J Korean Med Sci.  2007 Feb;22(1):48-56. 10.3346/jkms.2007.22.1.48.

Effects of Polyamines on Contractility of Guinea-Pig Gastric Smooth Muscle

• Kim YC
• Sim JH
• Kim YH
• Kwon SC
• Lee SJ
• Kim SR
• Kim DW
• Park SM
• Youn SJ
• Lee SJ
• Xing DG
• Xu WX
• Kim KW

• KMID: 1713229
• DOI: http://doi.org/10.3346/jkms.2007.22.1.48

Abstract

This study was designed to investigate the effects of polyamines on mechanical contraction and voltage-dependent calcium current (VDCC) of guinea-pig gastric smooth muscle. Mechanical contraction and calcium channel current (I(Ba)) were recorded by isometric tension recording and whole-cell patch clamp technique. Spermine, spermidine and putrescine inhibited spontaneous contraction of the gastric smooth muscle in a concentration-dependent manner. Spermine (2 mM) reduced high K+ (50 mM)-induced contraction to 16+/-6.4% of the control (n=9), and significantly inhibited I(Ba) in a reversible manner (p<0.05; IC50=0.8 mM). Pre- and post-treatment of tissue with spermine (2-5 mM, n=10) also inhibited acetylcholine (10 micrometer)-induced phasic contraction to 5+/-6.4% of the control. Inhibitory effect of spermine on I(Ba) was observed at a wide range of test potentials of current/voltage (I/V) relationship (p<0.05), and steady-state activation of I(Ba) was shifted to the right by spermine (p<0.05). Spermidine and putrescine (1 mM each) also inhibited I(Ba) to 51+/-5.7% and 81+/-5.3% of the control, respectively. And putrescine (1 mM) inhibited I(Ba) at whole tested potentials (p<0.05) without significant change of kinetics (p<0.05). Finally, 5 mM putrescine also inhibited high K+ -induced contraction to 53+/-7.1% of the control (n=4). These findings suggest that polyamines inhibit contractions of guinea-pig gastric smooth muscle via inhibition of VDCC.

Keyword

Muscle, Smooth; myocytes, Smooth Muscle; Gastrointestinal Tract; Calcium Current; Polyamines; Spermine; Spermidine; Putrescine

MeSH Terms

Pyloric Antrum/*drug effects/physiology
Potassium/pharmacology
Polyamines/*pharmacology
Muscle, Smooth/*drug effects/physiology
Muscle Contraction/*drug effects
Male
Guinea Pigs
Female
Calcium Channels/drug effects/physiology
Calcium/metabolism
Animals

Figure

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  Fig. 1 Structures of the endogenous polyamines. The structures of polyamines putrescine, spermidine and spermine are shown in A-C.

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  Fig. 2 Effect of polyamines on spontaneous contraction in guinea-pig gastric smooth muscle. Effects of spermine and putrescine on isometric contraction were studied by using vertical chamber system. Spermine produces relaxation in a concentration-dependent manner (A). In (B) and (C), concentration-response relationship of spermine and putrescine are summarized. Relative contractions at various concentrations of spermine and putrescine are plotted and fitted by the non-linear regression equation.

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  Fig. 3 Relationship between polyamines-induced relaxation and Ca2+ influx through VDCC. High K+ solution (50 mM) in the presence of nerve blockers produced biphasic contraction in guinea-pig gastric smooth muscle. Such high K+ (50 mM)-induced contraction is reduced by pretreatment of the cells with 2 mM spermine in (A). Pannel (B) shows summarized data of all tissues tested. In (C), Inhibitory effect of putrescine (5 mM) on high K+ (50 mM)-induced contraction is also summarized.

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  Fig. 4 Inhibitory effect of spermine on acetylchole (ACh)-induced contraction through modulation of VDCC. (A) 10 µM ACh produces transient initial and tonic contraction which is superimposed by phasic contraction in a reversible manner. ACh-induced contraction are summarized in (B). ACh-induced phasic contraction is inhibited by nicardipine 2 µM in (C). Pannel (D) shows summarized data of all tissues tested. Effect of spermine on ACh-induced phasic contraction was also studied. Spermine (2 mM) inhibited ACh-induced phasic contraction in (E). And (F) shows summarized data of all tissues tested.

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  Fig. 5 Inhibition of ACh-induced contraction by spermine in guinea-pig gastric smooth muscle. (A) Repetitive application of ACh (10 µM) in guinea-pig gastric smooth muscle produces contraction. Pretreatment of the muscle with 5 mM spermine reduced ACh-induced initial contraction and inhibited phasic contraction in a reversible manner. Pannel (B) shows summarized data of all tissues tested.

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  Fig. 6 Effect of polyamines on IBa in guinea-pig gastric myocytes. IBa was recorded under the condition in which extracellular Ca2+ was replaced by 10 mM Ba2+. IBa is inhibited by the application of spermine in (A) and (B). In (C), effects of polyamines (1 mM) on IBa at 0 mV were studied. IBa is decreased by the application of 1 mM each of spemine, spermidine and putrescine. (D, E) Concentration-response relationship of spermine and putrescine on IBa is summarized. Normalized responses by various concentrations of spermine and putrescine against the peak values obtained at 0 mV are plotted and fitted by the non-linear regression equation.

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  Fig. 7 Effects of polyamines on I/V relationship and kinetics of IBa in guinea-pig gastric myocytes. In (A) and (B), I/V relationship of IBa by spermine and putrescine in guinea-pig gastric myocytes are shown. Averaged responses of IBa in the presence and absence of spermine and putrescine are plotted. (C) and (D) show steady-state activation and inactivation curves for the cells exposed to none and polyamines. In (C), steady-state activation and inactivation curves for the cells exposed to none (▴,●) and spermine (▵,○) are shown. Spermine shifted values of half-activation to rightward (n=5, p<0.05). (D) shows steady-state activation and inactivation curves for the cells exposed to none (▴,●) and putrescine (▵,○). There is no significant difference in values of half-activation and half-inactivation (n=5, p>0.05).

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