Cromakalim effects of acetylcholine-induced changes in cytosolic calcium and tension in swine trachealis.

The effects of cromakalim, an ATP-sensitive K+ channel activator, on changes in cytosolic calcium concentration [( Ca++]i) and tension induced by acetylcholine (ACh; 0.1-10 microM) were examined in swine tracheal smooth muscle. Cromakalim (10 microM) hyperpolarized muscle cells by approximately 18 mV from -58 mV (resting membrane potential) to -76 mV. Cromakalim relaxed muscle contractions evoked by ACh at a concentration of 0.1 microM, but not at higher concentrations. Measurement of [Ca++]i using Fura-2 demonstrated that except at 0.1 microM ACh, cromakalim did not alter peak increases in [Ca++]i. At 0.1 microM ACh, the peak transient was decreased, but not eliminated. Cromakalim reduced steady-state increases in [Ca++]i at ACh less than or equal to 1 microM, but not 10 microM ACh. Tension was similarly affected. These data suggest that ACh-induced increases in steady-state [Ca++]i and tension are inhibited by cromakalim-induced hyperpolarization. The initial ACh-induced transient increase in [Ca++]i is not greatly altered. Cromakalim did not alter the transient peak tension and [Ca++]i relationship. The relationship between steady-state [Ca++]i/tension (EC50 = 321 nM) obtained for control, cromakalim inhibition and after glibenclamide reversal of cromakalim inhibition falls to the left of the peak transient [Ca++]i/tension relationship (EC50 = 587 nM). Thus, the Ca++ sensitivity of the contractile proteins during steady-state stimulation by ACh was increased from that at rest. We conclude that electromechanical coupling is important in ACh-induced contraction at concentrations less than 1 microM. Pharmacomechanical coupling with little or no sensitivity to changes in potential is important at higher ACh concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)     

The pathophysiology of tension pneumothorax in ventilated swine

It remains unclear as to whether the cardiovascular collapse observed in tension pneumothorax (TP) is strictly a mechanical pressure-related phenomenon or secondary to hypoxemia. This study describes the pathophysiologic changes associated with a surgically induced progressive TP in a ventilated swine model. With a balloon occlusion catheter surgically placed into the pleural space, progressive volumes of pneumothorax were created in six anesthetized pigs on positive-pressure ventilation. Air was introduced into the right hemithorax in 100-mL increments every 4–5 min, with measurements of heart rate (HR), mean arterial pressure (MAP), central venous pressure (CVP), mean intrapleural pressure (MIP), oxygen saturation (O₂%), arterial blood gas (ABG), and cardiac output (C.O.). With the induced progressive TP, results showed that O₂% measures decreased immediately and continued to decline throughout the experiment to levels below 50% prior to cardiovascular collapse. The MAP and HR remained relatively stable until approximately 57% total lung capacity progressive TP (600 mL) was reached. At this point, a significant decline in MAP and increase in HR was noted, indicating tension physiology. The C.O. showed a small but significant decrease after 200 mL of air was injected, with a progressive decline after this point. At >97% total lung capacity TP, lethal cardiovascular collapse occurred in all animals and was associated with an abrupt drop in C.O., HR, and MAP. There was a concurrent equalization of MIP with CVP at the point of collapse. Arterial blood gas measures correlated with O₂% trends during the trials. We conclude that the findings of this study support the alternative hypothesis that significant hypoxemia occurs early and precedes hypotension in ventilated animals with TP. Occlusive mechanical compression, suggested by equalization of MIP and CVP, is probably a late event.     

Prostaglandin E2 enhances acetylcholine-induced, Ca2+-dependent ionic currents in swine tracheal mucous gland cells

