Selective desensitization of cardiac beta adrenoceptors by prolonged in vivo infusion of catecholamines in rats

The effects of prolonged in vivo infusion of isoproterenol (400 micrograms/kg/hr) or norepinephrine (200 micrograms/kg/hr) from a minipump on the physiological reactivity and binding properties of cardiac beta and alpha-1 adrenoceptors were tested in rats. Infusion of either catecholamine significantly reduced the in vitro inotropic and chronotropic potency of isoproterenol in isolated left and right atria, respectively; desensitization was near maximal as early as after 2 hr of infusion. No significant change in the density of [3H]dihydroalprenolol-labeled beta receptors was evident at this time point in either atrial or ventricular tissue, although isoproterenol did decrease binding site density after 7 days of infusion. There was no change in the binding affinity or physiological blocking potency of dihydroalprenolol after isoproterenol infusion. The inotropic potency of phenylephrine in the presence of dihydroalprenolol was unaffected by infusion of either isoproterenol or norepinephrine and methoxamine failed to increase right atrial rate either in control or in isoproterenol-infused rats. There was also no change in the density and affinity of [3H]prazosin binding sites after isoproterenol infusion. These results indicate selective desensitization of cardiac beta receptors without changes in alpha-1 receptors by prolonged in vivo stimulation with catecholamines. This reaction pattern is different from the well documented effects of hypothyroidism, which include decreased sensitivity of cardiac beta and increased sensitivity of cardiac alpha-1 receptor-mediated responses in rats. Thus, the mechanisms responsible for altered receptor function in the two conditions appear to be different.     

Effects of prolonged isoproterenol infusion on cardiac and vascular responses to adrenoceptor agonists

In vitro responses of cardiac and vascular smooth muscle to both adrenoceptor agonists and phosphodiesterase inhibitors were studied in tissues from either saline- or isoproterenol-infused rats. After chronic isoproterenol infusion the sigmoidal relationship between concentration of acutely administered isoproterenol and inotropic response of cardiac muscle was shifted to the right; the maximum response was decreased by approximately 40%. Inotropic responses were attenuated further by the beta adrenoceptor antagonist, propranolol. By contrast, quantitatively comparable inotropic responses to phenylephrine were not altered after isoproterenol infusion. However, they were blocked by the selective alpha adrenoceptor antagonist, prazosin, but were not affected by propranolol. Inotropic effects of the phosphodiesterase inhibitor, isobutylmethylxanthine, were comparable in tissues from either saline- or isoproterenol-infused rats. Similar results were obtained in vascular tissues. Portal veins and aortas from isoproterenol-infused rats were less responsive to the acute relaxant properties of the beta adrenoceptor agonists, isoproterenol and salbutamol. However, as in cardiac muscle, relaxant effects to phosphodiesterase inhibitors (isobutylmethylxanthine and papaverine) were not attenuated. In addition, contraction to norepinephrine was comparable in tissues from either saline- or isoproterenol-infused rats. These data indicate that isoproterenol infusion attenuates beta adrenoceptor-mediated responses of vascular and cardiac muscle to similar degrees but does not alter responses to either alpha adrenoceptor agonists or phosphodiesterase inhibitors.     

Effects of calcium channel blockers on cloned cardiac K+ channels IKr and IKs

Cloned HERG and KvLQT1-IsK K+ channels have been expressed in mammalian cells and assayed as a target for calcium channel blockers. These channels generate the rapid and slow components of the cardiac delayed rectifier K+ current, and mutations can affect them that lead to long QT syndromes. HERG is blocked by bepridil (EC50 = 0.55 microM), verapamil (EC50 = 0.83 microM) and mibefradil (EC50 = 1.43 microM), whereas nitrendipine and diltiazem have negligible effects. Steady-state activation and inactivation parameters are shifted to more negative values in the presence of the blockers. Similarly, KvLQT1-IsK is inhibited by bepridil (EC50 = 10.0 microM) and mibefradil (EC50 = 11.8 microM), whilst being insensitive to nitrendipine, diltiazem or verapamil. This work may help to understand the mechanisms of action of verapamil in certain ventricular tachycardias as well as some of the deleterious adverse cardiac events associated with bepridil and mibefradil.     

