Picosecond Laser-Induced Photothermal Skin Damage Evaluation by Computational Clinical Trial.

BACKGROUND AND OBJECTIVES: Computational clinical trial (CCT) in the field of laser medicine promotes clinical application of novel laser devices, because this trial carried out based on numerical modeling of laser-tissue interactions and simulation of a series of treatment process. To confirm the feasibility of the computational clinical trial of skin treatment with a novel picosecond laser, this paper presents an evaluation method of the safety. STUDY DESIGN/MATERIALS AND METHODS: In this method, the light propagation and thermal diffusion process after ultrashort light pulse irradiation to a numerical skin model is calculated and the safety based on the photothermal damage is evaluated by computational modeling and simulation. As an example, the safety of a novel picosecond laser device was examined by comparing with several laser devices approved for clinical use. RESULTS: The ratio of the maximum thermal damage induced by picosecond laser irradiation was 1.2 × 10-2 % at the epidermis, while that caused by approved laser irradiation was 99 % at the capillary vessels. The numerical simulation demonstrated that less thermal damage was observed compared with the approved devices. The results show the safety simulated by photothermal damage calculation was consistent with the reported clinical trials. CONCLUSIONS: This computational clinical trial shows the feasibility of applying computational clinical trials for the safety evaluation of novel medical laser devices. In contrast to preclinical and clinical tests, the proposed computational method offers regulatory science for appropriately and quickly predicting and evaluating the safety of a novel laser device.

Effect of caffeine on mucus secretion and agonist-dependent Ca2+ mobilization in human gastric mucus secreting cells.

Caffeine is known to stimulate gastric acid secretion, but, the effects of caffeine on gastric mucus secretion have not been clarified. To elucidate the action of caffeine on gastric mucin-producing cells and its underlying mechanism, the effects of caffeine on mucus glycoprotein secretion and agonist-induced [Ca2+]i mobilization were examined in human gastric mucin secreting cells (JR-I cells). The measurement of [Ca2+]i using Indo-1 and the whole cell voltage clamp technique were applied. Mucus glycoprotein secretion was assessed by release of [3H]glucosamine. Caffeine by itself failed to increase [Ca2+]i and affect membrane currents, while it dose-dependently inhibited agonist (acetylcholine (ACh) or histamine)-induced [Ca2+]i rise, resulting in inhibiting activation of Ca2+-dependent K+ current (I(K.Ca)) evoked by agonists. The effect of caffeine was reversible, and the half maximal inhibitory concentration was about 0.5 mM. But, caffeine did not suppress [Ca2+]i rise and activation of I(K.Ca) induced by A23187 or inositol trisphosphate (IP3). Theophylline or 3-isobutyl-1-methyl-xanthine (IBMX) did not mimic the effect of caffeine. Caffeine failed to stimulate mucus secretion, while it significantly decreased ACh-induced mucus secretion. These results indicate that caffeine selectively inhibits agonist-mediated [Ca2+]i rise in human gastric epithelial cells, probably through the blockade of receptor-IP3 signaling pathway, which may affect the mucin secretion.

Troglitazone inhibits voltage-dependent calcium currents in guinea pig cardiac myocytes.

