Korean J Physiol Pharmacol.  2009 Jun;13(3):241-249. 10.4196/kjpp.2009.13.3.241.

Role of T-type Ca2+ Channels in the Spontaneous Phasic Contraction of Pregnant Rat Uterine Smooth Muscle

• Lee SE
• Ahn DS
• Lee YH

• KMID: 2071677
• DOI: http://doi.org/10.4196/kjpp.2009.13.3.241

Abstract

Although extracellular Ca2+ entry through the voltage-dependent Ca2+ channels plays an important role in the spontaneous phasic contractions of the pregnant rat myometrium, the role of the T-type Ca2+ channels has yet to be fully identified. The aim of this study was to investigate the role of the T-type Ca2+ channel in the spontaneous phasic contractions of the rat myometrium. Spontaneous phasic contractions and [Ca2+]i were measured simultaneously in the longitudinal strips of female Sprague-Dawley rats late in their pregnancy (on day 18~20 of gestation: term=22 days). The expression of T-type Ca2+ channel mRNAs or protein levels was measured. Cumulative addition of low concentrations (<1 micrometer) of nifedipine, a L-type Ca2+ channel blocker, produced a decrease in the amplitude of the spontaneous Ca2+ transients and contractions with no significant change in frequency. The mRNAs and proteins encoding two subunits (alpha1G, alpha1H) of the T-type Ca2+ channels were expressed in longitudinal muscle layer of rat myometrium. Cumulative addition of mibefradil, NNC 55-0396 or nickel induced a concentration-dependent inhibition of the amplitude and frequency of the spontaneous Ca2+ transients and contractions. Mibefradil, NNC 55-0396 or nickel also attenuated the slope of rising phase of spontaneous Ca2+ transients consistent with the reduction of the frequency. It is concluded that T-type Ca2+ channels are expressed in the pregnant rat myometrium and may play a key role for the regulation of the frequency of spontaneous phasic contractions.

Keyword

Calcium channels; Nickel; Mibefradil; NNC 55-0396; Spontaneous contractility

MeSH Terms

Animals
Benzimidazoles
Calcium Channels
Contracts
Cyclopropanes
Female
Humans
Mibefradil
Mice
Muscle, Smooth
Muscles
Myometrium
Naphthalenes
Nickel
Nifedipine
Pregnancy
Proteins
Rats
Rats, Sprague-Dawley
RNA, Messenger
Benzimidazoles
Calcium Channels
Cyclopropanes
Mibefradil
Naphthalenes
Nickel
Nifedipine
Proteins
RNA, Messenger

Figure

• 

  Fig. 1. Effect of removing external Ca2+ and nifedipine on spontaneous Ca2+ transients and contractions. (A, B) Effect of removing external Ca2+ (0-Ca2+) and 1 μM nifedipine on spontaneous Ca2+ transients and contractions. Representative recoding (C) and statistical evaluation (D) showing the concentration-response curve obtained by cumulative addition of low concentrations of nifedipine. Data are expressed as relative percentage of the control (amplitude before treatment of nifedipine). Results are expressed as mean±SEM of six experiments. ∗Control vs Nifedipine (p<0.05).

• 

  Fig. 2. Expression of mRNAs and proteins for α subunits (α1G and α1H) of T-type Ca2+ channel in longitudinal muscle layer of pregnant rat myometrium. Representative data of RT/PCR (A) and western blot (B). Immunoblots are representative of four independent preparations. The PCR was performed with 35 cycles and PCR products were followed by electrophoresis on a 1% agarose gel.

• 

  Fig. 3. Dose-response curve for the effect of T-type Ca2+ channel blockers on the spontaneous Ca2+ transients and contractions. (A1, B1, C1) Concentration-related reduction of the amplitude and frequency of spontaneous Ca2+ transients. (A2, B2, C2) concentration-related reduction of the amplitude and frequency of spontaneous contractions. Mibefradil (A), NNC 55-0396 (B), and nickel (C) were added cumulatively. Data are expressed as relative percentage of control (amplitude before treatment of blockers). Results are expressed as mean±SEM of seven experiments. ∗Control vs Blockers, ∗∗Amplitude vs Frequency (p<0.05).

• 

  Fig. 4. Representative recording for the effect of T-type Ca2+ channel blockers on spontaneous Ca2+ transients and contractions. Spontaneous Ca2+ transients (top) and contractions (bottom) before and during 1 μM mibefradil (A), 5 μM NNC 55-0396 (B), and 100 μM nickel (C) application. Data are representative of ten independent preparations.

• 

  Fig. 5. Effect of the T-type Ca2+ channel blockers on the frequency and slope of rising phase of spontaneous Ca2+ transients. Superimposed spontaneous Ca2+ transients under control conditions and in presence of 1 μM Mibefradil (A), 5 μM NNC 55-0396 (B), and 100 μM nickel (C), respectively. Each blocker was added in the bath solution after spontaneous Ca2+ transients were stable. Statistical evaluation (D) showing the Ca2+ ratio and frequency obtained by adding mibefradil, NNC 55-0396, and nifedipine. Data are expressed as relative percentage of control. Results are expressed as mean±SEM of six experiments.

• 

  Fig. 6. Effect of (A) nifedipine (1 μM), (B) mibefradil (1 μM), (C) NNC 55-0396 (5 μM), and (D) nickel (100 μM) on the 70 mM KCl-induced increase in [Ca2+]i and force. All drugs were added for 10 min before 70 mM KCl-induced contraction. Data are representative of seven~nine independent preparations.

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