Bepridil block of cardiac calcium and sodium channels

The effects of bepridil, a new Ca channel blocking agent with reported antiarrhythmic action, were examined in single isolated ventricular cells using whole-cell patch clamp techniques. Ca currents were studied in guinea-pig ventricular cells and Na currents were studied in cultured ventricular cells from neonatal rat, a preparation which is more suitable for Na current measurements than guinea pig. At low frequencies (0.1 Hz) and negative holding potentials (-50 mV for Ca currents and -100 mV for Na currents), bepridil produced a concentration-dependent decrease in both Ca and Na currents without any significant change in the current-voltage relations. Concentration-response curves for block of Ca and Na channels were fitted by a one-to-one drug-receptor occupancy model. Half-blocking concentrations (IC50) of bepridil were 5 x 10(-7) M for Ca currents and 3 X 10(-5) M for Na currents. Bepridil had no effect on the inwardly rectifying K current and the time-dependent outward current. The effects of bepridil on Ca and Na currents depend upon the holding potential. Inactivation curves of the Ca and Na currents were shifted to more negative potentials by the drug. The recovery of both the Ca and Na currents from inactivation was always prolonged by bepridil and the repriming of both currents usually displayed an added exponential component, attributed to slow release of the drug from the channels. The results indicate that bepridil, by inhibiting both Ca and Na currents, may have clinical usefulness in the treatment of certain ischemia-induced ventricular arrhythmias.     

Bolus thrombolytic infusion during prolonged refractory cardiac arrest of undiagnosed cause

Acute myocardial infarction (AMI) and pulmonary embolism (PE) account for about 70% of cardiac arrest. Although thrombolytic therapy is an effective therapy for both AMI and PE, it is not routinely recommended during cardiopulmonary resuscitation (CPR) for fear of life threatening bleeding complications. Numerous case reports and retrospective studies have suggested a beneficial effect of thrombolytics in cardiac arrest secondary to AMI and PE; however, we present a case of successful use of bolus thrombolytics during CPR in a patient with undifferentiated cardiac arrest (undiagnosed cause) after prolonged conventional resuscitation without success.     

Effects of prolonged adrenaline infusion and of mental stress on plasma minerals and parathyroid hormone

Summary. The role of the sympatho-adrenal system for the secretion of PTH in humans is not established. Previous studies on the effects of adrenaline on plasma mineral homeostasis have focused on injections or short-term infusions of adrenaline, and conflicting results concerning calcium and parathyroid hormone (PTH) responses have been reported. We therefore infused adrenaline or placebo continuously for 3 h to 10 healthy volunteers and studied several plasma minerals, as well as PTH levels. Venous plasma adrenaline concentrations increased to the upper physiological range (5 nmol l^-1) during adrenaline infusion. Another nine volunteers were exposed for 25 min to mental stress (a colour word conflict test; CWT), which causes marked circulatory changes and raises plasma catecholamine concentrations. Plasma ionized and total calcium, and magnesium concentrations were slowly and gradually reduced during infusion of adrenaline, but there was only a small increase in PTH. Plasma potassium was decreased by adrenaline within 30 min and thereafter did not change further during infusion. There was a marked but transient increase in the plasma free fatty acids concentrations, which were not related to the reduction of the calcium or magnesium levels. The adrenaline-induced decrements in calcium, magnesium and potassium, and increases in heart rates persisted 30 min after the infusion, despite a rapid decrease in plasma adrenaline concentrations within 5 min of termination of the infusion. Plasma phosphate concentrations were lowered during the first 90 min of adrenaline infusion, but after 3 h they had returned to baseline despite continued infusion. CWT induced small increments of the plasma ionized calcium and PTH concentrations. Plasma potassium levels were raised despite increases in plasma adrenaline at the beginning of the stress test, while phosphate values were reduced at the end of the test. Thus, long-lasting elevations of circulating adrenaline lower plasma ionized and total calcium, phosphate, magnesium and potassium, but the time courses for these changes differed markedly. Despite the reduction of plasma ionized calcium there was only little increase in PTH and thus no indication that sustained elevations of circulating adrenaline stimulates the secretion of PTH in vivo in humans. Responses to acute mental stress and adrenaline infusion differed qualitatively, indicating that adrenaline responses to stress are of minor importance in this respect.     

Effects of prolonged high phosphorus diet on phosphorus and calcium balance in rats

The amount of phosphorus contained in food as food additives is currently increasing and a high intake of phosphorus can cause various diseases. To determine the effects of a prolonged high phosphorus diet, here we investigated the phosphorus and calcium balance and expression of type IIa sodium-dependent phosphate transporter (Npt IIa) in mature rats. Wistar male rats (8-weeks old) were divided into five groups and fed diets containing 0.6% calcium plus 0.3, 0.6, 0.9, 1.2 or 1.5% phosphorus for 4 weeks. Urinary and fecal phosphorus excretions were significantly increased by the high phosphorus diets (from 0.6 to 1.5%), dependent on the amount of dietary phosphorus. The net absorption of intestinal phosphorus was also significantly increased by high phosphorus diets. As a result, a negative phosphorus balance was observed in rats given the 1.2% or 1.5% phosphorus diets. Serum parathyroid hormone and 1,25-dihydroxyvitamin D(3) concentrations were increased by high phosphorus diets. In addition, high phosphorus diets decreased the expression of Npt IIa mRNA and protein in the renal brush border membrane. Taken together, these results suggest that diets containing 1.2 or 1.5% phosphorus plus 0.6% calcium have potentially adverse effects on phosphorus homeostasis in mature rat.     

