Amino Acid Transport into Cultured Tobacco Cells: II. EFFECT OF CALCIUM

The effects of calcium ions on lysine transport into cultured Wisconsin-38 tobacco cells were examined. In the presence of calcium, lysine was transported at a relatively low rate for 30 to 40 minutes followed by a period of increasing rates and subsequent stabilization at a higher rate after 2 to 3 hours. In the absence of calcium, transport was uniformly low.     

Regulation of Sulfate Assimilation in Tobacco Cells: EFFECT OF NITROGEN AND SULFUR NUTRITION ON SULFATE PERMEASE AND O-ACETYLSERINE SULFHYDRYLASE

The effect of nitrogen and sulfur nutrition on sulfate permease and O-acetylserine sulfhydrylase was studied in tobacco cells.     

降低外淋巴钙浓度对耳蜗电位的影响

本实验在人工灌流条件下降低鼓阶外淋巴钙的浓度,观察其对听神经动作电位（ＣＡＰ）、耳蜗微音器电位（ＣＭ）以及蜗内直流电位（ＥＰ）的影响,以分析钙离子的作用机制。无钙液鼓阶灌流使ＣＡＰ、ＣＭ幅度可逆地下降,但不明显地改变ＣＡＰＩ／Ｏ曲线的非线性特征。无钙外淋巴灌流并不改变ＥＰ以及用缺氧法得到的负相ＥＰ（Ｎ－ＥＰ）,但使ＥＰ对强声给－撤时的快速变化趋于消失。本文讨论了这些耳蜗生物电改变所提示的钙离子作用的可能机制。     

钙调素拮抗剂与胞内钙离子浓度对离体培养的人蜕膜细胞活力的影响

应用妊娠 6～ 8周的人工流产蜕膜组织为材料进行离体人工蜕膜细胞培养 ,观察两种化学结构完全不同的特异性钙调素拮抗剂——三氟拉嗪、蝙蝠葛碱衍生物 ,以及 Ca2 + 螯合剂—— EGTA和 Ca2 +载体—— A2 3187对蜕膜细胞活力的影响。结果表明 :三氟拉嗪、蝙蝠葛碱衍生物与 EGTA对蜕膜细胞活力的抑制作用与药物的浓度及其作用时间密切相关 ,随着浓度加大及作用时间延长而增强。≥ 1 5 μmol/L的三氟拉嗪或≥ 2 5 μmol/L的蝙蝠葛碱衍生物 ,或2 mmol/L的 EGTA均可明显抑制蜕膜细胞活力。三氟拉嗪和蝙蝠葛碱衍生物作用 96小时后分别使细胞活力降至对照组的 8.7%和 1 2 .0 % ,EGTA作用 72小时后降至对照组的 2 8.6%。6μmol/L的 A2 3187可一定程度地刺激细胞活力上升 ,但这种刺激效应随作用时间的延长而逐渐减弱。EGTA可增强三氟拉嗪对细胞活力的抑制作用。由此推测 ,Ca2 + -钙调素系统可能直接参与子宫蜕膜的发育和维持 ,并在其中发挥重要作用。这可能也是钙调素拮抗剂抗早孕的主要机理之一。     

Amino Acid Transport into Cultured Tobacco Cells: I. LYSINE TRANSPORT

Lysine transport into suspension-cultured Wisconsin-38 tobacco cells was observed. Uptake was linear (up to 90 minutes) with respect to time and amount of tissue only after 4 to 6 hours preincubation in calcium-containing medium. The observed cellular accumulation of lysine was against a concentration gradient and not due to exchange diffusion. Transport was stimulated by low pH and characterized by a biphasic uptake isotherm with two K(m) values for lysine. System I (K(m) approximately 5 x 10(-6) molar; V(max) approximately 180 nanomoles per gram fresh weight per hour) and system II (K(m) approximately 10(-4) molar; V(max) approximately 1900 nanomoles per gram fresh weight per hour) were inhibited by N-ethylmaleimide and a variety of respiratory inhibitors. This inhibition was not due to increased efflux. In antagonism experiments, system I was inhibited most effectively by basic amino acids, followed by the sulfur amino acids. System I was only slightly inhibited by the neutral and aromatic amino acids and was not inhibited by the acidic amino acids aspartic and glutamic acids. Transport by system II was inhibited by all of the tested amino acids (including aspartic and glutamic acids) and analogs; however, this system was not inhibited by d-arginine. Neither system was strongly inhibited by d-lysine or the lysine analog S-2-aminoethyl-l-cysteine. Arginine was shown to be a competitive inhibitor of both systems with values for K(i) similar to the respective K(m) values.These studies suggest the presence of at least two amino acid permeases in W-38 tobacco cells.     

EFFECTS OF CALCIUM ION CONCENTRATION ON THE DEGENERATION OF AMPUTATED AXONS IN TISSUE CULTURE

Light and electron microscope studies were conducted on the nature of the degenerative changes in amputated nerve fibers of cultured rat sensory ganglia and on the effects of media with differing calcium concentrations upon these changes. With glucose-enriched Eagle's media (MEM) containing 1.6 mM calcium, the amputated myelinated and unmyelinated axons undergo a progressive granular disintegration of their axoplasm with collapse and fragmentation of myelin sheaths between 6 and 24 h after transection. With MEM containing only 25–50 µM calcium, the granular axoplasmic degeneration does not occur in transected fibers and they retain their longitudinal continuity and segmental myelin ensheathment for at least 48 h. Addition of 6 mM EGTA to MEM (reducing the estimated Ca⁺⁺ below 0.3 µM) results in the structural preservation of both microtubules and neurofilaments within transected axons. A transient focal swelling of amputated axons occurs, however, in cultures with normal and reduced calcium. These observations suggest that an alteration in the permeability of the axolemma is a crucial initiating event leading to axonal degenerative changes distal to nerve transection. The loss of microtubules and neurofilaments and the associated granular alterations of the axoplasm in transected fibers appears to result from the influx of calcium into the axoplasm.     