Airway submucosal gland cell (SMGC) secretions are under the control of various neurotransmitters and hormones. Interactions between different pathways, such as those mediated by cAMP and Ca(2+), in controlling mucus or electrolyte secretions are not well understood. Prostaglandin E(2) (PGE(2)) or forskolin has been shown to enhance acetylcholine (ACh)-induced short circuit current (I(SC)) in SMGC mucous cell monolayers. We show that PGE(2), by activating cAMP-dependent protein kinase A (PKA), enhanced ACh-induced, Ca(2+)-mediated current and changes in [Ca(2+)](i) in mucous cells. PGE(2) pretreatment sensitized ACh-induced I(SC) (DeltaI(SC)) by activating endoprostanoid (EP(2)) receptors. PKA inhibitors 14-22 amide PKI (PKI) and Rp-diastereomer (Rp) of cAMPs prevented the effect of PGE(2). Removing external Ca(2+) or pretreatment with the Ca(2+) entry blocker, SKF96365 [1-[beta-(3-(4-methoxyphenyl) propoxy)-4-methoxyphenethyl]-1H-imidazole hydrochloride1-[2-(4-methoxyphenyl)-2-[3-(4-methoxyphenyl) propoxy] ethyl] imidazole], shifted the concentration-response relationships for ACh to the right but did not abolish PGE(2)-induced sensitization of the ACh response. An inositol 1,4,5-trisphosphate (IP(3)) receptor antagonist and Ca(2+) entry blocker, 2-aminoethoxydiphenyl borate, abolished the ACh-induced response. Charybdotoxin, but not iberiotoxin (IbTX), inhibited the ACh-induced DeltaI(SC). Clotrimazole, but not IbTX, inhibited the ACh-induced serosal K(+) current. Under whole-cell patch clamp, ACh-induced K(+) and Cl(-) currents were coincident with increases in [Ca(2+)](i) in single mucous cells. PGE(2) or forskolin pretreatment did not induce current or [Ca(2+)](i) changes but enhanced ACh-induced currents, membrane hyperpolarization, and [Ca(2+)](i) changes. Intra-cellular dialysis with the PKA-catalytic subunit enhanced ACh-induced whole-cell current as well. These findings demonstrate that PGE(2), via EP(2) receptors and the cAMP/PKA pathway, activates Ca(2+) entry-independent mechanisms, possibly by increasing IP(3)-mediated Ca(2+) release, resulting in the sensitization of ACh-induced currents.     

Permissive role of prostanoids in acetylcholine-induced cerebral vasoconstriction

The present study was designed to test the hypothesis that prostanoids play a permissive role in acetylcholine-induced cerebral constriction. Pial arterioles of newborn pigs were observed using a closed cranial window. Pial arteriolar constriction induced by topical acetylcholine (10(-5) M) was blocked by indomethacin (5 mg/kg i.v.), but was restored when acetylcholine was coadministered with topical prostaglandin (PG) F2 alpha (1 ng/ml), U46619 (1 ng/ml) or PGH2 (100 ng/ml). The restored acetylcholine response was blocked by topical pirenzepine (10(-3) M), a muscarinic-1 antagonist. Constriction and ability of all three prostanoids to restore acetylcholine-induced constriction was blocked by SQ 29,548 (10(-4) M), a purported thromboxane A2/PGH2 receptor antagonist. Subthreshold concentrations of U46619 and PGF2 alpha (0.1 ng/ml) restored acetylcholine-induced constriction, whereas threshold and subthreshold concentrations of PGE2, platelet-activating factor and norepinephrine had no effect. Therefore, activation of the thromboxane A2/PGH2 receptor appears to be necessary for acetylcholine-induced constriction to occur. Thus, prostanoids appear to play a permissive role in acetylcholine-induced pial arteriolar constriction in newborn pigs.     

Methionine-enkephalin in migraine and tension headache. Differences between classic migraine, common migraine and tension headache, and changes during attacks

We measured methionine-enkephalin (MET) in platelets and platelet-poor-plasma of drug-free patients with classic migraine, common migraine or tension headache (TH). Migraineurs were studied both between and during attacks. TH patients were characterized by low platelet-MET and high plasma-MET levels, whereas migraine patients showed high platelet-MET and reduced plasma-MET levels. During migraine attacks both platelet and plasma-MET concentrations increased considerably. We suggest that determination of MET levels in platelets and plasma may be a useful marker to discriminate between TH and common migraine. The rise in MET during attacks may be directed against the ictal increase of serotonin.     

Taurocholate induces changes in rat cardiomyocyte contraction and calcium dynamics

