BACKGROUND: A correlation between a T-type voltage activated calcium channel (VACC) and pain mechanism has not yet been established. The purpose of this study is to find out the effect of ethosuximide and mibefradil, representative selective T-type VACC blockers on postoperative pain using an incisional pain model of rats. METHODS: After performing a plantar incision, rats were stabilized on plastic mesh for 2 hours. Then, the rats were injected with ethosuximide or mibefradil, intraperitoneally and intrathecally. The level of withdrawal threshold to the von Frey filament near the incision site was determined and the dose response curves were obtained. RESULTS: After an intraperitoneal ethosuximide or mibefradil injection, the dose-response curve showed a dose-dependent increase of the threshold in a withdrawal reaction. After an intrathecal injection of ethosuximide, the threshold of a withdrawal reaction to mechanical stimulation increased and the increase was dose-dependent. After an intrathecal injection of mibefradil, no change occurred in either the threshold of a withdrawal reaction to mechanical stimulation or a dose-response curve. CONCLUSIONS: The T-type VACC blockers in a rat model of postoperative pain showed the antihyperalgesic effect. This effect might be due to blockade of T-type VACC, which was distributed in the peripheral nociceptors or at the supraspinal level. Further studies of the effect of T-type VACC on a pain transmission mechanism at the spinal cord level would be needed.
Animals ; Calcium Channel Blockers* ; Calcium Channels* ; Calcium* ; Ethosuximide* ; Injections, Spinal ; Mibefradil* ; Models, Animal* ; Nociceptors ; Pain, Postoperative* ; Plastics ; Rats* ; Spinal Cord

OBJECTIVE : The sperm acrosome reaction is a Ca2+ -dependent exocytotic event that is triggered by adhesion to the mammalian egg's zona pellucida. Previous studies suggested a role of Ca2+ channels in acrosome reactions. This study was conducted to investigate the T-type calcium channel is operated in acrosome reaction of human spermatozoa. METHOD : Human semen samples were obtained from healthy donors with nomal criteria. The spermatozoa were divided into five groups: Group 1 were non-treated as a control; Group 2 where spermatozoa were exposed to 5 micrometer Ca2+ A23187 (Ca2+i); Group 3 where spermatozoa were exposed 5 micrometer Ca2+i and mibefradil; Group 4 where spermatozoa were exposed 5 micrometer Ca2+i and nifedipine, and Group 5 where spermatozoa were treated with 5 micrometer Ca2+i and both of mibefradil and nifedipine. Spermatozoa in all groups were retrieved after incubation for 15 and 30 minutes at 37degrees C. After staining with PSA-FITC, fluorescence was observed under a fluorescence microscope, and AR was evaluated on a total >100 spermatozoa/side. RESULT AND CONCLUSION : We observed on acrosome reaction inhibition rate in human spermatozoa the various of concentration of mibefradil, nifedipine. Maximum response was noted with 1.0 micrometer mibefradil and the decrease of acrosome reaction inhibition rate 45%. Nifedipine in acrosome reaction inhibition rate was only about 25%. The Ca2+i-induced AR of spermatozoa was significantly suppressed by mibefradil. Incidence of the suppression was depending on concentration of mibefradil. Results from the present study suggest that the human spermatozoa possess T-type channel. The observation that reversible inhibitor of T channels in male germ cells provides a new mechanism of contraceptive action.
Acrosome Reaction* ; Acrosome* ; Calcimycin ; Calcium Channels, T-Type ; Fluorescence ; Germ Cells ; Humans* ; Incidence* ; Male ; Mibefradil* ; Nifedipine ; Semen ; Spermatozoa* ; Tissue Donors ; Zona Pellucida

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.
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

