蜜蜂间接飞翔肌肌原纤维副肌球蛋白的鉴定

十五年前我们曾经提出蜜蜂间接飞翔肌粗肌丝大概是由一个贯穿整个肌小节,在A带是中空的内芯以及一层包含只位于A带的六根微丝之外套所组成。从分离出的微丝来看,它的构成单元很象是肌球     

蜜蜂间接飞翔肌肌原纤维副肌球蛋白的鉴定

十五年前我们曾经提出蜜蜂间接飞翔肌粗肌丝大概是由一个贯穿整个肌小节,在A 带是中空的内芯以及一层包含只位于A 带的六根微丝之外套所组成。从分离出的微     

蜜蜂间接飞翔肌粗肌丝的微丝结构

用能溶解肌球蛋白但不溶解副肌球蛋白的溶液(300 mM KCI,pH6.0)处理分离的蜜蜂间接飞翔肌粗肌丝,经数分钟后可以看到粗肌丝端头散开成为多根微丝,微丝数最多为7根。延长处理时间,可以见到粗肌丝中央部分只剩下直径约为5 nm的徽丝。实验结果支持我们以前提出的蜜蜂间接飞翔肌粗肌丝的结构模式,并指示贯穿肌小节、两端都和Z线相连的内芯至少部分由副肌球蛋白组成。只存在于A带的,由6根微丝形成的外套是由肌球蛋白分子组成。     

螫虾腹屈肌的肌球蛋白-副肌球蛋白丝

利用甘油梯度离心方法分离和纯化螯虾腹屈肌粗肌丝,电子显微镜照片显示粗肌丝上有数条纵行条纹,指示其可能由数根亚丝所组成。粗肌丝的 SDS-聚丙烯酰胺凝胶电泳表明其含有肌球蛋白和副肌球蛋白,肌球蛋白仅包含有二种轻链。副肌球蛋白类晶体呈针状,具有14.5nm 和72.5nm 的横纹周期。实验结果表明,螯虾腹屈肌粗肌丝是肌球蛋白-副肌球蛋白丝。     

蜜蜂飞翔肌收缩蛋白的电子显微镜鉴定

自五十年代肌丝滑行模型建立以来,关于脊椎动物骨胳肌的蛋白质成分,肌丝排列以及肌肉收缩时结构变化的研究取得了很大的进展。骨胳肌肌原纤维由粗、细肌丝有规律地排列所组成。对于肌肉收缩蛋白的选择性抽提,专一性抗体标记以及重组肌丝的研究,证实肌球蛋白存在于粗肌丝;肌动蛋白、原肌球蛋白和原宁蛋白存在于细肌丝(Huxley,A.F.,1957;Huxley,H.E;,1972)。昆虫间接飞翔肌的结构和生理特性有许多不同于脊椎动物骨胳肌的特点。蜜蜂飞翔肌肌原纤维虽然也包含有粗、细两     

蜜蜂間接飞翔肌的电生理观察

本文报告我們观察蜜蜂間接飞翔肌肌細胞的电反应和測量膜的一些电性貭的結果。膜电位的数值为35±7毫伏,峯电位的振幅为42±11毫伏。“自发”放电和通过头腹部两点刺激引起的反应伴有延續时間較长(数十至数百毫秒)的負后电位,在它的上面还往往重复出現一些峯电位。直接刺激肌肉引起的反应的負后电位較小,而且沒有重复反应。細胞膜的电阻为1—4×10~3欧姆厘米~2,电容为2—9微法拉/厘米~2。     

螯虾腹屈肌肌原纤维副肌球蛋白和原肌球蛋白的分离和结晶

螯虾Procambarus clarki腹屈肌肌原纤维副肌球蛋白和原肌球蛋白已经分离和结晶。在离子强度近于0.3和pH 6.0的溶液中透析,螯虾副肌球蛋白类晶体析出,与原肌球蛋白可以彼此完全分离。经含200mmol KCl,10mmol磷酸缓冲液,pH 6.0溶液或者50mmol BaCl_2 50mmol Tris.HCl_1 pH7.8溶液透析,可以得到副肌球蛋白针状类晶体,周期为14.5或72.5nm。原肌球蛋白溶液经含2％饱和硫酸铵,10mmol醋酸缓冲液,pH 5.1之溶液透析,呈现针状或梭形结晶,具有39nm周期。     

