5-Hydroxytryptamine Generates Tonic Inward Currents on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Pawan Kumar Shahi; Seok Choi; Dong Chuan Zuo; Cheol Ho Yeum; Pyung Jin Yoon; Jun Lee; Young Dae Kim; Chan Guk Park; Man Yoo Kim; Hye Rang Shin; Hyun Jung Oh; Jae Yeoul Jun

doi:10.4196/kjpp.2011.15.3.129

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Shahi, Choi, Zuo, Yeum, Yoon, Lee, Kim, Park, Kim, Shin, Oh, and Jun: 5-Hydroxytryptamine Generates Tonic Inward Currents on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Original Article

Korean J Physiol Pharmacol 2011;15(3):129-135.

Published online: 18 February 2011

DOI: https://doi.org/10.4196/kjpp.2011.15.3.129

5-Hydroxytryptamine Generates Tonic Inward Currents on Pacemaker Activity of Interstitial Cells of Cajal from Mouse Small Intestine

Pawan Kumar Shahi^1,^∗, Seok Choi^1,^∗, Dong Chuan Zuo¹, Cheol Ho Yeum¹, Pyung Jin Yoon¹, Jun Lee², Young Dae Kim², Chan Guk Park², Man Yoo Kim², Hye Rang Shin³, Hyun Jung Oh³, Jae Yeoul Jun^1,

¹Department of Physiology, College of Medicine, Chosun University, Gwangju 501-759, Korea

²Department of Internal Medicine, College of Medicine, Chosun University, Gwangju 501-759, Korea

³Department of Psychiatry, College of Medicine, Chosun University, Gwangju 501-759, Korea

Corresponding to: Jae Yeoul Jun, Department of Physiology, College of Medicine, Chosun University, 375, Seosuk-dong, Gwangju 501-759, Korea. (Tel) 82-62-230-6412, (Fax) 82-62-232-4943, (E-mail) jyjun@chosun.ac.kr

^∗ These authors contributed equally to this work.

Received 8 April 2011 Revised 13 June 2011 Accepted 14 June 2011

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

In this study we determined whether or not 5-hydroxytryptamine (5-HT) has an effect on the pacemaker activities of interstitial cells of Cajal (ICC) from the mouse small intestine. The actions of 5-HT on pacemaker activities were investigated using a whole-cell patch-clamp technique, intracellular Ca²⁺ ([Ca²⁺]_i) analysis, and RT-PCR in ICC. Exogenously-treated 5-HT showed tonic inward currents on pacemaker currents in ICC under the voltage-clamp mode in a dose-dependent manner. Based on RT-PCR results, we found the existence of 5-HT_2B, ₃, ₄, _and ₇ receptors in ICC. However, SDZ 205557 (a 5-HT₄ receptor antagonist), SB 269970 (a 5-HT₇ receptor antagonist), 3-tropanylindole – 3 – carboxylate methiodide (3-TCM; a 5-HT₃ antagonist) blocked the 5-HT-induced action on pacemaker activity, but not SB 204741 (a 5-HT_2B receptor antagonist). Based on [Ca²⁺]_i analysis, we found that 5-HT increased the intensity of [Ca²⁺]_i. The treatment of PD 98059 or JNK II inhibitor blocked the 5-HT-induced action on pacemaker activity of ICC, but not SB 203580. In summary, these results suggest that 5-HT can modulate pacemaker activity through 5-HT₃, ₄, _and ₇ receptors via [Ca²⁺]_i mobilization and regulation of mitogen-activated protein kinases.

Keywords: Interstitial cells of Cajal (ICC), Intestinal motility, Pacemaker currents, 5-hydroxytryptamine (5-HT)

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Fig. 1.

Effects of 5-HT on pacemaker potentials recorded in ICC from mouse small intestine. (A) shows the pacemaker potentials of ICC exposed to 5-HT (10 μM) in the current clamping mode (I=0). 5-HT induced membrane depolarization and inhibited the amplitude and frequency of pacemaker potential in ICC. The dot lines indicate the control resting membrane potentials levels. Responses to 5-HT are summarized in (B) and (C). Bars represent the means±SE. ^∗Asterisks indicate significantly different from controls (p<0.05). RMP, resting membrane potentials.

Fig. 2.

Effects of 5-HT on pacemaker currents recorded in cultured ICC from mouse small intestine. (A), (B) and (C) show pacemaker currents of ICC exposed to 5-HT (1, 10, or 100 μM) at a holding potential of –70 mV. Vertical solid line represents amplitude of pacemaker current and horizontal solid line represents duration of recording(s) pacemaker currents. The dot lines indicate the control resting current levels. (D) and (E) summarize the effects of 5-HT on pacemaker currents in ICC. Bars represent the means±SE. ^∗Asterisks indicate significantly different from controls (p<0.05). Con, control.

Fig. 3.

Agarose gels of the RT-PCR products of 5-HT receptor subtypes using separated ICC. Amplified cDNA prepared from pure ICC separated by magnetic cell separation and visualized in 2% gel. c-Kit – ICC marker, myosin – smooth muscle cells marker, PGP9.5 – neuronal cell marker.

Fig. 4.

Effects of 5-HT receptor subtype antagonists on 5-HT-induced responses on pacemaker currents in cultured ICC of the mouse small intestine. (A) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of 5-HT₄ receptor antagonist (SDZ 205557; 10 μM). (B) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of 5-HT₇ receptor antagonist (SB 269970; 10 μM). (C) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of 5-HT₃ receptor antagonist 3-TCM (10 μM). (D) Pacemaker currents of ICC exposed to the selective 5-HT_2B receptor antagonist SB 204741 (10 μM). Responses to 5-HT in presence of different receptor antagonists are summarized in (E). Bars represent the mean values±S.E. The dotted lines indicate the zero current levels. ^∗Asterisks indicate significantly different from the control (p<0.05). Con, control.

Fig. 5.

Effects of 5-HT on [Ca²⁺]_i oscillation in cultured ICC from mouse small intestine. (A) shows the basal and peak point of ICC image and (B) Ca²⁺ oscillation in the normal condition. (C) shows the basal and peak point of ICC image and (D) Ca²⁺ oscillation in the presence of 5-HT (10 μM). The interval of representative frame was 1 second and the exposure time of each frame was 500 ms.

Fig. 6.

Effects of various MAPK inhibitors on 5-HT induced responses on pacemaker currents in cultured ICC of the mouse small intestine. (A) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of PD 98059, a p42/44 MAPK inhibitor (10 μM). (B) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of SB 203580, a p38 MAPK inhibitor (10 μM). (C) Pacemaker currents of ICC exposed to 5-HT (10 μM) in the presence of JNK II inhibitor (10 μM). Responses to 5-HT in presence of different MAPK inhibitors are summarized in (D). Bars represent mean values±S.E. The dotted lines indicate the zero current levels. ^∗Asterisks indicate significantly different from control (p<0.05). Con, control.

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