BACKGROUND: It has been suggested that intracellular Ca2+ overload in cardiac myocytes leads to the development of diabetic cardiomyopathy. Troglitazone, an insulin-sensitizing agent, is a promising therapeutic agent for diabetes and has been shown to prevent diabetes-induced myocardial changes. To elucidate the underlying mechanism of troglitazone action on cardiac myocytes, the effects of troglitazone on voltage-dependent Ca2+ currents were examined and compared with classic Ca2+ antagonists (verapamil and nifedipine). METHODS AND RESULTS: Whole-cell voltage-clamp techniques were applied in single guinea pig atrial myocytes. Under control conditions with CsCl internal solution, the voltage-dependent Ca2+ currents consisted of both T-type (ICa,T) and L-type (ICa,L) Ca2+ currents. Troglitazone effectively reduced the amplitude of ICa,L in a concentration-dependent manner. Troglitazone also suppressed ICa,T, but the effect of troglitazone on ICa,T was less potent than that on ICa,L. The current-voltage relationships for ICa,L and the reversal potential for ICa,L were not altered by troglitazone. The half-maximal inhibitory concentration of troglitazone on ICa,L measured at a holding potential of -40 mV was 6.3 micromol/L, and 30 micromol/L troglitazone almost completely inhibited ICa,L. Troglitazone 10 micromol/L did not affect the time courses for inactivation of ICa,L and inhibited ICa,L mainly in a use-independent fashion, without shifting the voltage-dependency of inactivation. This effect was different from those of verapamil and nifedipine. Troglitazone also reduced isoproterenol- or cAMP-enhanced ICa,L. CONCLUSIONS: These results demonstrate that troglitazone inhibits voltage-dependent Ca2+ currents (T-type and L-type) and then antagonizes the effects of isoproterenol in cardiac myocytes, thus possibly playing a role in preventing diabetes-induced intracellular Ca2+ overload and subsequent myocardial changes.

Ouabainlike compound in hypertension associated with ectopic corticotropin syndrome.

Molecular mechanisms related to sodium retention have been implicated in the pathogenesis of hypertension. It is unclear how sodium retention leads to a rise in blood pressure, but ouabainlike compound may act as a final common pathway in sodium-induced hypertension. In ectopic corticotropin syndrome, hypertension has been attributed to cortisol inactivation overload, giving rise to mineralocorticoid-type hypertension. We sequentially measured plasma and urinary levels of ouabainlike compound over 2 months to evaluate its role in the hypertensive mechanisms in a 64-year-old man with this syndrome caused by lung cancer. His data included hypokalemia and increased cortisol concentrations, corticotropin levels, and urinary 17-hydroxycorticosteroid excretion. Plasma renin activity was suppressed. Plasma and urinary levels of ouabainlike compound were markedly increased concomitantly with high blood pressure. The maximum plasma level was 40-fold the normal range of the subject. After chemotherapy, ouabainlike compound levels gradually decreased in parallel with the decline in blood pressure and rise in potassium concentration. A correlation was observed between plasma and urinary levels of ouabainlike compound (P < .05). Plasma and urinary levels of ouabainlike compound correlated with systolic (P < .01) and diastolic (P < .05) pressures, respectively. The peak of ouabainlike compound in plasma and urine coincided with that of authentic ouabain on high-performance liquid chromatography. Ouabainlike compound derived from urine inhibited [3H]ouabain binding to human erythrocytes. These findings suggest that ouabainlike compound with biological activity could partly account for hypertension in ectopic corticotropin syndrome.

Periodicity of episode occurrences in rapid cycling affective disorders.

We analyzed the medical records of nine patients with severe rapid cycling affective disorders (RCAD), and determined the cycle of mania occurrences by calculating the period between two successive onsets of mania for each patient. Using the cycle as the index we devised a cycle-oriented diagram by dividing the observation period by the index cycle, and visually studied the mode of episode occurrences. In seven patients, the onset of mania generally followed the index cycle, but sometimes shifted from the day estimated by the index cycle. The shift seemed to be caused by a cycle other than the index cycle, which appeared temporarily in the course. The period between two successive onsets of mania was often several times longer than the index cycle length. In patients with RCAD, some sort of periodic rhythms may possibly control the course of episode occurrences on a continuing basis.

Influence of cellular incorporation of n-3 eicosapentaenoic acid on intracellular Ca2+ concentration and membrane potential in vascular smooth muscle cells.

