Interplay of Hydrogen Sulfide and Nitric Oxide on the Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

We studied whether nitric oxide (NO) and hydrogen sulfide (H2S) have an interaction on the pacemaker activities of interstitial cells of Cajal (ICC) from the mouse small intestine. The actions of NO and H2S on pacemaker activities were investigated by using the whole-cell patch-clamp technique and intracellular Ca2+ analysis at 30℃ in cultured mouse ICC. Exogenously applied (±)-S-nitroso-N-acetylpenicillamine (SNAP), an NO donor, or sodium hydrogen sulfide (NaHS), a donor of H2S, showed no influence on pacemaker activity (potentials and currents) in ICC at low concentrations (10 µM SNAP and 100 µM NaHS), but SNAP or NaHS completely inhibited pacemaker amplitude and pacemaker frequency with increases in the resting currents in the outward direction at high concentrations (SNAP 100 µM and NaHS 1 mM). Co-treatment with 10 µM SNAP plus 100 µM NaHS also inhibited pacemaker amplitude and pacemaker frequency with increases in the resting currents in the outward direction. ODQ, a guanylate cyclase inhibitor, or glibenclamide, an ATP-sensitive K+ channel inhibitor, blocked the SNAP+NaHS-induced inhibition of pacemaker currents in ICC. Also, we found that SNAP+NaHS inhibited the spontaneous intracellular Ca2+ ([Ca2+]i) oscillations in cultured ICC. In conclusion, this study describes the enhanced inhibitory effects of NO plus H2S on ICC in the mouse small intestine. NO+H2S inhibited the pacemaker activity of ICC by modulating intracellular Ca2+. These results may be evidence of a physiological interaction of NO and H2S in ICC for modulating gastrointestinal motility

Mechanisms of phytoestrogen biochanin A-induced vasorelaxation in renovascular hypertensive rats

Background: The plant-derived estrogen biochanin A is known to cause vasodilation, but its mechanism of action in hypertension remains unclear. This study was undertaken to investigate the effects and mechanisms of biochanin A on the thoracic aorta in two-kidney, one clip (2K1C) renovascular hypertensive rats. Methods: Hypertension was induced by clipping the left renal artery, and control age-matched rats were sham treated. Thoracic aortae were mounted in tissue baths to measure isometric tension. Results: Biochanin A caused concentration-dependent relaxation in aortic rings from 2K1C hypertensive and sham-treated rats, which was greater in 2K1C rats than in sham rats. Biochanin A-induced relaxation was significantly attenuated by removing the endothelium in aortic rings from 2K1C rats, but not in sham rats. Nω-Nitro-l-arginine methyl ester, a nitric oxide synthase inhibitor, or indomethacin, a cyclooxygenase inhibitor, did not affect the biochanin A-induced relaxation in aortic rings from 2K1C and sham rats. By contrast, treatment with glibenclamide, a selective inhibitor of adenosine triphosphate-sensitive K+ channels, or tetraethylammonium, an inhibitor of Ca2+-activated K+ channels, significantly reduced biochanin A-induced relaxation in aortic rings from both groups. However, 4-aminopyridine, a selective inhibitor of voltage-dependent K+ channels, inhibited the relaxation induced by biochanin A in 2K1C rats, whereas no significant differences were observed in sham rats. Conclusion: These results suggest that the enhanced relaxation caused by biochanin A in aortic rings from hypertensive rats is endothelium dependent. Vascular smooth muscle K+ channels may be involved in biochanin A-induced relaxation in aortae from hypertensive and normotensive rats. In addition, an endothelium-derived activation of voltage-dependent K+ channels contributes, at least in part, to the relaxant effect of biochanin A in renovascular hypertension

Activating of ATP-dependent K+ channels comprised of K(ir) 6.2 and SUR 2B by PGE2 through EP2 receptor in cultured interstitial cells of Cajal from murine small intestine