Obstetric cholestasis is characterized by raised bile acids, and can be complicated by intrauterine death. We have shown that the bile acid taurocholate causes loss of synchronous beating, bradycardia and cessation of contraction in cultured rat cardiomyocytes [Williamson, Gorelik, Eaton, Lab, de Swiet and Korchev (2001) Clin. Sci. 100, 363-369]. The aim of the present study was to investigate the effect of taurocholate on cardiomyocytes further. We demonstrated a reduced rate of contraction and proportion of beating cells when rat cardiomyocytes were exposed to increasing concentrations of taurocholate (0.1-3.0 mM); more marked at higher concentrations (P<0.001). Using scanning ion-conductance microscopy, we also demonstrated reduced amplitude of contraction and calcium transients with taurocholate. Our observations indicate that taurocholate affects calcium release from the sarcoplasmic reticulum and this parallels changes in contractile function. The relationship between the contraction amplitude and calcium transient is not linear, particularly at higher concentrations of taurocholate. We observed different effects in individual cultured neonatal cells; a reversible reduction in rate and amplitude of contraction in some, and irreversible oscillatory (fibrillatory) cessation of beating in others. The effects were more marked with higher concentrations. The contraction amplitude was also reduced in adult cardiomyocytes. The changes were reversible following removal of taurocholate in adult, but not in neonatal, cardiomyocytes exposed to higher concentrations (>0.3 mM) (P<0.001). In conclusion we have demonstrated that the bile acid taurocholate can cause different types of dysrhythmia in individual cardiomyocytes. These results provide further support for the hypothesis that obstetric cholestasis may produce cardiac-related sudden intrauterine death.     

心肺复苏后猪各主要脏器组织和超微结构的变化及对血流动力学的影响

目的 通过建立心搏骤停室颤-心肺复苏模型,观察心肺复苏后猪心肌、大脑、肺脏、肝脏及肾脏组织普通病理和超微结构的变化及对血流动力学的影响.方法 16头北京长白猪(29～35kg)随机(随机数字法)分为正常组(n=8)与复苏组(n=8).前组动物仅进行麻醉及气管插管,不予致室颤及心肺复苏.后组动物在建立心搏骤停室颤模型4 min后给予标准心肺复苏术,恢复自主循环(ROSC)10 min后以0.5 mL/(h·kg)的速度持续静脉泵入生理盐水直至复苏后6 h,并分别于致颤前,ROSC后0.5 h,2 h,4 h,6 h监测心排血量(CO)、左心室收缩期压力最大上升速率(+dp/dtmax)、左心室舒张期最大下降速率(-dp/dtmax)、心率(HR)、平均动脉压(MAP),两组动物均于复苏6 h后静脉麻醉下处死,速取其左心室心尖、大脑顶叶皮层、左肺、肝右叶、右肾上极组织行普通病理学及透射电镜检查.对实验数据用t检验的方法进行统计学分析.结果 复苏组在ROSC后各时间点,左心室收缩期压力最大上升速率、左心室舒张期最大下降速率、心排血量与致颤前相比均明显降低,差异具有统计学意义(P＜0.05),复苏组ROSC后各时间点HR较致颤前明显增加(P＜0.05),各时间点MAP之间比较差异无统计学意义(P＞0.05);通过对苏木精-伊红(HE)染色切片及透射电镜切片的观察,进行正常组与复苏组动物各主要脏器组织病理学比较,心搏骤停室颤-心肺复苏模型中猪的心肌、大脑、肺脏、肝脏及肾脏组织均存在不同程度的病理损伤,以心肌、大脑、肺脏的损伤更为严重.结论 心搏骤停-心肺复苏对机体各主要脏器均可以造成不同程度的形态学损伤及血流动力学影响,在ROSC成功6 h的模型中,以心肌、大脑及肺的损伤较重,肝、肾的损伤较轻.     

Perinatal changes in lung surfactant calcium measured in situ.

Calcium ion is thought to play a role in the structure and function of pulmonary surfactant after secretion into the alveolar space. Since fetal lung liquid calcium concentration is inadequate for this hypothesized role, at a time when optimal surfactant function is necessary for survival, we speculated that the necessary calcium is secreted with the surfactant material, i.e., in the lamellar body. Lungs from rat fetuses at 20, 21, and 22 d gestation, and also from newborn rats at 3-5 h, 1 and 3 d, were rapidly frozen, sectioned, freeze-dried, and examined cold (-100 degrees C) in a transmission electron microscope equipped with a fully quantitative energy-dispersive x-ray detector and analyzer. X-ray spectra were collected from the lamellar bodies and cytoplasm of type II cells at each time point. Lamellar body calcium concentration in the fetus was approximately twice that of the adult (70 +/- 4 vs. 37 +/- 2 mmol/kg dry wt +/- SEM, P less than 0.01), and it decreased rapidly after birth to adult levels. Apically located lamellar bodies in the fetus have a significantly higher calcium concentration than those in a perinuclear position (76 +/- 4 vs. 52 +/- 3, P less than 0.01). There is significant correlation of calcium and chloride concentrations in lamellar bodies, suggesting that factors responsible for the distribution of chloride, i.e., pH, may also be responsible for the accumulation of calcium by these organelles. These results show that mature calcium transport in lamellar bodies is achieved prenatally in the rat, and suggest that the calcium required for normal surfactant function at birth is secreted with the lamellar body.     