Long-term treatment with n-3 eicosapentaenoic acid (EPA) has been shown to exert hypotensive effects and have beneficial effects on atherosclerosis. To elucidate one of the underlying mechanisms of these effects, intracellular calcium concentration [Ca2+]i, and resting membrane potential were measured in rat vascular smooth muscle cells (A7r5 cell) treated with EPA, using Ca2+-sensitive dye fura-2 AM and the patch clamp technique. The alterations in fatty acid compositions of phospholipids and cell migration after treatment with EPA (30 microM) for 6 h-7 days were also examined. After treating cells with EPA, the EPA and DPA (docosapentaenoic acid) content of the phospholipid fraction (mol.%) increased in a time-dependent manner. Alternatively, arachidonic acid (AA) decreased, and then the ratio of EPA and AA (EPA/AA) increased significantly. The resting [Ca2+]i decreased from 170 +/- 46 nM (n = 16) in control cells to 123 +/- 29 nM (n = 16) in cells treated with EPA (30 microM) for 7 days. Vasopressin (100 nM), endothelin-1 (100 nM) and platelet-derived growth factor (PDGF 5 ng/ml) evoked an initial peak of [Ca2+]i, followed by a smaller sustained rise of [Ca2+]i in the presence of extracellular Ca2+. In EPA-treated cells, both the peak and the sustained rise of [Ca2+]i induced by these agonists decreased in comparison to the control cells. EPA treatment also decreased the transient [Ca2+]i rise evoked by these agonists in the absence of extracellular Ca2+. Under the current clamp condition, resting membrane potential was significantly higher in EPA-treated cells (-49.8 +/- 10.4 mV, n = 41) than in control cells (-44.6 +/- 7.4 mV, n = 41, P < 0.05), and the input resistance of the cell was lower in EPA-treated cells, while cell size and capacitance were not statistically different. In addition, long-term treatment with EPA for 7 days significantly inhibited PDGF-induced cell migration. These results suggest that cellular incorporation of n-3 eicosapentaenoic acid attenuates intracellular mechanisms related to changes of [Ca2+]i and affects membrane potential, thereby inhibiting migration of vascular smooth muscle cells. These actions of EPA may contribute to its vasorelaxant and antiatherosclerotic effects.

Inhibitory effects of omega-3 polyunsaturated fatty acids on receptor-mediated non-selective cation currents in rat A7r5 vascular smooth muscle cells.

1. The effects of omega-3 polyunsaturated fatty acids on receptor-mediated non-selective cation current (Icat) and K+ current were investigated in aortic smooth muscle cells from foetal rat aorta (A7r5 cells). The whole-cell voltage clamp technique was employed. 2. With a K(+)-containing solution, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA, 30 microM) produced an outward current at a holding potential of -40 mV. This response was inhibited by tetraethylammonium (20 mM) or Cs+ in the patch pipette solution, and the reversal potential of the EPA-induced current followed the K+ equilibrium potential in a near Nernstian manner. 3. Under conditions with a Cs(+)-containing pipette solution, both vasopressin and endothelin-1 (100 nM) induced a long-lasting inward current at a holding potential of -60 mV. The reversal potential of these agonist-induced currents was about +0 mV, and was not significantly altered by the replacement of the extracellular or intracellular Cl+ concentration, suggesting that the induced current was a cation-selective current (Icat). 4. La3+ and Cd2+ (1 mM) completely abolished these agonist-induced Icat, but nifedipine (10 microM) failed to inhibit it significantly. 5. omega-3 polyunsaturated fatty acids (3-100 microM), EPA, DHA and docosapentaenoic acids (DPA), inhibited the agonist-induced Icat in a concentration-dependent manner. The potency of the inhibitory effect was EPA > DHA > DPA, and the half maximal inhibitory concentration (IC50) of EPA was about 7 microM. 6. Arachidonic and linoleic acids (10, 30 microM) showed a smaller inhibitory effect compared to omega-3 fatty acids. Also, oleic and stearic acids (30 microM) did not show a significant inhibitory effect on Icat. 7. A similar inhibitory action of EPA was observed when Icat was activated by intracellularly applied GTP gamma S in the absence of agonists, suggesting that the site of action of omega-3 fatty acids is not located on the receptor. 8. These results demonstrate that omega-3 polyunsaturated fatty acids can activate a K+ current and also effectively inhibit receptor-mediated non-selective cation currents in rat A7r5 vascular smooth muscle cells. Thus, the data suggest that omega-3 fatty acids may play an important role in the regulation of vascular tone.