Effects of ranolazine on cloned cardiac kv4.3 potassium channels

The effects of ranolazine, an antianginal drug, on potassium channel Kv4.3 were examined by using the whole-cell patch-clamp technique. Ranolazine inhibited the peak amplitude of Kv4.3 in a reversible, concentration-dependent manner with an IC(50) of 128.31 μM. The activation kinetics were not significantly affected by ranolazine at concentrations up to 100 μM. Applications of 10 and 30 μM ranolazine had no effect on the fast and slow inactivation of Kv4.3. However, at concentrations of 100 and 300 μM ranolazine caused a significant decrease in the rate of fast inactivation, and at a concentration of 300 μM it caused a significant decrease in the rate of slow inactivation, resulting in a crossover of the current traces during depolarization. The Kv4.3 inhibition by ranolazine increased steeply between -20 and +20 mV. In the full activation voltage range, however, no voltage-dependent inhibition was found. Ranolazine shifted the voltage dependence of the steady-state inactivation of Kv4.3 in the hyperpolarizing direction in a concentration-dependent manner. The apparent dissociation constant (K(i)) for ranolazine for interacting with the inactivated state of Kv4.3 was calculated to be 0.32 μM. Ranolazine produced little use-dependent inhibition at frequencies of 1 and 2 Hz. Ranolazine did not affect the time course of recovery from the inactivation of Kv4.3. The results indicated that ranolazine inhibited Kv4.3 and exhibited a low affinity for Kv4.3 channels in the closed state but a much higher affinity for Kv4.3 channels in the inactivated state.     

Effects of tertatolol on post- and prejunctional beta adrenoceptors

The effects of tertatolol, a new and powerful beta adrenoceptor blocking drug, on post- and prejunctional beta receptors were investigated; canine vascular tissues (saphenous veins, coronary arteries and splenic arteries) and guinea-pig trachea and atria were used. At concentrations below 10(-5) M, tertatolol did not alter basal tension or contractile responses to electrical stimulation, norepinephrine, K+ or prostaglandin F2 alpha; at doses at or above 10(-5) M the drug-evoked contractions which were reduced by phentolamine and were absent in denervated veins. Tertatolol at 10(-5) M and 3 X 10(-5) M augmented the basal efflux of [3H] norepinephrine in saphenous veins labeled with the 3H-transmitter. In veins, 10(-5) M of tertatolol depressed the contractions caused by electrical stimulation without affecting those to exogenous norepinephrine; this concentration of the drug also inhibited the stimulation-induced overflow of [3H]norepinephrine. The major part of the present study was designed to test the beta receptor blocking properties of tertatolol and to compare its effects with those of propranolol. Tertatolol inhibited, in a concentration-dependent manner, the relaxations caused by isoproterenol in saphenous veins, splenic arteries and coronary arteries and the relaxations evoked by norepinephrine and epinephrine in coronary arteries; the potency of tertatolol was higher than that of propranolol. In trachea and right atria of the guinea-pig, tertatolol inhibited, in a concentration-dependent manner, the dose-response curves to isoproterenol; the relative potency of tertatolol was higher than that of propranolol. In dog saphenous veins, previously incubated with [3H]norepinephrine, tertatolol (10(-7)M) blocked the increased stimulation-evoked overflow of the 3H-transmitter induced by isoproterenol.(ABSTRACT TRUNCATED AT 250 WORDS)     