伸展程度不同的蜜蜂间接飞翔肌肌原纤维A带和I带蛋白浓度的比例

本文报告伸展程度不同的蜜蜂間接飞翔肌肌原纤維A带和I带蛋白浓度的比例。当肌小节长度从3.0微米增加到4.7微米,蛋白浓度之比稍稍上升。实驗結果和根据Hodge提出的肌原纤維由一組蛋白細絲組成的模型計算的結果大体相符。     

意蜂三个品系间的基因差异

用聚丙烯酰胺凝胶等电聚焦(1EF)电泳对意蜂(Apis mellifera ligustica)三个品系(渐农大A系, 湖北意蜂和引进的原意蜂)的成年工蜂进行了苹果酸脱氢酶 (MDH) 同工酶的测定。它们的MDH图谱均有三个区带(MDHI、MDHll和MDHIll),其中MDHII是由三个等位基因(a、b、c) 编码的,应有六种基因型(a/a、b/b、c/c、a/b、b/c和a/c),但本实验室测定的360余头个体中,均宋 出现b/b型.浙农大A系出现了除b/b型以外的其它五种基因型,而其它两个晶系都仅出现了c/c, a/c和b/c等三种基因型。对结果进行独立性检验,浙农大A系的MDHII的基因型频率、基因频率以及 杂合度等与其它两个晶系均有很显著的差异,这说明它与这两个晶系已有了相当程度的遗传分化,因此,这个结果可以为浙农大A系的建立提供一个遗传和生化的依据。     

THE ELONGATION OF MYOFIBRILS FROM THE INDIRECT FLIGHT MUSCLE OF DROSOPHILA

Myofibrils which lengthen by several per cent in the presence of ATP and magnesium ions were prepared by teasing indirect flight muscle of Drosophila in solutions containing ethylenediaminetetraacetate. A study was made of the hydrogen ion, magnesium ion, ATP, and potassium chloride concentrations with which this effect could be observed. The lack of elongation with pyrophosphate and several nucleoside triphosphates suggests that the lengthening is ATP specific. A relaxing factor system comparable to that described for rabbit muscle was not demonstrable, as elongated fibrils did not shorten with calcium ions, carnosine, or digitonin.     

LOCALIZATION OF MYOSIN FILAMENTS IN SMOOTH MUSCLE

Thick myosin filaments, in addition to actin filaments, were found in sections of glycerinated chicken gizzard smooth muscle when fixed at a pH below 6.6. The thick filaments were often grouped into bundles and run in the longitudinal axis of the smooth muscle cell. Each thick filament was surrounded by a number of thin filaments, giving the filament arrangement a rosette appearance in cross-section. The exact ratio of thick filaments to thin filaments could not be determined since most arrays were not so regular as those commonly found in striated muscle. Some rosettes had seven or eight thin filaments surrounding a single thick filament. Homogenates of smooth muscle of chicken gizzard also showed both thick and thin filaments when the isolation was carried out at a pH below 6.6, but only thin filaments were found at pH 7.4. No Z or M lines were observed in chicken gizzard muscle containing both thick and thin filaments. The lack of these organizing structures may allow smooth muscle myosin to disaggregate readily at pH 7.4.     

THE ORGANIZATION OF FLIGHT MUSCLE FIBERS IN THE ODONATA

The cytological organization of flight muscle fibers of Odonata has been investigated. These fibers, in representatives of the Zygoptera and Anisoptera, have been compared and found to be similar, except that, in the former, pairs of lamellar fibrils, rather than single fibrils, alternate with the mitochondria. In each instance, in these synchronous muscles, the actin filaments of the myofibrils are found to lie opposite to and midway between pairs of myosin filaments—a configuration previously reported in asynchronous flight muscle fibers. The disposition of the T system and sarcoplasmic reticulum membranes in glutaraldehyde-fixed anisopteran muscle is described in detail: the T system tubules are shown to be radially continuous across the fiber, and are derived as openmouthed invaginations from the surface cell-membrane. The detailed organization of the dyad junctions between these tubules and the adjoining cisternae of the sarcoplasmic reticulum is described. The accessibility of the T system interior to diffusion exchange with the general extracellular milieu has been investigated by studies on the penetration of ferritin into the fiber: molecules of this marker have been found to diffuse solely along the T system tubules, and their presence in the tubule extremities adjoining the centrally placed nuclei confirms the morphological evidence suggesting that these tubules provide open diffusion channels extending across the radius of the fiber. The possible physiological role of these membrane components and their distribution in synchronous muscles of insects and vertebrates and in asynchronous insect flight muscle are discussed.     

