利用普通电极导管测定在体心脏单相动作电位的研究

采用吸引电极记录的在体心脏单相动作电位(MAP)与同时用玻璃微电极记录的单个心肌细胞的跨膜动作电位(TAP)具有同样的形态。然而,因吸引部位心肌损伤、出血、MAP波形畸变、振幅下降等,限制了实验,尤其是临床研究的应用。1983年国外有人利用特制的接触电极导管稳定地记录了人的在体心脏MAP。我们利用普通国产电极导管也较理想地测定到了兔和人的在体心脏MAP。现介绍如下。     

利用接触电极记录人或动物在体心脏的单相动作电位

单相动作电位(monophasic action potential,MAP)与单个心肌细胞的跨膜动作电位(transmembrane action Potential,TAP)具有同样的形态和间期,能较精确地反映心肌细胞的去极化和复极化过程。以往采用吸引电极(Suetion electrode)容易造成吸引部位的心肌损伤、出血,并引起波形畸变、振幅下降,持续时间也短,限制了它的临床     

测定人和动物心脏单相动作电位的新方法

用吸引电极记录人和动物在体心脏的单相动作电位(MAP),虽可获得心肌细胞群电活动的某些信息,但由于吸引电极对心肌细胞的损伤,因而限制了临床应用。本文介绍用接触电极导管代替吸引电极记录MAP的方法。对接触电极的制备、实验程序、MAP的形态、测量、特征以及应用价值均作了简要说明。     

单相动作电位技术在猫在体心脏触发性活动研究中的应用

应用接触电极记录心脏单相动作电位(MAP)是80年代发展起来的新技术。MAP是心肌局部细胞群的电活动,它与单细胞跨膜动作电位(TAP)具有基本相同的形态、间期和复极化过程。自1985年开始,很多研究者将其用于在体心脏触发性活动的研究。触发性活动是近年来提出的心律失常发生机制的新概念,包括早期后去极化(EAD)、延时性     

在体心脏后去极化及触发性心律失常的细胞电生理学研究

应用漂浮微电极记录跨膜动作电位(TAP)和接触电极记录单相动作电位(MAP)两项技术,研究了 CsCl 诱发的猫在体心脏的触发性活动。结果表明,静脉注射 CsCl(0.5mmol/kg)后10s,左心室外膜 TAP 和 MAP 的3相中晚期出现早期后去极化(EAD),在30s 时,TAP上 EAD 的振幅为25.6±9.3mV,MAP 上的 EAD 振幅为3.4±1.3mV。EAD 表现为拖尾、平台、凸起三种不同形态。在两例猫中出现延迟性后去极化 DAD,其在 TAP 和 MAP上的振幅分别为13.0±5.3mV 和3.3±0.omV。MAP 上后去极化的形态与 TAP 极为相似。CsCl 的重复注射可诱发室性早搏、室性心动过速等多种心律失常,根据后去极化的发生与否以及后去极化和室性心搏的耦联间期的关系,可将心律失常分为两种类型:一种是由记录部位的后去极化引起的触发性心律失常;另一类可能由非记录部位的后去极化或其它机制引起。     

离体灌流心脏希氏束电图记录方法

为稳定地记录离体心脏希氏束电图,我们研制成一种新型的二合一插管电极,利用这种电极可同时完成Ｌａｎｇｅｎｄｏｒｆｆ法心脏灌流及希氏束电位记录。心脏取自成年家兔或幼狗,与Ｌａｎｇｅｎｄｏｒｆｆ法相同,首先经主动脉逆行插入这种插管电极实现心脏灌流,然后在示波器监视下小心转动插管,当电极贴近希氏束部位时,即可记录到离体灌流心脏的希氏束电位。实践证明：这种方法操作简便,组织损伤小,波型记录稳定。如配合心电等     