The interstitial cells of Cajal (ICC) are pacemaker cells in gastrointestinal tract and generate an electrical rhythm in gastrointestinal muscles. We investigated the possibility that PGE(2) might affect the electrical properties of cultured ICC by activating ATP-dependent K(+) channels and, the EP receptor subtypes and the subunits of ATP-dependent K(+) channels involved in these activities were identified. In addition, the regulation of intracellular Ca(2+) ([Ca(2+)](i)) mobilization may be involved the action of PGE(2) on ICC. Treatments of ICC with PGE(2) inhibited electrical pacemaker activities in the same manner as pinacidil, an ATP-dependent K(+) channel opener and PGE(2) had only a dose-dependent effect. Using RT-PCR technique, we found that ATP-dependent K(+) channels exist in ICC and that these are composed of K(ir) 6.2 and SUR 2B subunits. To characterize the specific membrane EP receptor subtypes in ICC, EP receptor agonists and RT-PCR were used: Butaprost (an EP(2) receptor agonist) showed the actions on pacemaker currents in the same manner as PGE(2). However sulprostone (a mixed EP(1) and EP(3) agonist) had no effects. In addition, RT-PCR results indicated the presence of the EP(2) receptor in ICC. To investigate cAMP involvement in the effects of PGE(2) on ICCs, SQ-22536 (an inhibitor of adenylate cyclase) and cAMP assays were used. SQ-22536 did not affect the effect of PGE(2) on pacemaker currents, and PGE(2) did not stimulate cAMP production. Also, we found PGE(2) inhibited the spontaneous [Ca(2+)](i) oscillations in cultured ICC. These observations indicate that PGE(2) alters pacemaker currents by activating the ATP-dependent K(+) channels comprised of K(ir) 6.2-SUR 2B in ICC and this action of PGE(2) are through EP(2) receptor subtype and also the activation of ATP-dependent K(+) channels involves intracellular Ca(2+) mobilization

Phentolamine Inhibits the Pacemaker Activity of Mouse Interstitial Cells of Cajal by Activating ATP-Sensitive K(+) Channels

The aim of this study was to clarify if phentolamine has proven effects on the pacemaker activities of interstitial cells of Cajal (ICC) from the mouse small intestine involving the ATP-sensitive K(+) channels and adrenergic receptor. The actions of phentolamine on pacemaker activities were investigated using whole-cell patch-clamp technique and intracellular Ca(2+) analysis at 30 degrees C in cultured mouse intestinal ICC. ICC generated spontaneous pacemaker currents at a holding potential of -70 mV. Treatment with phentolamine reduced the frequency and amplitude of the pacemaker currents and increased the resting outward currents. Moreover, under current clamping (I = 0), phentolamine hyperpolarized the ICC membrane and decreased the amplitude of the pacemaker potentials. We also observed that phentolamine inhibited spontaneous [Ca(2+)](i) oscillations in ICC. The alpha-adrenergic drugs prazosin, yohimbine, methoxamine, and clonidine had no effect on ICC intestinal pacemaker activity and did not block phentolamine-induced effects. Phentolamine-induced effects on the pacemaker currents and the pacemaker potentials were significantly inhibited by ATP sensitive K(+) channel blocker glibenclamide, but not by TEA, apamin, or 4-aminopyridine. In addition, the NO synthase inhibitor, L-NAME and the guanylate cyclase inhibitor, ODQ were incapable of blocking the phentolamine-induced effects. These results demonstrate that phentolamine regulates the pacemaker activity of ICC via ATP-sensitive K(+) channel activation. Phentolamine could act through an adrenergic receptor- and also through NO-independent mechanism that involves intracellular Ca(2+) signaling.Epperson A, 2000, AM J PHYSIOL-CELL PH, V279, pC529Deka DK, 2004, EUR J PHARMACOL, V492, P13, DOI 10.1016/j.ejphar.2004.03.057Silva LFG, 2005, INT J IMPOT RES, V17, P27, DOI 10.1038/sj.ijir.3901269Rodrigo GC, 2005, CURR PHARM DESIGN, V11, P1915Jun JY, 2005, BRIT J PHARMACOL, V144, P242, DOI 10.1038/sj.bjp.0706074Kito Y, 2005, AM J PHYSIOL-CELL PH, V288, pC710, DOI 10.1152/ajpcell.00361.2004Choi S, 2006, CELL PHYSIOL BIOCHEM, V18, P187Sanders KM, 2006, ANNU REV PHYSIOL, V68, P307, DOI 10.1146/annurev.physiol.68.040504.094718Ward SM, 2006, J PHYSIOL-LONDON, V576, P675, DOI 10.1113/jphysiol.2006.117390Park CG, 2007, N-S ARCH PHARMACOL, V376, P175, DOI 10.1007/s00210-007-0187-1Hoy M, 2001, J BIOL CHEM, V276, P924Vemulapalli S, 2001, FUNDAM CLIN PHARM, V15, P1Jun JY, 2001, AM J PHYSIOL-CELL PH, V281, pC857Brayden JE, 2002, CLIN EXP PHARMACOL P, V29, P312Jun JY, 2004, BRIT J PHARMACOL, V141, P670, DOI 10.1038/sj.bjp.0705665Mannhold R, 2004, MED RES REV, V24, P213, DOI 10.1002/med.10060Suzuki H, 2000, J PHYSIOL-LONDON, V525, P105Proks P, 1997, P NATL ACAD SCI USA, V94, P11716Kubo M, 1997, J PHYSIOL-LONDON, V503, P489Shepherd RM, 1996, BRIT J PHARMACOL, V119, P911MURPHY ME, 1995, J PHYSIOL-LONDON, V486, P47HUIZINGA JD, 1995, NATURE, V373, P347Rustenbeck I, 1995, EXP CLIN ENDOCR DIAB, V103, P42WARD SM, 1994, J PHYSIOL-LONDON, V480, P91ZHANG L, 1994, AM J PHYSIOL, V267, pG494WILDE AAM, 1994, CARDIOVASC RES, V28, P847QUAYLE JM, 1994, J PHYSIOL-LONDON, V475, P9JONAS JC, 1992, BRIT J PHARMACOL, V107, P8SCHWIETERT R, 1992, EUR J PHARMACOL, V211, P87SMALL RC, 1992, BRAZ J MED BIOL RES, V25, P983PLANT TD, 1991, BRIT J PHARMACOL, V104, P385DUNNE MJ, 1991, BRIT J PHARMACOL, V103, P1847HOFFMANN BB, 1991, PHARMACOL BASIS THER, pCH10PLANT TD, 1990, BRIT J PHARMACOL, V101, P115