Antiarrhythmic effect and its underlying ionic mechanism of 17beta-estradiol in cardiac myocytes.

1. The effects of oestrogens on action potential and membrane currents were examined in single guinea-pig atrial myocytes. 2. 17Beta-estradiol (3-10 microM) shortened the action potential duration without significant changes in the resting membrane potential. E-4031 (1 microM) markedly prolonged the action potential duration and induced early afterdepolarization, and 17beta-estradiol (10 microM) abolished it. 3. When cells were perfused in isoproterenol-containing solution, action potentials due to abnormal automaticity caused by membrane depolarization developed, and were also inhibited by 17beta-estradiol. 4. Under voltage clamp conditions, the voltage-dependent Ca2+ currents consisted of both T-(I(Ca,T)) and L-type (I(Ca,L)). 17Beta-estradiol reduced I(Ca,L) concentration-dependently, while it (10 microM) suppressed I(Ca,T) only by approximately 10%. 17Beta-estradiol did not affect time courses of I(Ca,L) inactivation, but it shifted the steady-state inactivation curve to more negative potentials. 5. 17Beta-estradiol (10 microM) did not affect the time-dependent K+ current (I(K)), referred to as I(Kr) and I(Ks) and inwardly rectifying K+ current. However, 17beta-estradiol (30 microM) or diethylstilbestrol (10 microM) inhibited K+ currents. 6. DES and ethinylestradiol (EES) also suppressed I(Ca,L), but testosterone and progesterone failed to inhibit I(Ca,L) The potency of the inhibitory effect on I(Ca,L) was DES> EES> 17beta-estradiol. 7. 17Beta-estradiol and DES also inhibited the cyclic AMP-enhanced I(Ca,L), but cyclic GMP in the pipette or pretreatment of L-NAME could not block the effects of oestrogen on I(Ca,L). 8 These results suggest that oestrogen specifically has antiarrhythmic effects, possibly by acting the L-type Ca2+ channels. The antiarrhythmic effects of oestrogens may contribute to the cardioprotective actions of oestrogens.

Primary biliary cirrhosis among atomic bomb survivors in Nagasaki, Japan.

Despite rapid progress in methods for analyzing radiation effects, much remains to be learned about the mechanisms and processes of radiation-induced immunological dysfunction. Among 17,899 sera obtained from atomic bomb survivors in Nagasaki, Japan, sera from 484 participants who complied with a reexamination for alkaline phosphatase (ALP) were tested for antimitochondrial antibody (AMA) by indirect immunofluorescence, and autoantibodies against 2-oxo-acid dehydrogenase complex (2-OADC) by immunoblotting to investigate the prevalence of primary biliary cirrhosis (PBC). Of these 484 sera, 28 (5.8%) were seropositive for AMA. The 484 participants were divided into three groups according to distance from the hypocenter: 72 who were exposed within 1999 m (closest group), 368 from 2000 to 5999 m (intermediate distant group), and 44 outside 6000 m (distant group). The positivity rates for AMA in these three groups were 6/72 (8.3%), 22/368 (6.0%), and 0/44 (0%), respectively (P =.08). Furthermore, high titers ( > 1:320) of AMA were observed in 3/6 (50%) AMA-positive sera from the closest group, in contrast to 4/22 (18%) from the intermediate distant group, although there was no significant correlation between AMA titer and distance from the hypocenter (P =.07). Of these 28 AMA-positive sera, 11 (39%) were from participants who had already been diagnosed with PBC, and 25 (89%) contained antibodies against at least one component of 2-OADC enzymes by immunoblotting. Therefore, the prevalence of PBC was estimated to be at least 615 cases per million (792 per million women). Our results suggest that the prevalence of PBC in atomic bomb survivors in Nagasaki is higher than that reported for the general population in Japan, and a further survey of the environmental factors, including radiation exposure, that predispose to PBC would be needed for understanding this disease of unknown etiology.