用单相动作电位研究触发性心律失常

触发性心律失常(triggered arrhythmia)以冲动起源异常机制为特征受到临床的普通重视。近年来,接触电极或接触电极导管记录人或动物心内、外膜单相动作电位新技术的出现,为在整体动物或人心脏上获得触发性活动提供了前提。目前,单相动作电位(mono-phasic action Potential,MAP)已用于冠状动脉搭桥术中的局部心肌缺血、心肌电生理     

低能量心脏除颤的实验研究

本研究用22只犬建立实验模型。除颤脉冲采用非周期性衰减波,通过四种不同的除颤电极系统向心脏发放:1.双极导管电极;2.多极导管电极;3.导管—杯状心外膜电极;4.导管一体表电极。共除颤151次,成功133次,其中小于30J(焦耳)的成功除颤93次。上述各种电极在除颤能量小于30J时以导管—杯状心外膜电极的除颤成功率为最高,达84.5％。测量9只犬的导管除颤电极心肌阻抗值为431±117(?)(欧姆)。研制了一种多道除颤信号的A/D、D/A接口,在微型计算机上进行了除颤脉冲的波形分析和除颤能量的计算等试验。     

在体兔心急性心肌缺血时的复极后不应性

本实验观察了开胸麻醉家兔急性心肌缺血时的不应期改变。用吸引电极记录的单相动作电位(MAP)确定机能不应期(FRP)及 FRP 超过 MAP 复极90%时程(MAPD_(90))的复极后不应性(PRR)。阻断冠脉左室支以后,缺血区不应期表现出两种完全不同的变化:缺血中心区不应期延长并超过 MAPD_(90),出现 PRR;缺血周边区不应期则通常较对照缩短。利用自体颈总动脉的动脉血进行灌流的兔心实验表明,当100%阻断灌流血液时,缺血中心区出现PRR,但是当50%阻断时,同一部位心肌的不应期却表现为缩短。上述结果提示:PRR 是在严重心肌缺血情况下出现的。同一时刻测定缺血中心区、周边区和非缺血区的功能不应期,结果表明,冠脉阻断后不应期离散程度的明显增大是与当时 PRR 的出现密切相关的。因此,PRR 的存在可以认为是造成急性心肌缺血时不应期离散的重要原因,而后者一般认为是异致折返性心律失常的重要因素。     

高血压大鼠心肌肥大及逆转过程中相关因素的探讨

目的：探讨在心肌肥大及逆转过程中收缩压(SBP)、舒张压(DBP)、平均动脉压(MAP)、神经肽Y(NPY)等与左心室肥大的关系。方法：血压和心率用生物信号分析系统记录;NPY用放射免疫法测定,用SPSS软件求出了相关系数和回归方程。结果：SBP、DBP、MAP、心肌匀浆中NPY与心系数(LVW／BW)呈正相关,血液中NPY和心率(HR)与心系数不相关。结论：血压升高是导致左室肥大的因素之一,收缩压的影响大于舒张压;SBP、DBP、MAP、心肌匀浆中NPY与心系数(LVW/BW)有相关的趋势。     

Localization and characterization of the binding site for the regulatory subunit of type II cAMP-dependent protein kinase on MAP2

Microtubule-associated protein 2 (MAP2) binds, and is a substrate for, type II cAMP-dependent protein kinase. The structural domain in MAP2 that binds the regulatory subunit (RII) of protein kinase II was identified by expressing fragments of a human MAP2 cDNA in E. coli using the pATH11 vector. Fusion proteins were resolved by SDS-PAGE and transferred to nitrocellulose. The filters were probed with purified bovine heart or brain RII, anti-RII monoclonal antibodies, and 125I-labeled protein A. Binding of RII was localized to a 31 amino acid sequence near the N-terminus of the MAP2 molecule. Fusion proteins containing this fragment bound both heart and brain RIIs in a concentration-dependent manner, but bound heart RII with a higher apparent affinity than brain RII. The amino acid sequence of this fragment (DRETAEEVSARIVQVVTAEAVAVLKGEQEKE) is totally conserved between human and mouse MAP2, suggesting an important role for the RII binding site of MAP2 in neuronal function.     