Effects of prostaglandin F2α on small intestinal interstitial cells of Cajal

AIM: To explore the role of prostaglandin F2α (PGF2α)) on pacemaker activity in interstitial cells of Cajal (ICC) from mouse small intestine

Action of imipramine on activated ATP-sensitive K(+) channels in interstitial cells of Cajal from murine small intestine

Tricyclic antidepressants have been widely used for the treatment of depression and as a therapeutic agent for the altered gastrointestinal (GI) motility of irritable bowel syndrome (IBS). The aim of this study was to clarify whether antidepressants directly modulate pacemaker currents in cultured interstitial cells of Cajal (ICC). We used the whole-cell patch-clamp techniques at 30 degrees C in cultured ICC from the mouse small intestine. Treatment of pinacidil, an ATP-sensitive K(+) channel opener, in the ICC using the current clamping mode, produced hyperpolarization of the membrane potential and decreased the amplitude of the pacemaker potentials. With the voltage clamp mode, we observed a decrease in the frequency and amplitude of pacemaker currents and increases in the resting outward currents. These effects of pinacidil on pacemaker potentials and currents were completely suppressed by glibenclamide, an ATP-sensitive K(+) channel blocker. Also, with the current clamp mode, imipramine blocked the affect of pinacidil on the pacemaker potentials. Observations of the voltage clamp mode with imipramine, desipramine and amitryptyline suppressed the action of pinacidil in the ICC. Next, we examined whether protein kinase C (PKC) and the G protein are involved in the action of imipramine on pinacidil induced pacemaker current inhibition. We used chelerythrine, a potent PKC inhibitor and GDPbetaS, a nonhydrolyzable guanosine 5-diphosphate (GDP) analogue that permanently inactivates GTP-binding proteins. We found that pretreatment with chelerythrine and intracellular application of GDPbetaS had no influence on the blocking action of imipramine on inhibited pacemaker currents by pinacidil. We conclude that imipramine inhibited the activated ATP-sensitive K(+) channels in ICC. This action does not appear to be mediated through the G protein and protein kinase C. Furthermore, this study may suggest another possible mechanism for tricyclic antidepressants related modulation of GI motility

Bradykinin modulates pacemaker currents through bradykinin B2 receptors in cultured interstitial cells of Cajal from the murine small intestine

1. We studied the modulation of pacemaker activities by bradykinin in cultured interstitial cells of Cajal (ICC) from murine small intestine with the whole-cell patch-clamp technique. Externally applied bradykinin produced membrane depolarization in the current-clamp mode and increased tonic inward pacemaker currents in the voltage-clamp mode. 2. Pretreatment with bradykinin B1 antagonist did not block the bradykinin-induced effects on pacemaker currents. However, pretreatment with bradykinin B2 antagonist selectively blocked the bradykinin-induced effects. Also, only externally applied selective bradykinin B2 receptor agonist produced tonic inward pacemaker currents and ICC revealed a colocalization of the bradykinin B2 receptor and c-kit immunoreactivities, but bradykinin B1 receptors did not localize in ICC. 3. External Na(+)-free solution abolished the generation of pacemaker currents and inhibited the bradykinin-induced tonic inward current. However, a Cl(−) channel blocker (DIDS) did not block the bradykinin-induced tonic inward current. 4. The pretreatment with Ca(2+)-free solution and thapsigargin, a Ca(2+)-ATPase inhibitor in endoplasmic reticulum, abolished the generation of pacemaker currents and suppressed the bradykinin-induced action. 5. Chelerythrine and calphostin C, protein kinase C inhibitors or naproxen, an inhibitor of cyclooxygenase, did not block the bradykinin-induced effects on pacemaker currents. 6. These results suggest that bradykinin modulates the pacemaker activities through bradykinin B2 receptor activation in ICC by external Ca(2+) influx and internal Ca(2+) release via protein kinase C- or cyclooxygenase-independent mechanism. Therefore, the ICC are targets for bradykinin and their interaction can affect intestinal motility