咪唑安定长期镇静对重症脓毒症病人免疫平衡的影响

目的 观察外科ICU(SICU)严重腹内感染病人使用咪唑安定镇静48 h 对促炎/抗炎平衡的影响,为SICU镇静药物的选用提供理论依据。方法 28例符合重型脓毒症诊断的外科严重腹内感染病人术后随机分为咪唑安定组和对照组,另设健康组5例,咪唑安定组给予咪唑安定镇静,时间超过48 h,对照组使用生理盐水,检测两组病人入SICU即刻和48 h后血清IFN-γ、TNF-α、IL-10、单核细胞表面人类白细胞抗原-DR(HLA-DR)表达。结果 两组病人入SICU后即刻各细胞因子均明显高于健康组,HLA-DR表达显著低于健康组,组间无差别,咪唑安定平均负荷量0.063 mg/kg,维持剂量0.054 mg·kg-1·h-1。48 h后咪唑安定组TNF-α、IL-10较入院明显下降,IFN-γ变化不显著,单核细胞HLA-DR表达有恢复趋势(P=0.057)。对照组IFN-γ、TNF-α、IL-10、HLA-DR均较入院时无显著差异,组间比较IL-10、HIA-DR差异有显著性。结论 咪唑安定长期镇静不仅可发挥其作用平稳、可控性强、遗忘作用好等药理特点,而且在抑制TNF-α等促炎因子、减轻SIRS损伤的同时,降低抗炎因子IL-10分泌、促进单核细胞HLA-DR表达的细胞免疫功能恢复、改善免疫平衡。在外科高危病人术后恰当应用咪唑安定可能对病人预后产生有利影响。     

Alpha-1 adrenoceptors and calcium in isolated canine coronary arteries

Experiments were designed to define the postjunctional alpha adrenoceptor subtype(s) in large canine coronary arteries and to determine the dependency of contractions due to their activation upon the entry of extracellular calcium. Rings of left circumflex coronary artery were mounted at their optimal length for isometric tension recording in organ chambers filled with physiological salt solution. Phenylephrine and cirazoline were full agonists relative to norepinephrine. Methoxamine was a partial agonist relative to norepinephrine whereas clonidine, xylazine, B-HT 920 and B-HT 933 produced minimal contractions. Prazosin competitively inhibited the contractile response to phenylephrine (pA2 = 8.6), whereas rauwolscine caused a noncompetitive inhibition and was more than 100 times less potent than prazosin at inhibiting the response to phenylephrine. Similar results were obtained using norepinephrine (in the presence of propranolol) as the agonist. The calcium-entry blockers nimodipine, verapamil and diltiazem inhibited contractions caused by norepinephrine, phenylephrine and cirazoline. Removal of extracellular calcium abolished the response to cirazoline. These results suggest that in large canine coronary arteries: 1) only alpha-1 adrenoceptors are present postjunctionally and 2) responses due to alpha-1 adrenoceptor activation are dependent upon extracellular calcium.     

Effects of irbesartan on cloned potassium channels involved in human cardiac repolarization

We studied the effects of irbesartan, a selective angiotensin II type 1 receptor antagonist, on human ether-a-go-go-related gene (HERG), KvLQT1+minK, hKv1.5, and Kv4.3 channels using the patch-clamp technique. Irbesartan exhibited a low affinity for HERG and KvLQT1+minK channels (IC(50) = 193.0 +/- 49.8 and 314.6 +/- 85.4 microM, respectively). In hKv1.5 channels, irbesartan produced two types of block, depending on the concentration tested. At 0.1 microM, irbesartan inhibited the current in a time-dependent manner (22 +/- 3.9% at +60 mV). The blockade increased steeply with channel activation increasing at more positive potentials. However, at 10 microM, irbesartan induced a time-independent blockade that occurred in the range of potentials of channel opening, reaching its maximum at approximately 0 mV, and remaining unchanged at more positive potentials (24.0 +/- 1.0% at +60 mV). In Kv4.3 currents, irbesartan produced a concentration-dependent block, which resulted in two IC(50) values (1.0 +/- 0.1 nM and 7.2 +/- 0.6 microM). At 1 microM, it inhibited the peak current and accelerated the time course of inactivation, decreasing the total charge crossing the membrane (36.6 +/- 7.8% at +50 mV). Irbesartan shifted the inactivation curve of Kv4.3 channels, the blockade increasing as the amount of inactivated channels increased. Molecular modeling was used to define energy-minimized dockings of irbesartan to hKv1.5 and HERG channels. In conclusion, irbesartan blocks Kv4.3 and hKv1.5 channels at therapeutic concentrations, whereas the blockade of HERG and KvLQT1+minK channels occurred only at supratherapeutic levels. In hKv1.5, a receptor site is apparent on each alpha-subunit of the channel, whereas in HERG channels a common binding site is present at the pore.     

