Enhancement of calcium transport in Caco-2 monolayer through PKCζ-dependent Cav1.3-mediated transcellular and rectifying paracellular pathways by prolactin

2009 ◽  
Vol 296 (6) ◽  
pp. C1373-C1382 ◽  
Author(s):  
Narongrit Thongon ◽  
La-iad Nakkrasae ◽  
Jirawan Thongbunchoo ◽  
Nateetip Krishnamra ◽  
Narattaphol Charoenphandhu
Keyword(s):  
Protein Kinase ◽  
Calcium Channel ◽  
Calcium Transport ◽  
Calcium Absorption ◽  
Protein Biosynthesis ◽  
Ussing Chamber ◽  
Calcium Flux ◽  
Channel Blockers ◽  
Voltage Dependent ◽  
Calbindin D

Previous investigations suggested that prolactin (PRL) stimulated the intestinal calcium absorption through phosphoinositide 3-kinase (PI3K), protein kinase C (PKC), and RhoA-associated coiled-coil forming kinase (ROCK) signaling pathways. However, little was known regarding its detailed mechanisms for the stimulation of transcellular and voltage-dependent paracellular calcium transport. By using Ussing chamber technique, we found that the PRL-induced increase in the transcellular calcium flux and decrease in transepithelial resistance of intestinal-like Caco-2 monolayer were not abolished by inhibitors of gene transcription and protein biosynthesis. The PRL-stimulated transcellular calcium transport was completely inhibited by the L-type calcium channel blockers (nifedipine and verapamil) and plasma membrane Ca2+-ATPase (PMCA) inhibitor (trifluoperazine) as well as small interfering RNA targeting voltage-dependent L-type calcium channel Cav1.3, but not TRPV6 or calbindin-D9k. As demonstrated by 45Ca uptake study, PI3K and PKC, but not ROCK, were essential for the PRL-enhanced apical calcium entry. In addition, PRL was unable to enhance the transcellular calcium transport after PKCζ knockdown or exposure to inhibitors of PKCζ, but not of PKCα, PKCβ, PKCε, PKCμ, or protein kinase A. Voltage-clamping experiments further showed that PRL markedly stimulated the voltage-dependent calcium transport and removed the paracellular rectification. Such PRL effects on paracellular transport were completely abolished by inhibitors of PI3K (LY-294002) and ROCK (Y-27632). It could be concluded that the PRL-stimulated transcellular calcium transport in Caco-2 monolayer was mediated by Cav1.3 and PMCA, presumably through PI3K and PKCζ pathways, while the enhanced voltage-dependent calcium transport occurred through PI3K and ROCK pathways.

Effects of 1,25(OH)2D3 and calcium channel blockers on cecal calcium transport in the rat

1985 ◽  
Vol 248 (6) ◽  
pp. G676-G681 ◽  
Author(s):  
M. J. Favus ◽  
E. Angeid-Backman
Keyword(s):  
Vitamin D ◽  
Calcium Channel ◽  
Calcium Transport ◽  
Calcium Absorption ◽  
Channel Blockers ◽  
Dihydroxyvitamin D3 ◽  
High Concentrations ◽  
Absorptive Flux ◽  
Mucosal Side