Genesis of the monophasic action potential: role of interstitial resistance and boundary gradients

The extracellular potential at the site of a mechanical deformation has been shown to resemble the underlying transmembrane action potential, providing a minimally invasive way to access membrane dynamics. The biophysical factors underlying the genesis of this signal, however, are still poorly understood. With the use of data from a recent experimental study in a murine heart, a three-dimensional anisotropic bidomain model of the mouse ventricular free wall was developed to study the currents and potentials resulting from the application of a point mechanical load on cardiac tissue. The applied pressure is assumed to open nonspecific pressure-sensitive channels depolarizing the membrane, leading to monophasic currents at the electrode edge that give rise to the monophasic action potential (MAP). The results show that the magnitude and the time course of the MAP are reproduced only for certain combinations of local or global intracellular and interstitial resistances that form a resting tissue length constant that, if applied over the entire domain, is smaller than that required to match the wave speed. The results suggest that the application of pressure not only causes local depolarization but also changes local tissue properties, both of which appear to play a critical role in the genesis of the MAP.     

在体心脏后去极化及触发性心律失常细胞电生理学研究

CsCl triggered activities in cat heart in vivo were studied by using floating microelectrode and contact electrode to record transmembrane and monophasic action potentials (TAP and MAP). Ten seconds after CsCl (0.5 m mol/kg, i.v.), early after depolarization (EAD) appeared in the middle-later period of phase 3 in both TAP and MAP. Thirty seconds after CsCl, the amplitude of TAP-EAD was 25.6 +/- 9.3 mV and that of MAP-EAD was 3.4 +/- 1.3 mV. The potential changes of the EADs could be divided into three kinds, i.e. the "tail", the "plateau" and "peak" types. Delayed after depolarization (DADs) could also be induced by CsCl in the phase 4 of the TAP and MAP in two cats. The amplitudes of TAP-DAD and MAP-DAD were 13.0 +/- 5.3 mV and 3.3 +/- 0.6 mV respectively. The types of the afterdepolarizations in MAP were very similar to those in TAP. The ventricular extrasystole and/or tachycardias could be induced by repeated injections of CsCl. According to the occurrence of after depolarization (AD) and the relationship between the coupling interval of the AD and that of the ventricular beat, two kinds of generation of arrhythmias were suggested, i.e. one triggered by AD of the myocardium under the electrode and the other induced by AD originating from the other sites of the myocardium.     

Transmural recording of monophasic action potentials

To investigate the possibility of transmural recording of repolarization through the ventricular wall, KCl monophasic action potential (MAP) electrodes positioned along plunge needles were developed and tested. The MAP electrode consists of a silver wire surrounded by agarose gel containing KCl, which slowly eluted into the adjacent tissue to depolarize it. In six dogs, a plunge needle containing three KCl MAP electrodes was inserted into the left ventricle to simultaneously record from the subepicardium, midwall, and subendocardium. In six pigs, eight plunge needles containing three KCl MAP electrodes and two plunge needles containing similar electrodes except for the absence of KCl were inserted into the ventricles. In three guinea pig papillary muscles, a KCl electrode was used to record MAPs along with two microelectrodes for recording transmembrane potentials. Transmural MAP recordings could be made for >1 h in dogs and >2 h in pigs with a significant decrease in MAP amplitude over time but without a significant change in MAP duration. With the electrodes without KCl in pigs, the injury potentials subsided in <30 min. When the pacing rate was changed to alter the action potential duration and refractory period in dogs, the MAP duration correlated with the local effective refractory period (r = 0.94). The time course of the MAP duration recorded with a KCl MAP electrode in guinea pig papillary muscles corresponded well with that of the transmembrane potential recorded with an adjacent microelectrode. It is possible to record transmural repolarization of the ventricles with KCl MAP electrodes on plunge needles. The MAP is caused by the KCl rather than being a nonspecific injury potential.     

