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The possible roles of hyperpolarization-activated cyclic nucleotide channels in regulating pacemaker activity in colonic interstitial cells of Cajal.

Shahi PK, Choi S, Zuo DC, Kim MY, Park CG, Kim YD, Lee J, Park KJ, So I, Jun JY - J. Gastroenterol. (2013)

Bottom Line: CsCl, ZD7288, zatebradine, clonidine (HCN channel blockers), and genistein (a tyrosine kinase inhibitor) suppressed the pacemaker activity.In recordings of spontaneous intracellular Ca(2+) [Ca(2+)]i oscillations, rolipram and 8-bromo-cAMP increased [Ca(2+)]i oscillations, whereas SQ-22536, CsCl, ZD7288, and genistein decreased [Ca(2+)]i oscillations.HCN channels in colonic ICCs are tonically activated by basal cAMP production and participate in regulation of pacemaking activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, College of Medicine, Chosun University, Sesuk-dong, Dong-gu, Gwangju, 501-759, Korea.

ABSTRACT

Background: Hyperpolarization-activated cyclic nucleotide (HCN) channels are pacemaker channels that regulate heart rate and neuronal rhythm in spontaneously active cardiac and neuronal cells. Interstitial cells of Cajal (ICCs) are also spontaneously active pacemaker cells in the gastrointestinal tract. Here, we investigated the existence of HCN channel and its role on pacemaker activity in colonic ICCs.

Methods: We performed whole-cell patch clamp, RT-PCR, and Ca(2+)-imaging in cultured ICCs from mouse mid colon.

Results: SQ-22536 and dideoxyadenosine (adenylate cyclase inhibitors) decreased the frequency of pacemaker potentials, whereas both rolipram (cAMP-specific phosphodiesterase inhibitor) and cell-permeable 8-bromo-cAMP increased the frequency of pacemaker potentials. CsCl, ZD7288, zatebradine, clonidine (HCN channel blockers), and genistein (a tyrosine kinase inhibitor) suppressed the pacemaker activity. RT-PCR revealed expression of HCN1 and HCN3 channels in c-kit and Ano1 positive colonic ICCs. In recordings of spontaneous intracellular Ca(2+) [Ca(2+)]i oscillations, rolipram and 8-bromo-cAMP increased [Ca(2+)]i oscillations, whereas SQ-22536, CsCl, ZD7288, and genistein decreased [Ca(2+)]i oscillations.

Conclusions: HCN channels in colonic ICCs are tonically activated by basal cAMP production and participate in regulation of pacemaking activity.

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The effects of cAMP-related drugs and hyperpolarization-activated cyclic nucleotide (HCN) channel blockers on spontaneous intracellular Ca2+ oscillations in cultured colonic ICCs. Cell permeable 8-bromo-cAMP (100 μM) (a) and rolipram (100 μM) (b) increased the frequency of intracellular Ca2+ oscillations. In contrast SQ-22536 (100 μM) (c), dideoxyadenosine (100 μM) (d), ZD7288 (10 μM) (e) and genistein (10 μM) (f) decreased the frequency of intracellular Ca2+ oscillations
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Fig6: The effects of cAMP-related drugs and hyperpolarization-activated cyclic nucleotide (HCN) channel blockers on spontaneous intracellular Ca2+ oscillations in cultured colonic ICCs. Cell permeable 8-bromo-cAMP (100 μM) (a) and rolipram (100 μM) (b) increased the frequency of intracellular Ca2+ oscillations. In contrast SQ-22536 (100 μM) (c), dideoxyadenosine (100 μM) (d), ZD7288 (10 μM) (e) and genistein (10 μM) (f) decreased the frequency of intracellular Ca2+ oscillations

Mentions: In cultured intestinal ICCs, the activity of pacemaker channels was found to be closely coupled with spontaneous [Ca2+]i oscillations [31]. Thus, to evaluate whether HCN channels activation is coupled with spontaneous [Ca2+]i oscillations, we measured spontaneous [Ca2+]i oscillations in cultured colonic ICCs that are connected with cell cluster and treated with 8-bromo-cAMP (100 μM; n = 6), rolipram (100 μM; n = 6), SQ-22536 (100 μM; n = 6), CsCl (5 mM; n = 6), ZD7288 (10 μM; n = 6), and genistein (10 μM; n = 4). ICCs were loaded with fluo3-AM and spontaneous [Ca2+]i oscillations were observed in a time series. The frequency of [Ca2+]i oscillations was 7.3 ± 1.6 cycles/5 min in the control (n = 34). Both 8-bromo-cAMP and rolipram increase spontaneous [Ca2+]i oscillations (Fig. 6a, b). In contrast, SQ-22536, CsCl, ZD7288, and genistein all decreased spontaneous [Ca2+]i oscillations (Fig. 6c–f). The values of the frequency induced by the above drugs were significantly different from control values. These results suggested that the basal activation of HCN channels may be coupled with spontaneous [Ca2+]i oscillations (Fig. 6g).Fig. 6


The possible roles of hyperpolarization-activated cyclic nucleotide channels in regulating pacemaker activity in colonic interstitial cells of Cajal.