A preliminary double-blind study on the efficacy of carbamazepine in prophylaxis of manic-depressive illness.

A preliminary double-blind controlled study on the prophylactic effect of carbamazepine on recurrent manic-depressive psychotics was conducted with 22 patients using an inert placebo in ten subjects as a control drug. Carbamazepine in the dosage of 200-600 mg was administered for 1 year. Among the 22 carbamazepine subjects, carbamazepine was found to be effective in 60% of the cases and inert placebo in 22.2% (U-test, P less than 0.10). It is suggested that carbamazepine is a useful drug for the prophylaxis of manic-depressive illness.

Severe primary hyperparathyroidism in a neonate having a parent with hypercalcemia: treatment by total parathyroidectomy and simultaneous heterotopic autotransplantation.

Neonatal primary hyperparathyroidism is a life-threatening disease because of marked hypercalcemia and severe respiratory distress caused by the hypoplastic thorax and occasional rib fractures. We report a 29-day-old girl treated by total parathyroidectomy and simultaneous autotransplantation of parathyroid tissue (one fifth of each of the two glands) in the femoral quadriceps muscle near the groin. At the time of operation, all four of the parathyroid glands were markedly enlarged, and their total weight was 900 mg. Part of the resected parathyroid tissue was cryopreserved for further autotransplantation should hypoparathyroidism develop. Two years six months after surgery, the infant was well and had normal levels of serum calcium and immunoreactive parathyroid hormone in the absence of any supplementary treatment. Asymptomatic hypercalcemia in the presence of abnormally low fractional excretion of calcium was found in the father. Based on our experience and review of the literature, we recommend total parathyroidectomy, autotransplantation, and cryopreservation for the neonate with primary hyperparathyroidism.

The role of wet-dog shakes during amygdaloid electrical and methionine-enkephalin kindling in the rat.

The role of wet-dog shakes (WDS) in the kindling phenomenon was investigated in the rat using amygdala (AM) electrical kindling (E-K group), methionine-enkephalin (ME) chemical kindling (ME-K group) and ME-chemical kindling after the completion of electrical kindling (E-M group). The AM electrical kindling was carried out with 200 microA stimulation. Repeated microinjections of 10 micrograms ME into the AM were given for ME chemical kindling. EEG and behavioral seizures were recorded from the beginning of the electrical stimulation or ME microinjection to 2 min after the end of the after-discharge (AD). The mean number of WDS in ME-K and E-M groups during the chemical kindling, in contrast to that in E-K group, was significantly decreased as the kindling stages progressed. In the ME-K group, WDS completely disappeared when the stage developed into stage 5. In all the three groups, the maximum incidence of WDS in each stage appeared near the termination of AD, which was accordant with the end of the convulsive seizures. These results suggest that WDS may be associated with the kindling stage and the end of the convulsive seizure or the AD. Furthermore, the disappearance of WDS could be a behavioral index of the fully kindled state in some kinds of kindling models.

Effects of pirmenol on action potentials and membrane currents in single atrial myocytes.