Receptor Tyrosine and MAP Kinase are Involved in Effects of H(2)O(2) on Interstitial Cells of Cajal in Murine Intestine

Hydrogen peroxide (H(2)O(2)) is involved in intestinal motility through changes of smooth muscle activity. However, there is no report as to the modulatory effects of H(2)O(2) on interstitial cells of Cajal (ICC). We investigated the H(2)O(2) effects and signal transductions to determine whether the intestinal motility can be modulated through ICC. We performed whole-cell patch clamp in cultured ICC from murine intestine and molecular analyses. H(2)O(2) hyperpolarized the membrane and inhibited pacemaker currents. These effects were inhibited by glibenclamide, an inhibitor of ATP-sensitive K(+) (K(ATP)) channels. The free radical scavenger catalase inhibited the H(2)O(2)-induced effects. MAFP and AACOCF(3) (a cytosolic phospholipase A(2) inhibitors) or SC-560 and NS-398 (a selective COX-1 and 2 inhibitor) or AH6809 (an EP(2) receptor antagonist) inhibited the H(2)O(2)-induced effects. PD98059 (a mitogen activated/ERK-activating protein kinase inhibitor) inhibited the H(2)O(2)-induced effects, though SB-203580 (a p38 MAPK inhibitor) or a JNK inhibitor did not affect. H(2)O(2)-induced effects could not be inhibited by LY-294002 (an inhibitor of PI(3)-kinases), calphostin C (a protein kinase C inhibitor), or SQ-22536 (an adenylate cyclase inhibitor). Adenoviral infection analysis revealed H(2)O(2) stimulated tyrosine kinase activity and AG 1478 (an antagonist of epidermal growth factor receptor tyrosine kinase) inhibited the H(2)O(2)-induced effects. These results suggest H(2)O(2) can modulate ICC pacemaker activity and this occur by the activation of K(ATP) channels through PGE(2) production via receptor tyrosine kinase-dependent MAP kinase activation

Receptor tyrosine and MAP kinase are involved in effects of H(2)O(2) on interstitial cells of Cajal in murine intestine