Shahi PK, Choi S, Zuo DC, Kim MY, Park CG, Kim YD, Lee J, Park KJ, So I, Jun JY - J. Gastroenterol. (2013)

The effects of cAMP-related drugs and hyperpolarization-activated cyclic nucleotide (HCN) channel blockers on spontaneous intracellular Ca2+ oscillations in cultured colonic ICCs. Cell permeable 8-bromo-cAMP (100 μM) (a) and rolipram (100 μM) (b) increased the frequency of intracellular Ca2+ oscillations. In contrast SQ-22536 (100 μM) (c), dideoxyadenosine (100 μM) (d), ZD7288 (10 μM) (e) and genistein (10 μM) (f) decreased the frequency of intracellular Ca2+ oscillations
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Related In: Results  -  Collection

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Fig6: The effects of cAMP-related drugs and hyperpolarization-activated cyclic nucleotide (HCN) channel blockers on spontaneous intracellular Ca2+ oscillations in cultured colonic ICCs. Cell permeable 8-bromo-cAMP (100 μM) (a) and rolipram (100 μM) (b) increased the frequency of intracellular Ca2+ oscillations. In contrast SQ-22536 (100 μM) (c), dideoxyadenosine (100 μM) (d), ZD7288 (10 μM) (e) and genistein (10 μM) (f) decreased the frequency of intracellular Ca2+ oscillations
Mentions: In cultured intestinal ICCs, the activity of pacemaker channels was found to be closely coupled with spontaneous [Ca2+]i oscillations [31]. Thus, to evaluate whether HCN channels activation is coupled with spontaneous [Ca2+]i oscillations, we measured spontaneous [Ca2+]i oscillations in cultured colonic ICCs that are connected with cell cluster and treated with 8-bromo-cAMP (100 μM; n = 6), rolipram (100 μM; n = 6), SQ-22536 (100 μM; n = 6), CsCl (5 mM; n = 6), ZD7288 (10 μM; n = 6), and genistein (10 μM; n = 4). ICCs were loaded with fluo3-AM and spontaneous [Ca2+]i oscillations were observed in a time series. The frequency of [Ca2+]i oscillations was 7.3 ± 1.6 cycles/5 min in the control (n = 34). Both 8-bromo-cAMP and rolipram increase spontaneous [Ca2+]i oscillations (Fig. 6a, b). In contrast, SQ-22536, CsCl, ZD7288, and genistein all decreased spontaneous [Ca2+]i oscillations (Fig. 6c–f). The values of the frequency induced by the above drugs were significantly different from control values. These results suggested that the basal activation of HCN channels may be coupled with spontaneous [Ca2+]i oscillations (Fig. 6g).Fig. 6

Bottom Line: CsCl, ZD7288, zatebradine, clonidine (HCN channel blockers), and genistein (a tyrosine kinase inhibitor) suppressed the pacemaker activity.In recordings of spontaneous intracellular Ca(2+) [Ca(2+)]i oscillations, rolipram and 8-bromo-cAMP increased [Ca(2+)]i oscillations, whereas SQ-22536, CsCl, ZD7288, and genistein decreased [Ca(2+)]i oscillations.HCN channels in colonic ICCs are tonically activated by basal cAMP production and participate in regulation of pacemaking activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, College of Medicine, Chosun University, Sesuk-dong, Dong-gu, Gwangju, 501-759, Korea.

ABSTRACT

Background: Hyperpolarization-activated cyclic nucleotide (HCN) channels are pacemaker channels that regulate heart rate and neuronal rhythm in spontaneously active cardiac and neuronal cells. Interstitial cells of Cajal (ICCs) are also spontaneously active pacemaker cells in the gastrointestinal tract. Here, we investigated the existence of HCN channel and its role on pacemaker activity in colonic ICCs.

Methods: We performed whole-cell patch clamp, RT-PCR, and Ca(2+)-imaging in cultured ICCs from mouse mid colon.

Results: SQ-22536 and dideoxyadenosine (adenylate cyclase inhibitors) decreased the frequency of pacemaker potentials, whereas both rolipram (cAMP-specific phosphodiesterase inhibitor) and cell-permeable 8-bromo-cAMP increased the frequency of pacemaker potentials. CsCl, ZD7288, zatebradine, clonidine (HCN channel blockers), and genistein (a tyrosine kinase inhibitor) suppressed the pacemaker activity. RT-PCR revealed expression of HCN1 and HCN3 channels in c-kit and Ano1 positive colonic ICCs. In recordings of spontaneous intracellular Ca(2+) [Ca(2+)]i oscillations, rolipram and 8-bromo-cAMP increased [Ca(2+)]i oscillations, whereas SQ-22536, CsCl, ZD7288, and genistein decreased [Ca(2+)]i oscillations.

Conclusions: HCN channels in colonic ICCs are tonically activated by basal cAMP production and participate in regulation of pacemaking activity.

Show MeSH
Related in: MedlinePlus