Effect of acute changes in renal arterial blood flow on urine oxygen tension in dogs

The relationships between urine oxygen tension (PuO2), PaO2, and acute changes in renal arterial blood flow (RBF) were studied in 19 anesthetized dogs. Indwelling sensors that measure continuously PO2 were inserted into the femoral artery and the ureter. RBF was measured by an electromagnetic flowmeter placed over the renal artery. PuO2 increased significantly from 36 to 72 torr during a stepwise increase in PaO2 from 70 to 180 torr. RBF was decreased in a stepwise fashion from a baseline value of 5.51 to 4.16, 2.13, and finally to .20 ml/kg.min by aortic constriction. PuO2 decreased significantly from a baseline value of 72 torr to 66, 57, and finally to 23 torr. The correlation coefficient between RBF and PuO2 was .84, which was significantly higher than that between RBF and femoral arterial pressure or that between RBF and urinary flow rate. This study demonstrates that PuO2 is a sensitive indicator of acute RBF changes in normal, healthy dogs.     

Change mechanisms in EMG biofeedback training: cognitive changes underlying improvements in tension headache

Forty-three college students suffering from recurrent tension headache were randomly assigned to 1 of 4 electromyographic (EMG) biofeedback training conditions. Although all subjects were led to believe they were learning to decrease frontal EMG activity, actual feedback was contingent on decreased EMG activity for half of the subjects and increased EMG activity for the other half. Within these 2 groups, subjects also viewed bogus video displays designed to convince them they were achieving large (high success) or small (moderate success) reductions in EMG activity. Regardless of actual changes in EMG activity, subjects receiving high-success feedback showed substantially greater improvement in headache activity (53%) than subjects receiving moderate success feedback (26%). Performance feedback was also related to changes in locus of control and self-efficacy. Changes in these 2 cognitive variables during biofeedback training were also correlated with reductions in headache activity following treatment, whereas changes in EMG activity exhibited during training were uncorrelated with outcome. These results suggest that the effectiveness of EMG biofeedback training with tension headache may be mediated by cognitive changes induced by performance feedback and not primarily by reductions in EMG activity.     

高渗条件下大鼠下丘脑神经元和星形胶质细胞内钙的动态变化

背景既往有很多关于培养神经元或胶质细胞内钙的研究.然而关于高渗条件下共同观察神经元和胶质细胞内钙变化的研究目前较少.目的探讨高渗刺激后大鼠下丘脑神经元和星形胶质细胞(AST)内钙的变化.设计随机对照实验研究.单位一所大学医院的神经外科和一所大学神经科学研究所的实验室.材料实验在南方医科大学医院神经外科和解放军第四军医大学全军神经科学研究所完成.新生两三天的SD大鼠2只和孕18 d大鼠胚胎6只,清洁级,由解放军第四军医大学实验动物中心提供.方法体外联合培养的神经元和AST经钙荧光指示剂Fluo-3负载后,用激光扫描共聚焦显微镜测定高渗NaCl溶液影响前后的细胞内钙水平随时间变化情况.主要观察指标联合培养的神经元和AST的形态观察及等渗和高渗状态下神经元和AST内Ca2+含量测定.结果高渗刺激使培养神经元和AST内钙水平先升高后降低,最后维持在比高渗刺激前稍高的静息钙水平上.AST钙水平达到峰值的速度略快于神经元.结论神经元和AST内Ca2+在渗透压信息传递过程起作用.     