Electrophysiological effects of pirmenol hydrochloride (pirmenol) were investigated in single atrial myocytes obtained from rabbit and guinea-pig hearts by using a whole-cell clamp technique. Under current clamp conditions, pirmenol (2-30 microM) prolonged action potential duration in a concentration-dependent manner without affecting resting membrane potential in rabbit atrial myocytes. However, in the presence of 4-aminopyridine (4 mM), pirmenol (10 microM) failed to prolong the action potential duration further. Pirmenol also suppressed acetylcholine-induced hyperpolarization and action potential duration shortening, resulting in a significant prolongation of the action potential duration in the presence of acetylcholine. Under voltage clamp conditions, pirmenol (1-1000 microM) inhibited transient outward current (I(to)) in a concentration-dependent manner. The concentration for half-maximal inhibition (IC50) of pirmenol on I(to) was about 18 microM. Pirmenol did not show the use and frequency dependent inhibition of I(to). The voltage dependence of the steady-state inactivation of I(to) and the recovery from inactivation were not significantly affected by pirmenol. Pirmenol accelerated the inactivation of I(to) and blocked I(to) as an exponential function of time, consistent with a time-dependent open channel blockade. Pirmenol (30 microM) did not affect the inwardly rectifying K+ current significantly, but it decreased the voltage-dependent L-type Ca2+ current by about 20%. In guinea-pig atrial myocytes, both acetylcholine and adenosine induced a specific K+ current activated by GTP-binding proteins. Pirmenol suppressed both the acetylcholine- and adenosine-induced K+ current effectively. The IC50 of pirmenol for acetylcholine- and adenosine-induced current was about 1 and 8 microM, respectively. The present results suggest that pirmenol prolongs the action potential duration by primarily inhibiting the transient outward current in atrial myocytes. In addition, since pirmenol inhibits acetylcholine- and adenosine-induced K+ current, pirmenol may effectively prolong the action potential duration in atrial myocytes under various physiological conditions as in the whole heart or ischemia.

Extracellular Mg2+ inhibits receptor-mediated Ca(2+)-permeable non-selective cation currents in aortic smooth muscle cells.

The effects of extracellular Mg2+ on receptor-mediated Ca(2+)-permeable non-selective cation currents were investigated in a cultured aortic smooth muscle cell line (A7r5) from rat thoracic aorta, using the whole-cell voltage-clamp technique. Under the Cs(+)-containing internal solution, both vasopressin and endothelin-1 (100 nM) activated a long-lasting inward current with a high noise level. The reversal potential of these agonists-induced current was approximately +0 mV, and was not significantly altered by the replacement of [Cl-]i or [Cl-]o, suggesting that the inward current was a cation-selective channel. La3+ and Cd2+ (1 mM) almost completely abolished the vasopressin or endothelin-induced non-selective cation current; however, nifedipine (10 microM) failed to inhibit it significantly. Extracellular Mg2+ (3-20 mM) also markedly inhibited the vasopressin- or endothelin-induced non-selective cation current in a concentration-dependent manner. When a non-hydrolysable GTP-analogue, GTP gamma S (1 mM), was applied from the patch pipette, the non-selective cation current was gradually activated even in the absence of agonist (vasopressin or endothelin-1), probably due to the direct activation of GTP-binding proteins coupled to the receptors. Extracellular Mg2+ (3-20 mM) also suppressed the activation of non-selective cation current induced by GTP gamma S, suggesting that the inhibitory sites of Mg2+ are not located on the receptors. These results suggest that extracellular Mg2+ inhibits receptor-mediated non-selective cation current, which may contribute to the relaxation effects of Mg2+ in vascular smooth muscle cells.

17beta-Estradiol inhibits the voltage-dependent L-type Ca2+ currents in aortic smooth muscle cells.

To elucidate the mechanisms of estrogens-induced relaxation effects on vascular smooth muscle cells, the effects of estrogens and the related hormones were examined in cultured rat thoracic aortic smooth muscle cell lines (A7r5), using the whole-cell voltage clamp technique. The patch pipette was filled with 140 mM CsCl- or KCl-containing internal solution. With CsCl-internal solution, 17beta-estradiol and synthetic estrogens, ethynylestradiol and diethylstilbestrol (0.1-30 mu M) inhibited the Ba2+ inward current (IBa) through the voltage-dependent L-type Ca2+ channel in a concentration-dependent and reversible manner. The potency of the inhibitory effects on IBa was 17beta-estradiol < ethynylestradiol < diethylstilbestrol. 17beta-Estradiol (10 mu M) appeared to reduce the maximal conductance of IBa with only a slight shift of voltage-dependency of inactivation and to affect IBa in a use-independent fashion. On the other hand, testosterone and progesterone (30 mu M) failed to affect IBa. At a holding potential of -40 mV, both vasopressin and endothelin-1 (100 nM) activated a long-lasting inward current. After endothelin-1 (100 nM) activated the current, the additional application of vasopressin (100 nM) could not induce it furthermore, suggesting that each agonist activates the same population of the channels. The reversal potential of the current was about 0 mV and was not significantly altered by replacement of [Cl-]i or [Cl-]0 and the inward current was also observed even when extracellular cations are Ca2+, proposing that it was a Ca2+-permeable non-selective cation channel (IN.S.). La3+ or Cd2+ (1 nM) completely abolished IN.S., however, nifedipine (10 mu M) failed to inhibit it at all. Diethylstilbestrol (1-30 mu M) suppressed the IN.S. evoked by both endothelin-1 and vasopressin in a concentration-dependent manner, while 17beta-estradiol, ethynylestradiol, progesterone and testosterone (30 mu M) failed to inhibit it significantly. In addition, at a holding potential of +0 mV, 17beta-estradiol by itself did not affect the holding currents, and did not inhibit K+ currents evoked by endothelin-1 or vasopressin, possibly due to the Ca2+ release from the storage sites. These results suggest that 17beta-estradiol may play a role in regulating vascular tone, selectively by inhibiting the voltage-dependent L-type Ca2+ current in vascular smooth muscle cells.