Developmental regulation of cardiac MAP4 protein expression

It has been shown that the level of expression of microtubule-associated protein 4 (MAP4) mRNAs changes throughout neonatal heart development [Chapin SJ, et al. 1995. Biochemistry 34:2289]. In the present study, both immunofluorescence and western blotting methods were used to monitor MAP4 protein expression levels in the developing heart. By both methods, it was shown that the levels of total MAP4 protein were maximal during the first postnatal week, and then declined progressively to adulthood. In addition, four major electrophoretic species that reacted with MAP4-specific antibodies (called bands 1-4) were observed in all heart tissue samples. Three of the four bands decreased in abundance throughout postnatal development, but at different rates. The fourth band remained relatively constant in abundance with increasing postnatal age. To determine if phosphorylation events might contribute to this heterogeneity, western blotting experiments using phospho-specific antibodies and phosphatase digestion of extract samples were performed. No phosphorylation-specific antibody staining was observed and no significant changes were demonstrated in the bands after phosphatase treatment, implying that the observed complexity was due mainly to alternative start site or differential isoform expression. Finally, it was discovered that cardiomyocyte MAP4 associated with drug- and cold-stable microtubules in early neonatal myocytes. Thus, the complex regulation of MAP4 protein expression may play a key role in the functional differentiation of myocyte microtubules during heart development.     

Electrical and mechanical response in biventricular mechanical alternans.

Mechanical alternans of various degrees is produced by rapid heart rates, slower rates in failing hearts and can be brought about by a single extra systole. It has also been shown that the two ventricles may exhibit different degrees of mechanical alternation. The present study was planned to clarify the possible mechanism inducing this latter phenomenon. For this reason myocardial tension was recorded simultaneously from the two ventricles as well as through a miniature strain gage capable of measuring electrogram and myocardial tension of a small area -- just adjacent to a stimulating electrode. The heart was driven at a steady heart rate through one electrode and very late premature beats were applied at various coupling times at another site through an electrode attached to the miniature strain gage. It was found that the degree of mechanical alternans is markedly different at the sites of measurements in either ventricle. These changes could be related to the time interval elapsed between the application of the electrical stimulus and the occurrence of the mechanical response.     

Arabidopsis microtubule-associated protein MAP65-3 cross-links antiparallel microtubules toward their plus ends in the phragmoplast via its distinct C-terminal microtubule binding domain

Plant cytokinesis is brought about by the phragmoplast, which contains an antiparallel microtubule (MT) array. The MT-associated protein MAP65-3 acts as an MT-bundling factor that specifically cross-links antiparallel MTs near their plus ends. MAP65 family proteins contain an N-terminal dimerization domain and C-terminal MT interaction domain. Compared with other MAP65 isoforms, MAP65-3 contains an extended C terminus. A MT binding site was discovered in the region between amino acids 496 and 588 and found to be essential for the organization of phragmoplast MTs. The frequent cytokinetic failure caused by loss of MAP65-3 was not rescued by ectopic expression of MAP65-1 under the control of the MAP65-3 promoter, indicating nonoverlapping functions between the two isoforms. In the presence of MAP65-3, however, ectopic MAP65-1 appeared in the phragmoplast midline. We show that MAP65-1 could acquire the function of MAP65-3 when the C terminus of MAP65-3, which contains the MT binding site, was grafted to it. Our results also show that MAP65-1 and MAP65-3 may share redundant functions in MT stabilization. Such a stabilization effect was likely brought about by MT binding and bundling. We conclude that MAP65-3 contains a distinct C-terminal MT binding site with a specific role in cross-linking antiparallel MTs toward their plus ends in the phragmoplast.     