Subconductance activity induced by quinidine and quinidinium in purified cardiac sarcoplasmic reticulum calcium release channels

This study examined the effects of quinidine, quinine, and the quaternary quinidine derivative, quinidinium, on the conductance and activity of purified cardiac sarcoplasmic reticulum calcium release channels/ryanodine receptors (RyR) incorporated into planar lipid bilayers. Quinidine (50-500 microM) reduced the single-channel open probability in a voltage- and concentration-dependent manner. Reduction of channel activity was evident only at positive holding potentials where current flow is from the cytoplasmic to luminal side of the channel and when the drug was present only on the cytoplasmic face of the channel. A more pronounced effect was the appearance of a subconductance state at positive potentials. Single channel recordings and dose-response experiments revealed that at least two quinidine molecules were involved in reduction of the RyR activity. The permanently charged quinidinium compound produced nearly identical effects as quinidine when present only on cytoplasmic side of the channel, suggesting the positive-charged form of quinidine is responsible for the effects on the channel. There was no stereospecificity in the effects of quinidine because the levoisomer, 100 microM quinine, produced a similar subconductance activity of the channel. Ryanodine modification of the channel prevented subconductance activity. These findings suggest that the quinidine-induced subconductance activity may be the result of a partial occlusion of the channel pore interfering with ion conduction. Modification of the channel by ryanodine alters quinidine binding to the channel through a conformational change in protein structure.     

吡拉西坦对缺氧大鼠海马神经元钙激活钾通道的作用

目的：研究吡拉西坦对缺氧大鼠海马神经元钙激活钾通道的作用，以揭示吡拉西坦抗缺血性脑损伤的电生理途径。 方法：实验于2004—03／2005—06在泸州医学院心肌电生理研究室完成。应用膜片钳制技术分别记录吡拉西坦和缺氧对海马神经元钙激活钾通道的单通道电流活动的影响，并经P-clamp软件进行微机采样、储存数据和数据的分析处理。 结果：（1）吡拉西坦对钙激活钾通道具有明显的激活作用，随着药物浓度（2．5—7．5mmol／L）的增加，通道开放概率明娃增加，由用药前的0．032&;#177;0．010增加到0．272&;#177;0．038（P〈0.01，n=6），伴随有通道平均关闭时间明显缩短，而电流幅值及平均开放时间无明显变化。（2）应用氰化钠20μmol／L造成细胞急性缺氧。在缺氧早期（5-15min）通道开放概率增加，由缺氧前的0．024&;#177;0．009增加至0．090&;#177;0．026（P〈0．01，n=6），而缺氧后期通道开放概率明显降低，表明缺氧时间对通道的开放概率有明显影响。（3）吡拉西坦能明显增加缺氧神经元钙激活钾通道的开放概率，而以缺氧20min时为最明显，缺氧40min时为最低，即由对照组的0．038&;#177;0.008逐渐增加至最高时的0．148&;#177;0．060，然后又逐渐降低至最低时的0．033&;#177;0．005（P〈0．01，n=6）。 结论：吡拉西坦对海马锥体神经元大电导钙激活钾通道具有明显的激活作用。吡拉西坦可能通过激活钙激活钾通道等机制发挥对缺氧神经元的保护作用。     

吡拉西坦对缺氧大鼠海马神经元钙激活钾通道的作用

目的:研究吡拉西坦对缺氧大鼠海马神经元钙激活钾通道的作用,以揭示吡拉西坦抗缺血性脑损伤的电生理途径。方法:实验于2004-03/2005-06在泸州医学院心肌电生理研究室完成。应用膜片钳制技术分别记录吡拉西坦和缺氧对海马神经元钙激活钾通道的单通道电流活动的影响,并经P-clamp软件进行微机采样、储存数据和数据的分析处理。结果:①吡拉西坦对钙激活钾通道具有明显的激活作用,随着药物浓度(2.5～7.5mmol/L)的增加,通道开放概率明显增加,由用药前的0.032±0.010增加到0.272±0.038(P<0.01,n=6),伴随有通道平均关闭时间明显缩短,而电流幅值及平均开放时间无明显变化。②应用氰化钠20μmol/L造成细胞急性缺氧,在缺氧早期(5～15min)通道开放概率增加,由缺氧前的0.024±0.009增加至0.090±0.026(P<0.01,n=6),而缺氧后期通道开放概率明显降低,表明缺氧时间对通道的开放概率有明显影响。③吡拉西坦能明显增加缺氧神经元钙激活钾通道的开放概率,而以缺氧20min时为最明显,缺氧40min时为最低,即由对照组的0.038±0.008逐渐增加至最高时的0.148±0.060,然后又逐渐降低至最低时的0.033±0.005(P<0.01,n=6)。结论:吡拉西坦对海马锥体神经元大电导钙激活钾通道具有明显的激活作用。吡拉西坦可能通过激活钙激活钾通道等机制发挥对缺氧神经元的保护作用。