To determine whether calcium transport across rat cecum is vitamin D dependent, we measured in vitro bidirectional calcium fluxes under short-circuited conditions across cecum from rats that were vitamin D deficient, vitamin D replete, or vitamin D deficient or vitamin D replete and injected with either 10, 25, or 75 ng of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] daily for 4 days before study. Vitamin D deficiency decreased net calcium absorption (Jnet) by reducing the mucosal-to-serosal absorptive flux (Jm----s) from 168 +/- 18 to 33 +/- 5 nmol X cm-2 X h-1 (mean +/- SE, P less than 0.0001). Twenty-five nanograms of 1,25(OH)2D3 raised Jm----s to 124 +/- 17 nmol X cm-2 X h-1, not different from values in vitamin D-replete rats. Although active calcium absorption by cecum appears to respond to vitamin D, calcium Jm----s is near maximal under normal conditions, and further stimulation follows only pharmacological doses of 1,25(OH)2D3. The in vitro addition of the calcium channel blocker verapamil (5 X 10(-5) M) to the mucosal side of cecum from vitamin D-replete rats reduced calcium Jm----s, but lower concentrations of verapamil or nitrendipine (10(-5) to 10(-9) M) did not reduce calcium Jm----s. The lack of inhibition by low concentrations of channel blockers suggest that the plasma membrane channels for calcium translocation across intestinal epithelium may not be analogous to voltage-dependent calcium channels in excitable tissue. The inhibition of cecal calcium transport that was blocked by high concentrations of verapamil may represent a nonspecific effect of the agent.

Prolactin directly stimulates transcellular active calcium transport in the duodenum of female rats

2001 ◽  
Vol 79 (5) ◽  
pp. 430-438 ◽  
Author(s):  
Narattaphol Charoenphandhu ◽  
Liangchai Limlomwongse ◽  
Nateetip Krishnamra
Keyword(s):  
Calcium Transport ◽  
Calcium Absorption ◽  
Direct Action ◽  
Female Rats ◽  
Calcium Flux ◽  
Total Calcium ◽  
Solvent Drag ◽  
Control Value ◽  
Voltage Dependent ◽  
Active Calcium

Prolactin has been postulated to be a novel calcium-regulating hormone during pregnancy and lactation. It stimulates both passive and active duodenal calcium transport in several experimental models. Our study was performed on sexually mature female Wistar rats (200–250 g) to study the direct action of prolactin on calcium transport in the duodenum using the Ussing chamber technique. To evaluate the effect of prolactin on total calcium transport in the duodenum, we intraperitoneally injected rats with 0.4, 0.6, and 0.8 mg/kg prolactin. The total calcium transport was divided into voltage-dependent, solvent drag-induced, and transcellular active fluxes by applying short-circuit current and by mucosal glucose replacement with mannitol. The effect of prolactin on each flux was studied separately. Finally, to evaluate the direct action of prolactin on duodenal transcellular active flux, we directly exposed duodenal segments to prolactin that had been added to the serosal solution with or without calcium transport inhibitors. We found that 0.6 and 0.8 mg/kg prolactin ip significantly increased the total mucosa–to–serosa calcium flux from the control value (nmol·hr–1·cm–2) of 34.53 ± 6.81 to 68.07 ± 13.53 (P < 0.05) and 84.43 ± 19.72 (P < 0.01), respectively. Prolactin also enhanced the solvent drag-induced calcium flux and transcellular active calcium flux, but not the voltage-dependent calcium flux. The duodenal segments directly exposed to 200, 400, and 800 ng/mL prolactin showed a significant increase in the transcellular active calcium absorption in a dose-dependent manner, i.e., from the control value (nmol·hr–1·cm–2) of 2.94 ± 0.47 to 5.45 ± 0.97 (P < 0.01), 8.09 ± 0.52 (P < 0.001), and 18.42 ± 2.92 (P < 0.001), respectively. Its direct action was inhibited by mucosal exposure to 50 µM lanthanum chloride, a calcium transporter protein competitor, and serosal exposure to 0.1 mM trifluoperazine, a Ca2+-ATPase inhibitor. These studies demonstrate that the duodenum is a target organ of prolactin, which enhances transcellular active calcium transport.Key words: calcium absorption, duodenum, prolactin, solvent drag, transcellular calcium transport.

scholarly journals Impaired renal calcium absorption in mice lacking calcium channel β3 subunits

2009 ◽  
Vol 87 (7) ◽  
pp. 522-530 ◽  
Author(s):  
José F. Bernardo ◽  
Clara E. Magyar ◽  
W. Bruce Sneddon ◽  
Peter A. Friedman
Keyword(s):  
Plasma Membrane ◽  
Transgenic Mice ◽  
Calcium Channels ◽  
Calcium Channel ◽  
Sodium Excretion ◽  
Calcium Transport ◽  
Calcium Absorption ◽  
Wild Type ◽  
Distal Tubules ◽  
Calbindin D