Indomethacin attenuates post-suspension hypotension in Sprague-Dawley rats

Orthostatic hypotension is a serious condition that is sometimes manifested in astronauts during standing postflight. These observations may be related to impairment of autonomic function and/or excessive production of endothelium-dependent relaxing factors. To evaluate the role of the cyclooxygenase inhibitor indomethacin as a countermeasure against the post-suspension reduction in mean arterial pressure (MAP), we examined the cardiovascular responses to 7-day 30 degrees tail-suspension and a subsequent 6-hr post-suspension period in conscious male Sprague-Dawley rats. Indomethacin (2 mg/kg) or saline were administered intravenously prior to release from suspension and at 2 and 4 hrs post-suspension. Direct MAP and heart rate were determined prior to suspension, daily and every 2 hrs post-suspension. During suspension, MAP did not change, in contrast, during post-suspension; it decreased compared to parallel non-suspended, untreated animals. There were no significant changes in heart rate. The reduction in MAP post-suspension was associated with significant increases in plasma prostacyclin. Indomethacin attenuated the observed post-suspension reduction in MAP and reduced plasma prostacyclin levels. Also, the baroreflex sensitivity for heart rate was modified by indomethacin--the MAP threshold for baroreflex activation was raised and the effective MAP range expanded. Thus, the post suspension reduction in mean arterial pressure may be due to overproduction of vasodilatory prostaglandins and/or other endothelium-dependent relaxing factors and alteration in baroreflex activity.     

Cardiovascular responses to apneic facial immersion during altered cardiac filling.

The hypothesis that reduced cardiac filling, as a result of lower body negative pressure (LBNP) and postexercise hypotension (PEH), would attenuate the reflex changes to heart rate (HR), skin blood flow (SkBF), and mean arterial pressure (MAP) normally induced by facial immersion was tested. The purpose of this study was to investigate the cardiovascular control mechanisms associated with apneic facial immersion during different cardiovascular challenges. Six subjects randomly performed 30-s apneic facial immersions in 6.0 +/- 1.2 degrees C water under the following conditions: 1) -20 mmHg LBNP, 2) +40 mmHg lower body positive pressure (LBPP), 3) during a period of PEH, and 4) normal resting (control). Measurements included SkBF at one acral (distal phalanx of the thumb) and one nonacral region of skin (ventral forearm), HR, and MAP. Facial immersion reduced HR and SkBF at both sites and increased MAP under all conditions (P < 0.05). Reduced cardiac filling during LBNP and PEH significantly attenuated the absolute HR nadir observed during the control immersion (P < 0.05). The LBPP condition did not result in a lower HR nadir than control but did result in a nadir significantly lower than that of the LBNP and PEH conditions (P < 0.05). No differences were observed in either SkBF or MAP between conditions; however, the magnitude of SkBF reduction was greater at the acral site than at the nonacral site for all conditions (P < 0.05). These results suggest that the cardiac parasympathetic response during facial immersion can be attenuated when cardiac filling is compromised.     

A MAP kinase docking site is required for phosphorylation and activation of p90(rsk)/MAPKAP kinase-1

Activation of the various mitogen-activated protein (MAP) kinase pathways converts many different extracellular stimuli into specific cellular responses by inducing the phosphorylation of particular groups of substrates. One important determinant for substrate specificity is likely to be the amino-acid sequence surrounding the phosphorylation site; however, these sites overlap significantly between different MAP kinase family members. The idea is now emerging that specific docking sites for protein kinases are involved in the efficient binding and phosphorylation of some substrates [1] [2] [3] [4]. The MAP kinase-activated protein (MAPKAP) kinase p90 rsk contains two kinase domains [5]: the amino-terminal domain (D1) is required for the phosphorylation of exogenous substrates whereas the carboxy-terminal domain (D2) is involved in autophosphorylation. Association between the extracellular signal-regulated kinase (Erk) MAP kinases and p90(rsk) family members has been detected in various cell types including Xenopus oocytes [6] [7] [8], where inactive p90(rsk) is bound to the inactive form of the Erk2- like MAP kinase p42(mpk1). Here, we identify a new MAP kinase docking site located at the carboxyl terminus of p90(rsk). This docking site was required for the efficient phosphorylation and activation of p90(rsk) in vitro and in vivo and was also both necessary and sufficient for the stable and specific association with p42(mpk1). The sequence of the docking site was conserved in other MAPKAP kinases, suggesting that it might represent a new class of interaction motif that facilitates efficient and specific signal transduction by MAP kinases.