实验猪肺栓塞前后死腔、分流及气体交换的研究

目的 研究肺栓塞死腔、肺内分流及气体交换的变化特点,探讨肺栓塞发生的病理生理学机制及意义.方法 建立肺栓塞模型,将8只健康幼猪于栓塞前30 min,栓塞后0、30、60和120 min 5个时间点,对生理死腔(VDphy)、肺泡死腔(VDalv)、气道死腔(VDaw)、肺内分流(Qs/Qt)、肺动脉压(PAP)、动脉血氧分压(PaO2)、二氧化碳分压(PaC02)及肺泡动脉氧分压差(PA-aDO2)进行测定.结果 研究发现VDphy、VDalv、Qs/Qt、PAP、PA-aDO2栓塞后较栓塞前明显增大(P＜0.01).PaO2栓塞后较栓塞前明显减小(P＜0.05和P＜0.01).VDaw、PaCO2栓塞前后差异无统计学意义(P＞0.05).SNK检验VDphy、VDalv栓塞后60 min和120 min较30min减小;PAP于栓塞后30 min、60 min、120 min较0 min减小;其余各组栓塞后各时间点差异无统计学意义.血液动力学指标栓塞前后无明显变化.结论 肺栓塞发生后,机体发生了一系列病理生理学变化,即死腔和肺内分流增大,氧分压减小,肺泡动脉氧分压差增大,而二氧化碳分压保持不变.     

Effect of calcium antagonists on basal and digitalis-dependent changes in splanchnic and systemic hemodynamics

We investigated the effects of the calcium antagonists verapamil and tiapamil on basal splanchnic and systemic hemodynamics and digoxin-induced vasoconstriction in 16 healthy men, using the hepatic venous catheter technique, the thermodilution method, and systolic time intervals. After a baseline period, verapamil or tiapamil were given by a primed-constant infusion in six of 16 subjects each, and hemodynamic changes were determined. Thereafter digoxin (1 mg) was concomitantly infused and hemodynamic changes were observed over 105 minutes. Control trials with digoxin alone were performed in seven of 16 subjects. Four of 16 subjects received only placebo. Digoxin provoked an increase in systolic blood pressure and splanchnic vascular resistance. Estimated splanchnic blood flow, mean pulmonary artery pressure, and total electromechanical systole decreased. Verapamil did not change basal hemodynamic parameters. Tiapamil decreased diastolic blood pressure and systemic and splanchnic vascular resistance. Both drugs attenuated the vasoconstricting effect of digoxin on the splanchnic vascular bed. Results indicate that calcium antagonists might be beneficial in treatment of digitalis-induced splanchnic vasospasm.     

Foscarnet-induced changes in plasma concentrations of total and ionized calcium and magnesium in HIV-positive patients

The time course and magnitude of foscarnet-induced changes in plasma concentrations of total and ionized calcium and magnesium were investigated in 13 male HIV-positive patients who had no active cytomegalovirus-associated disease. The patients had a mean age of 36 years (range 25-49 years) and a mean CD4 cell count of 550 cells/mm3 (range 130-1280 cells/mm3). Peak (mean +/- SD) plasma concentrations of foscarnet (0.89+/-0.10 mmol/l) were seen at the end of the period of drug infusion (90 mg/kg of foscarnet was infused over 2 hours) and declined with a terminal half-life of 5.7+/-0.7 hours. Plasma concentrations of total calcium declined over an 8-hour period, with the lowest concentration occurring after 4 hours (baseline: 2.29+/-0.09 mmol/l; lowest: 2.18+/-0.07 mmol/l; P < 0.001). By contrast, the lowest plasma concentration of ionized calcium occurred after 2 hours (baseline: 1.25+/-0.04 mmol/l; lowest: 0.99+/-0.05 mmol/l; P < 0.001), before gradually recovering to baseline levels over the next 10 hours. The mean maximal decrease in total calcium was 0.11+/-0.06 mmol/l, compared with 0.26+/-0.04 mmol/l for ionized calcium (P < 0.001). Plasma concentrations of total magnesium declined from 0.79+/-0.06 mmol/l (baseline) to 0.74+/-0.04 mmol/l (P < 0.05) after 4 hours and remained at this level after 8 hours. However, plasma concentrations of ionized magnesium fell steeply from 0.56+/-0.03 mmol/l to 0.39+/-0.03 mmol/l at 2 hours (P < 0.001), followed by a gradual recovery over the next 10 hours. The mean maximal decrease in total magnesium was 0.05+/-0.08 mmol/l, compared with 0.18+/-0.03 mmol/l (P < 0.001) for ionized magnesium. In summary, we found that foscarnet-induced changes in the plasma concentrations of total calcium and magnesium were dissociated from the corresponding changes in ionized calcium and magnesium. The maximal decreases in the plasma concentrations of total calcium and magnesium were smaller in magnitude and occurred much later than did the changes in ionized calcium and magnesium. The relative changes in the plasma concentration of ionized magnesium were greater than those of ionized calcium, indicating that foscarnet binds preferentially to the magnesium ion.