Transgenic mice lacking calcium channel β3 subunits (CaVβ3) were used to determine the involvement of a multimeric calcium channel in mediating stimulated renal calcium absorption. We measured the ability of calcium channel β3 subunit-null (CaVβ3−/−) and wild-type (CaVβ3+/+) mice to increase renal calcium absorption in response to the calcium-sparing diuretic chlorothiazide (CTZ). Control rates of fractional sodium excretion were comparable in CaVβ3−/− and CaVβ3+/+ mice and CTZ increased sodium excretion similarly in both groups. CTZ enhanced calcium absorption only in wild-type CaVβ3+/+ mice. This effect was specific for diuretics acting on distal tubules because both CaVβ3−/− and CaVβ3+/+ mice responded comparably to furosemide. The absence of β3 subunits resulted in compensatory increases of TrpV5 calcium channels, the plasma membrane Ca-ATPase, NCX1 Na/Ca exchanger protein, and calbindin-D9k but not calbindin-D28k. We conclude that TrpV5 mediates basal renal calcium absorption and that a multimeric calcium channel that includes CaVβ3 mediates stimulated calcium transport.

Calcium transport in turtle bladder

1987 ◽  
Vol 253 (6) ◽  
pp. R917-R921
Author(s):  
S. Sabatini ◽  
N. A. Kurtzman
Keyword(s):  
Calcium Transport ◽  
Calcium Absorption ◽  
Short Circuit ◽  
Open Circuit ◽  
Calcium Flux ◽  
Bladder Epithelium ◽  
Control Value ◽  
Sodium Calcium ◽  
Turtle Bladder ◽  
Absorptive Flux

Unidirectional 45Ca fluxes were measured in the turtle bladder under open-circuit and short-circuit conditions. In the open-circuited state net calcium flux (JnetCa) was secretory (serosa to mucosa) and was 388.3 +/- 84.5 pmol.mg-1.h-1 (n = 20, P less than 0.001). Ouabain (5 X 10(-4) M) reversed JnetCa to an absorptive flux (serosal minus mucosal flux = -195.8 +/- 41.3 pmol.mg-1.h-1; n = 20, P less than 0.001). Amiloride (1 X 10(-5) M) reduced both fluxes such that JnetCa was not significantly different from zero. Removal of mucosal sodium caused net calcium absorption; removal of serosal sodium caused calcium secretion. When bladders were short circuited, JnetCa decreased to approximately one-third of control value but remained secretory (138.4 +/- 54.3 pmol.mg-1.h-1; n = 9, P less than 0.025). When ouabain was added under short-circuit conditions, JnetCa was similar in magnitude and direction to ouabain under open-circuited conditions (i.e., absorptive). Tissue 45Ca content was approximately equal to 30-fold lower when the isotope was placed in the mucosal bath, suggesting that the apical membrane is the resistance barrier to calcium transport. The results obtained in this study are best explained by postulating a Ca2+-ATPase on the serosa of the turtle bladder epithelium and a sodium-calcium antiporter on the mucosa. In this model, the energy for calcium movement would be supplied, in large part, by the Na+-K+-ATPase. By increasing cell sodium, ouabain would decrease the activity of the mucosal sodium-calcium exchanger (or reverse it), uncovering active calcium transport across the serosa.