Effects of azelastine on membrane currents in tracheal smooth muscle cells isolated from the guinea-pig.

Azelastine [4-(p-chlorobenzyl)-2-(hexahydro-1-methyl -1H-azepin-4-yl)-1-(2H)-phthalazinone hydrochloride], an anti-allergic agent, inhibited the high K(+)-induced contraction in tracheal smooth muscle cells isolated from the guinea-pig. In order to investigate the ionic mechanisms, we examined the effects of azelastine on membrane currents, using the tight-seal whole cell voltage clamp technique. Azelastine (1-100 microM) caused an inhibition of the Ba2+ inward current (IBa) through the voltage-dependent L-type Ca2+ channel in a concentration-dependent manner. The inhibitory effect of azelastine on IBa was fully reversible. The IC50 value for azelastine-induced inhibition of IBa was approximately 8 microM, and 100 microM azelastine completely suppressed IBa. Azelastine exerted mainly a tonic block of IBa but did not show use dependence. Azelastine (10 microM) shifted the quasi-steady-state inactivation curve of IBa to more negative membrane potentials by approximately -20 mV, suggesting that the inhibitory effect of azelastine on IBa was voltage-dependent. In addition, azelastine produced inhibitory actions on other membrane currents (i.e. the voltage-dependent transient outward K+ current and the Ca(2+)-activated oscillatory K+ current) at doses higher than 10 microM. These results suggest that azelastine inhibits the voltage-dependent L-type Ca2+ current in single tracheal smooth muscle cells, which may contribute to the anti-allergic actions of azelastine in airways.

Omega-3 polyunsaturated fatty acids--modulation of voltage-dependent L-type Ca2+ current in guinea-pig tracheal smooth muscle cells.

Omega-3 polyunsaturated fatty acids have been reported to be associated with favorable changes in the respiratory system. To determine one of the mechanisms for this effect, membrane currents were recorded in guinea-pig tracheal myocytes by using the whole-cell voltage clamp technique. Without EGTA in the patch pipette containing the Cs-internal solution, command voltage pulses positive to +0 mV from a holding potential of -60 mV elicited a voltage-dependent L-type Ca2+ current (I(Ca x L)) and a subsequent outward current. Upon repolarization, slowly decaying inward tail currents were recorded. The outward currents and the inward tail current were enhanced by methyl-1,4,-dihydro-2,6-dimethyl-3-nitro-4-(2-trigluromethylphenyl )-pyridine-5-carboxylate, and blocked by Cd2+ or nifedipine. Inclusion of EGTA (5 mM) in the patch pipette also abolished these currents, indicating that they were Ca2+-dependent. When [Cl-]o or [Cl-]i was changed, the reversal potential of these currents shifted, thus behaving like a Cl(-)-sensitive ion channel. 4,4'-Diisothiocyanatostilbene-2,2'-disulphonic acid. a Cl- channel blocker, inhibited the currents. The omega-3 polyunsaturated fatty acids eicosapentaenoic acid (3-30 microM) and docosahexaenoic acid (30 microM) suppressed I(Ca x L) and then inhibited I(Ca x Cl) in a reversible manner. Similar inhibitory effects of eicosapentaenoic acid on I(Ca x L) were observed with 5 mM EGTA in the patch pipette. Neurokinin A (1 microM) and caffeine (10 mM) also transiently activated I(Cl x Ca), probably due to Ca2+ release from Ca2+ storage sites. Pretreatment of the cells with eicosapentaenoic acid markedly suppressed the activation of I(Cl x Ca) by neurokinin A or caffeine. These results suggest that omega-3 polyunsaturated fatty acids inhibit voltage-dependent L-type Ca2+ currents and also Ca2+-activated Cl- currents in tracheal smooth muscle cells from the guinea-pig, which may play a role in modulation of tracheal smooth muscle tone.