OBJECTIVE: To observe the effect of mibefradil on skeletal muscle mass, function and structure in obese mice. METHODS: Fifteen 6-week-old C57BL/6 mice were randomized equally into normal diet group (control group), high-fat diet (HFD) group and high-fat diet +mibefradil intervention group (HFD +Mibe group). The grip strength of the mice was measured using an electronic grip strength meter, and the muscle content of the hindlimb was analyzed by X-ray absorptiometry (DXA). Triglyceride (TG) and total cholesterol (TC) levels of the mice were measured with GPO-PAP method. The cross-sectional area of the muscle fibers was observed with HE staining. The changes in the level of autophagy in the muscles were detected by Western blotting and immunofluorescence assay, and the activation of the Akt/mTOR signaling pathway was detected with Western blotting. RESULTS: Compared with those in the control group, the mice in HFD group had a significantly greater body weight, lower relative grip strength, smaller average cross sectional area of the muscle fibers, and a lower hindlimb muscle ratio (P < 0.05). Immunofluorescence assay revealed a homogenous distribution of LC3 emitting light red fluorescence in the cytoplasm in the muscle cells in HFD group and HFD+Mibe group, while bright spots of red fluorescence were detected in HFD group. In HFD group, the muscular tissues of the mice showed an increased expression level of LC3 II protein with lowered expressions of p62 protein and phosphorylated AKT and mTOR (P < 0.05). Mibefradil treatment significantly reduced body weight of the mice, lowered the expression level of p62 protein, and increased forelimb grip strength, hindlimb muscle ratio, cross-sectional area of the muscle fibers, and the expression levels of LC3 II protein and phosphorylated AKT and mTOR (P < 0.05). CONCLUSION Mibefradil treatment can moderate high-fat diet-induced weight gain and improve muscle mass and function in obese mice possibly by activating AKT/mTOR signal pathway to improve lipid metabolism and inhibit obesityinduced autophagy.
Animals ; Body Weight ; Diet, High-Fat ; Mibefradil/metabolism* ; Mice ; Mice, Inbred C57BL ; Mice, Obese ; Muscle, Skeletal/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; TOR Serine-Threonine Kinases/metabolism*

Voltage dependent calcium channel (VDCC), one of the most important regulator of Ca2+ concentration in neuron, play an essential role in the central processing of nociceptive information. The present study investigated the antinociceptive effects of L, T or N type VDCC blockers on the formalin-induced orofacial inflammatory pain. Experiments were carried out on adult male Sprague-Dawley rats weighing 220-280 g. Anesthetized rats were individually fixed on a stereotaxic frame and a polyethylene (PE) tube was implanted for intracisternal injection. After 72 hours, 5% formalin (50 microL) was applied subcutaneously to the vibrissa pad and nociceptive scratching behavior was recorded for nine successive 5 min intervals. VDCC blockers were administered intracisternally 20 minutes prior to subcutaneous injection of formalin into the orofacial area. The intracisternal administration of 350 or 700 microg of verapamil, a blocker of L type VDCC, significantly decreased the number of scratches and duration in the behavioral responses produced by formalin injection. Intracisternal administration of 75 or 150 microg of mibefradil, a T type VDCC blocker, or 11 or 22 microg of cilnidipine, a N type VDCC blocker, also produced significant suppression of the number of scratches and duration of scratching in the first and second phase. Neither intracisternal administration of all VDCC blockers nor vehicle did not affect in motor dysfunction. The present results suggest that central VDCCs play an important role in orofacial nociceptive transmission and a targeted inhibition of the VDCCs is a potentially important treatment approach for inflammatory pain originating in the orofacial area.
Adult ; Animals ; Calcium ; Calcium Channel Blockers ; Calcium Channels ; Calcium Channels, L-Type ; Calcium Channels, N-Type ; Calcium Channels, T-Type ; Dihydropyridines ; Facial Pain ; Formaldehyde ; Humans ; Injections, Subcutaneous ; Male ; Mibefradil ; Neurons ; Pain Measurement ; Polyethylene ; Rats ; Rats, Sprague-Dawley ; Verapamil