Hydrogen peroxide (H(2)O(2)) is involved in intestinal motility through changes of smooth muscle activity. However, there is no report as to the modulatory effects of H(2)O(2) on interstitial cells of Cajal (ICC). We investigated the H(2)O(2) effects and signal transductions to determine whether the intestinal motility can be modulated through ICC. We performed whole-cell patch clamp in cultured ICC from murine intestine and molecular analyses. H(2)O(2) hyperpolarized the membrane and inhibited pacemaker currents. These effects were inhibited by glibenclamide, an inhibitor of ATP-sensitive K+ (K(ATP)) channels. The free-radical scavenger catalase inhibited the H(2)O(2)-induced effects. MAFP and AACOCF(3) (a cytosolic phospholipase A(2) inhibitors) or SC-560 and NS-398 (a selective COX-1 and 2 inhibitor) or AH6809 (an EP(2) receptor antagonist) inhibited the H(2)O(2)-induced effects. PD98059 (a mitogen activated/ERK-activating protein kinase inhibitor) inhibited the H(2)O(2)-induced effects, though SB-203580 (a p38 MAPK inhibitor) or a JNK inhibitor did not affect. H(2)O(2)-induced effects could not be inhibited by LY-294002 (an inhibitor of PI(3)-kinases), calphostin C (a protein kinase C inhibitor) or SQ-22536 (an adenylate cyclase inhibitor). Adenoviral infection analysis revealed H(2)O(2) stimulated tyrosine kinase activity and AG 1478 (an antagonist of epidermal growth factor receptor tyrosine kinase) inhibited the H(2)O(2)-induced effects. These results suggest H(2)O(2) can modulate ICC pacemaker activity and this occur by the activation of K(ATP) channels through PGE(2) production via receptor tyrosine kinase-dependent MAP kinase activation.Song HJ, 2005, J PHARMACOL EXP THER, V312, P391, DOI 10.1124/jpet.104.074401Shimojima N, 2005, PHARMACOLOGY, V74, P95, DOI 10.1159/000084021Cao WB, 2004, J PHARMACOL EXP THER, V311, P60, DOI 10.1124/jpet.104.068023Lin YF, 2004, P NATL ACAD SCI USA, V101, P7799, DOI 10.1073/pnas.0402496101Ward SM, 2004, NEUROGASTROENT MOTIL, V16, P112Prasad M, 2004, AM J PHYSIOL-CELL PH, V286, pC671, DOI 10.1152/ajpcell.00137.2003Jun JY, 2004, BRIT J PHARMACOL, V141, P670, DOI 10.1038/sj.bjp.0705665Jabbour HN, 2003, J CLIN ENDOCR METAB, V88, P4481, DOI 10.1210/jc.2003-030297Cao WB, 2003, AM J PHYSIOL-GASTR L, V285, pG86, DOI 10.1152/ajpgi.00156.2002Polosukhina D, 2003, AM J NEPHROL, V23, P380, DOI 10.1159/000073984Stork PJS, 2002, TRENDS CELL BIOL, V12, P258Vrees MD, 2002, ARCH SURG-CHICAGO, V137, P439Pai R, 2002, NAT MED, V8, P289Xiao ZL, 2002, AM J PHYSIOL-GASTR L, V282, pG300Pearson G, 2001, ENDOCR REV, V22, P153Yang ZW, 1999, N-S ARCH PHARMACOL, V360, P646Narumiya S, 1999, PHYSIOL REV, V79, P1193Tabet F, 2005, J HYPERTENS, V23, P2005Cheng L, 2005, GASTROENTEROLOGY, V129, P1675, DOI 10.1053/j.gastro.2005.09.008Cao W, 2005, AM J PHYSIOL-CELL PH, V289, pC1408, DOI 10.1152/ajpcell.00073.2005Faussone-Pellegrini MS, 2006, J CELL MOL MED, V10, P20Choi S, 2006, CELL PHYSIOL BIOCHEM, V18, P187Choi S, 2006, LIFE SCI, V78, P2322, DOI 10.1016/j.lfs.2005.09.032Popescu LM, 2006, EUR J PHARMACOL, V546, P177, DOI 10.1016/j.ejphar.2006.06.068Yin JY, 2006, DIGEST DIS SCI, V51, P1818, DOI 10.1007/s10620-006-9313-zCao WB, 2006, AM J PHYSIOL-GASTR L, V291, pG672, DOI 10.1152/ajpgi.00110.2006OZAKI H, 2005, INFLAMMOPHARMACOLOGY, V13, P103Jun JY, 2005, BRIT J PHARMACOL, V144, P242, DOI 10.1038/sj.bjp.0706074Daniel EE, 2005, NEUROGASTROENT MOTIL, V17, P355, DOI 10.1111/j.1365-2982.2005.00639.xKataoka K, 2005, J GASTROENTEROL, V40, P610, DOI 10.1007/s00535-005-1595-yLuttrell LM, 1999, CURR OPIN CELL BIOL, V11, P177Hackel PO, 1999, CURR OPIN CELL BIOL, V11, P184Lu G, 1999, GASTROENTEROLOGY, V116, P884Koh SD, 1998, J PHYSIOL-LONDON, V513, P203Zhang JH, 1998, AM J RESP CELL MOL, V19, P324Thomsen L, 1998, NAT MED, V4, P848Sanders KM, 1998, AM J PHYSIOL-GASTR L, V275, pG1Abe MK, 1998, AM J RESP CELL MOL, V18, P562Lu G, 1997, AM J PHYSIOL-GASTR L, V273, pG1233Myers BS, 1997, AM J PHYSIOL-GASTR L, V273, pG928Hawes BE, 1996, J BIOL CHEM, V271, P12133GOLDHILL JM, 1995, AM J PHYSIOL-GASTR L, V268, pG823HUIZINGA JD, 1995, NATURE, V373, P347CROSS DAE, 1994, BIOCHEM J, V303, P21WARD SM, 1994, J PHYSIOL-LONDON, V480, P91GRISHAM MB, 1994, LANCET, V344, P859ZHANG L, 1994, AM J PHYSIOL, V267, pG494BLENNERHASSETT MG, 1992, AM J PHYSIOL, V262, pG1041REDDY SN, 1991, GASTROENTEROLOGY, V101, P1289