Chemical kindling with Met-enkephalin and transfer between chemical and electrical kindling.

The role played by Met-enkephalin (ME) in epileptic seizures was investigated, using 57 ME kindled rats and 10 saline injected control rats. Repeated microinjection of 10 micrograms ME into the right amygdala (AM) of male Wistar rats led to development of generalized convulsions. One week after the completion of ME kindling, 1 or 2 electrical stimulations (200-400 microA, 60 Hz, 1 sec) of the right AM of ME kindled rats resulted in generalized convulsions in 5 rats. The duration of after-discharge (AD) in the first generalized convulsion induced by electrical AM stimulation in the ME kindled rats was significantly longer than that in the first generalized convulsion induced by electrical stimulation in the saline treated control rats (P less than 0.05). One week after the completion of ME kindling, naloxone (10 or 20 mg/kg, i.p.) given 10 min before the infusion of ME into the other 3 ME kindled rats attenuated both convulsive behavior and electrographical seizures. With the progress of convulsive behavior, the frequency of wet-dog shakes (WDS) tended to decrease and was significantly lower after ME injection in the first stage 5 seizures than after the first ME injection (P less than 0.01). These results strongly suggest that ME has a potent epileptogenic effect on the rat brain which is caused by the opioid receptors. There are some differences between chemical kindling with ME and electrical kindling as indicated by the development of the AD duration and the WDS frequency.

Synchronized disappearance of serum HCV-RNA, anti-U1 RNP, anti-La/SS-B, and anti-Scl-70 in a patient with chronic hepatitis.

The authors report a rare case of chronic hepatitis in whom normalization of serum aminotransferases was associated with disappearance of serum hepatitic C virus (HCV)-ribonucleic acid (RNA), anti-U1 RNP, anti-La/SS-B, and anti-Scl-70 antibodies without treatment of interferon or corticosteroids. A 27-year-old Japanese woman was diagnosed with chronic hepatitis C, with positive anti-nuclear antibody, anti-U1 RNP, anti-La/SS-B, and anti-Scl-70 antibodies. Histopathologic examination of a liver biopsy specimen showed a periportal interface hepatitis with a predominantly lymphoplasmacytic necroinflammatory infiltrate and lobular hepatitis. After two-year treatment with ursodeoxycholic acid (UDCA), serum aminotransferases normalized and serum HCV-RNA, anti-U1 RNP, anti-La/SS-B, and anti-Scl-70 antibodies disappeared. It was unclear whether disappearance of HCV-RNA was spontaneous, due to some immunomodulating effects of UDCA, or other unknown mechanism, but host immune response may be associated with HCV elimination.

Surgical management of piriform sinus fistula.

Four cases of acute cervical abscess resulting from infection through the left piriform sinus fistula are described. The fistula seems to be related to the cause of infection in many of previously reported cases of acute suppurative thyroiditis or "recurrent lateral cervical fistula." Complete removal of the fistula is essential to a permanent cure, and the guide of a Fogarty catheter through the fistula is useful at operation for patients with repeated infection.