Sprague Dawley Rats ◽  
Intracellular Ca ◽  
Resting Muscle ◽  
Krebs Henseleit Buffer

The purpose of the present investigation was to establish a method for estimating intracellular Ca2+ concentrations ([Ca2+]i) in isolated rat epitrochlearis muscles. Epitrochlearis muscles excised from 4-wk-old male Sprague-Dawley rats were loaded with a fluorescent Ca2+indicator, fura 2-AM, for 60–90 min at 35°C in oxygenated Krebs-Henseleit buffer. After fura 2 loading and subsequent 20-min incubation, the intensities of 500-nm fluorescence, induced by 340- and 380-nm excitation lights (Ftotal340 and Ftotal380), were measured. The fluorescences specific to fura-2 (Ffura 2340 and Ffura 2380) were calculated by subtracting the non-fura 2-specific component from Ftotal340 and Ftotal380, respectively. The ratio of Ffura 2340 to Ffura 2380 was calculated as R, and the change in the ratio from the baseline value (ΔR) was used as an index of the change in [Ca2+]i. In resting muscle, ΔR was stable for 60 min. Incubation for 20 min with caffeine (3–10 mM) significantly increased ΔR in a concentration-dependent manner. Incubation with hypoxic Krebs-Henseleit buffer for 10–60 min significantly elevated ΔR, depending on the duration of the incubation. Incubation with 50 μM N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide for 20 min significantly elevated ΔR ( P < 0.05). No significant increases in ΔR were observed during incubation for 20 min with 2 mM 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside or with 2 mU/ml insulin. These results demonstrated that, by using the fura 2-AM fluorescence method, the changes in [Ca2+]i can be monitored in the rat epitrochlearis muscle and suggest that the method can be utilized to observe quantitative information regarding [Ca2+]i that may be involved in contraction- and hypoxia-stimulated glucose transport activity in skeletal muscle.

scholarly journals Sec1p directly stimulates SNARE-mediated membrane fusion in vitro

2004 ◽  
Vol 167 (1) ◽  
pp. 75-85 ◽  
Author(s):  
Brenton L. Scott ◽  
Jeffrey S. Van Komen ◽  
Hassan Irshad ◽  
Song Liu ◽  
Kirilee A. Wilson ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Membrane Fusion ◽  
Membrane Trafficking ◽  
Snare Complex ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Bud Neck ◽  
Precise Role

Sec1 proteins are critical players in membrane trafficking, yet their precise role remains unknown. We have examined the role of Sec1p in the regulation of post-Golgi secretion in Saccharomyces cerevisiae. Indirect immunofluorescence shows that endogenous Sec1p is found primarily at the bud neck in newly budded cells and in patches broadly distributed within the plasma membrane in unbudded cells. Recombinant Sec1p binds strongly to the t-SNARE complex (Sso1p/Sec9c) as well as to the fully assembled ternary SNARE complex (Sso1p/Sec9c;Snc2p), but also binds weakly to free Sso1p. We used recombinant Sec1p to test Sec1p function using a well-characterized SNARE-mediated membrane fusion assay. The addition of Sec1p to a traditional in vitro fusion assay moderately stimulates fusion; however, when Sec1p is allowed to bind to SNAREs before reconstitution, significantly more Sec1p binding is detected and fusion is stimulated in a concentration-dependent manner. These data strongly argue that Sec1p directly stimulates SNARE-mediated membrane fusion.

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