LOCALIZATION AND PROPERTIES OF THE CHOLINESTERASE IN CRUSTACEAN MUSCLE

Cholinesterase (ChE) activity is present in crustacean muscle extracts. However, since acetylcholine (ACh) is not a neuromuscular transmitter in these animals, the role and exact localization of ChE was unknown. The histochemical localization of the enzyme was studied in whole muscle and in the sarcoplasmic reticulum fraction of the extract, 50-µm frozen sections of glutaraldehyde-fixed crayfish tail flexor muscle were incubated with acetylthiocholine (ATC) as substrate, and examined under the electron microscope. After some modifications in published techniques, dense deposits were found associated with the sarcolemma, sarcolemmal invaginations, and transverse tubules. No deposits were found in 10^-4 M eserine, or if butyrylthiocholine (BTC) was substituted for ATC. The vesicles in the sarcoplasmic reticulum fraction which demonstrate the activity must represent minced bits of these membranes. Using a spectrophotometric method, the kinetics of the crustacean muscle enzyme was compared to the acetylcholinesterase (AChE) on mammalian red blood cells and in the lobster ventral nerve cord. Surprisingly, and contrary to previous reports, the crustacean muscle enzyme did not demonstrate substrate inhibition. While a number of similarities to AChE were found, this lack of substrate inhibition makes questionable an unequivocal similarity with classical AChE.     

CYTOCHEMICAL LOCALIZATION OF LACTIC DEHYDROGENASE IN WHITE SKELETAL MUSCLE

The limitations of the conventional histochemical methods for localization of lactic dehydrogenase (LDH) in white skeletal muscle have been analyzed quantitatively. It is demonstrated that more than 80 per cent of LDH diffuses into the incubation medium within the first 10 minutes of incubation. Furthermore, it is confirmed that the addition of phenazine methosulfate (PMS) to the ingredients of the histochemical reaction for LDH increases substantially the capacity of the white muscle extract to reduce Nitro-BT. Based on these observations, a modified method of cytochemical localization of LDH has been developed. This method prevents the leakage of LDH from tissue sections by the application of all the ingredients of the histochemical reaction to tissue sections in a thin gelatin film. The incubation mixture contains PMS so that the staining system is independent of tissue diaphorase. The application of this method to the adductor magnus muscle of the rabbit revealed a fine reticulum in the sarcoplasm of all muscle fibers, in addition to the staining of mitochondria. The distribution of the staining suggests that LDH is localized in the sarcoplasmic reticulum.     

IMMUNOHISTOCHEMICAL LOCALIZATION OF CONTRACTILE PROTEINS IN LIMULUS STRIATED MUSCLE

Limulus paramyosin and myosin were localized in the A bands of glycerinated Limulus striated muscle by the indirect horseradish peroxidase-labeled antibody and direct and indirect fluorescent antibody techniques. Localization of each protein in the A band varied with sarcomere length. Antiparamyosin was bound at the lateral margins of the A bands in long (~ 10.0 µ) and intermediate (~ 7.0 µ) length sarcomeres, and also in a thin line in the central A bands of sarcomeres, 7.0–~6.0 µ. Antiparamyosin stained the entire A bands of short sarcomeres (<6.0). Conversely, antimyosin stained the entire A bands of long sarcomeres, showed decreased intensity of central A band staining except for a thin medial line in intermediate length sarcomeres, and was bound only in the lateral A bands of short sarcomeres. These results are consistent with a model in which paramyosin comprises the core of the thick filament and myosin forms a cortex. Differential staining observed using antiparamyosin and antimyosin at various sarcomere lengths and changes in A band lengths reflect the extent of thick-thin filament interaction and conformational change in the thick filament during sarcomeric shortening.     

大鼠颌下腺及其离体培养细胞脱氢表雄酮（DHEA）的免疫组织化学定位

用免疫组织化学ABC法,研究了颌下腺及无血清培养的颌下腺上皮细胞DHEA的定位,结果显示,大鼠颌下腺的浆液性腺泡的上皮细胞及各级导管上皮细胞均呈DHEA免疫反应阳性,无血清培养腺上皮细胞也呈DHEA免疫反应阳性,阳性物质分布于胞质,胞核呈阴性反应,此结果提示：大鼠颌下腺能自身合成DHEA,DHEA对消化功能可能具有重要的调节作用。