Epithelial calcium transporter expression in human duodenum

2001 ◽  
Vol 280 (2) ◽  
pp. G285-G290 ◽  
Author(s):  
Natalie F. Barley ◽  
Alison Howard ◽  
David O'Callaghan ◽  
Stephen Legon ◽  
Julian R. F. Walters
Keyword(s):  
Vitamin D ◽  
Calcium Channel ◽  
Calcium Absorption ◽  
Apical Membrane ◽  
Calcium Influx ◽  
Basolateral Membrane ◽  
Cdna Probe ◽  
Vitamin D Metabolites ◽  
Expression Of Genes ◽  
Calbindin D

Calcium absorption in intestine and kidney involves transport through the apical membrane, cytoplasm, and basolateral membrane of the epithelial cells. Apical membrane calcium influx channels have recently been described in rabbit (epithelial calcium channel, ECaC) and rat (calcium transport protein, CaT1). We amplified from human duodenum a 446-base partial cDNA probe (ECAC2) having a predicted amino acid similarity of 97% to rat CaT1. Duodenum, but not ileum, colon, or kidney, expressed a 3-kb transcript. A larger transcript was also found in placenta and pancreas, and a different, faint transcript was found in brain. In duodenal biopsies from 20 normal volunteers, expression varied considerably but was not significantly correlated with vitamin D metabolites. This signal correlated with calbindin-D9k ( r = 0.48, P< 0.05) and more strongly with the plasma membrane calcium ATPase PMCA1 ( r = 0.83, P < 0.001). These data show that although individual variations in calcium channel transcripts are not vitamin D dependent, expression of genes governing apical entry and basolateral extrusion are tightly linked. This may account for some of the unexplained variability in calcium absorption.

Ca fluxes across duodenum and colon of spontaneously hypertensive rats: effect of 1,25(OH)2D3

1986 ◽  
Vol 251 (2) ◽  
pp. F278-F282 ◽  
Author(s):  
U. Gafter ◽  
S. Kathpalia ◽  
D. Zikos ◽  
K. Lau
Keyword(s):  
Calcium Transport ◽  
Spontaneously Hypertensive Rats ◽  
Calcium Absorption ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Calcium Flux ◽  
Hypertensive Rats ◽  
Spontaneously Hypertensive ◽  
Wistar Kyoto

Calcium absorption by spontaneously hypertensive rats (SHR) was variably reported to be different from normotensive Wistar-Kyoto (WKY) controls. Furthermore, blunted responsiveness to the intestinal effects of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] has also been postulated. To evaluate this hypothesis, calcium fluxes were measured by the Ussing technique across duodenum and descending colon with or without prior 1,25(OH)2D3 treatment. Duodenal mucosal-to-serosal calcium flux (Jm----s) (44.9 vs. 52.4 nmol X cm-2 X h-1), serosal-to-mucosal flux (Js----m) (25.6 vs. 28.4 nmol X cm-2 X h-1), and net flux (Jnet) were comparable. 1,25(OH)2D3 increased duodenal Jm----s in both SHR and WKY groups (95.2 and 86.8 nmol X cm-2 X h-1). Js----m was lower in SHR (26.1 vs. 35.6 nmol X cm-2 X h-1, P less than 0.01), although the tendency for a higher Jnet in SHR (68.6 vs. 51.2 nmoles X cm-2 X h-1) was statistically insignificant. Short-circuit current was higher in the colon of SHR, both before and after 1,25(OH)2D3, suggesting increased sodium transport. Basal colonic Jnet was virtually zero in both groups but comparably increased by 1,25(OH)2D3 because of stimulation in only Jm----s. Prevention of hypertension by hydralazine since the 4th wk of age did not alter the findings compared with the hypertensive SHR, suggesting calcium transport rates were unaffected by hypertension. These data indicate that in vitro, duodenal, and colonic active calcium transport by the SHR is similar to WKY. Their normal responses to 1,25(OH)2D3 do not support the hypothesis of intestinal resistance.

scholarly journals Bio-inspired voltage-dependent calcium channel blockers

2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Tingting Yang ◽  
Lin-Ling He ◽  
Ming Chen ◽  
Kun Fang ◽  
Henry M. Colecraft
Keyword(s):  
Calcium Channel ◽  
Calcium Channel Blockers ◽  
Channel Blockers ◽  
Voltage Dependent Calcium Channel ◽  
Voltage Dependent
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