凝血酶引致的血小板内钙,镁离子浓度及分布状态的变化

我们使用荧光探针ｆｕｒａ２、ｍａｇ－ｆｕｒａ２和ｆｌｕｏ３测定了凝血酶引致的血小板凝聚过程中细胞内钙、镁离子浓度的变化及分布状态。在０．５Ｕ／ｍｌ凝血酶作用下,血小板细胞内钙离子浓度呈双时相变化。血小板细胞群中细胞内钙离子浓度呈正态分布。伴随血小板凝聚时细胞内钙离子浓度增加,血小板细胞内游离镁离子浓度也明显增加,提示镁离子在血小板凝聚中有重要的作用。     

心肌缺氧对大鼠心肌细胞内游离钙浓度的作用

本实验用ｆｕｒａ－２ 荧光技术研究离体鼠心缺氧期间心肌细胞内游离钙浓度（Ｃａ＾２＋）增加的机制。实验结果为：（１）Ｋｒｅｂｓ－Ｈｅｎｓｅｌｅｉｔ（Ｋ－Ｈ）液缺氧灌流离体鼠心脏引起（Ｃａ＾２＋）先慢后快地增国（２）无钙液氧灌流,仍能引起（Ｃａ蓿玻黾印＃ǎ常┖ǎǎ保薄粒保埃蓿担恚铮欤蹋┑模耍纫喝毖豕嗔鳎ǎ茫幔蓿玻┑谋浠嗨莆薷埔喝毖豕嗔鳎伊秸咭鸬模ǎ茫幔蓿玻┰黾樱飨缘陀?     

EFFECTS OF TAURODEOXYCHOLATE AND CALCIUM ON CONTRACTILITY OF NEWT EGGS

Injection of taurodeoxycholate (TDOC) alone or in combination with 10⁻⁵ M Ca²⁺(Ca-EGTA buffer) into newt eggs induced a cup- or furrow-like depression on the egg surface. Reduction of the Ca²⁺ concentration inhibited the response. These findings imply that TDOC induces a cytoplasmic contraction associated with the membrane and that a micromolar Ca²⁺ concentration is required for this process. Injection of 10⁻⁵ M Ca²⁺(Ca-EGTA buffer) alone had no effect. On the basis of these findings, the roles of TDOC and Ca²⁺ in induction of contraction were discussed.     

EFFECTS OF GUAIACOL AND HEXYLRESORCINOL IN THE PRESENCE OF BARIUM AND CALCIUM

Guaiacol was applied at two spots on the same cell of Nitella. At one spot it was dissolved in 0.01 M NaCl, at the other in 0.01 M CaCl₂ or BaCl₂. The effect was practically the same in all cases, i.e. a similar change of P.D. in a negative direction, involving a more or less complete loss of P.D. (depolarization). When hexylresorcinol was used in place of guaiacol the result was similar. That Ca⁺⁺ and Ba⁺⁺ do not inhibit the effect of these organic depolarizing substances may be due to a lack of penetration of Ca⁺⁺ and Ba⁺⁺. The organic substances penetrate more rapidly and their effect is chiefly on the inner protoplasmic surface which is the principal seat of the P.D.     

不同钙离子浓度对鲎腹神经光感受器二种光碰击的影响

在生理盐水中用持续的弱光（６＊１０＾６－９＊１０＾６个光子／ｃｍ＾２）刺激鲎的腹神经光感受器细胞可得到二种类型的碰击,一种称谓“标准光碰击”或“Ｃ２碰击”,它有对称的形状,信号衰退的指０数常数大于０．０１／毫秒。     

ADAPTATION OF CUTANEOUS TACTILE RECEPTORS : VI. INHIBITORY EFFECTS OF POTASSIUM AND CALCIUM

1. Both solutions of Ringer plus fifteen times the normal K content, and solutions of Ringer plus fifteen times the normal Ca content markedly hasten the adaptation of single freely branching axon endings in frog''s skin to repetitive air puff stimuli. 2. The K effect is produced more rapidly than is that of Ca. The K effect is reversible by washing with Ringer''s solution, while the Ca effect is not. The Ca inhibition can, however, be reversed and recovery effected by washing with K rich solutions. 3. Evidence is discussed which indicates that Ca probably plays no rôle in normal adaptation, and the experiments are interpreted as substantiating the hypothesis of adaptation due to K.     

Dark Formation of Chlorophyll in Cultured Tobacco Cells

Dark accumulation of chlorophyll was demonstrated in calluscells of Nicotiana glutinosa. The chlorophyll content of dark-growncallus cells was dependent on the agar concentration in theculture medium: the content was significantly higher at lowerconcentrations of agar. Some properties of the dark-formed chlorophyllare described. (Received June 25, 1983; Accepted December 2, 1983)     

Membrane Potentials of Heterotrophically Cultured Tobacco Cells

The electrophysiological properties of the membrane of Nicotianatabacum var. Sarnsun cultured cells were determined using amicroelectrode technique in standard medium containing 1 mMKC1, 1 mM NaCl and 1 mu CaCl₂ at pH 7. Tobacco callus was derivedfrom the pith (E_m=–104.4%16.2 mV). The membrane potentialsof the callus cells did not show a symmetrical Gaussian distributionbut were scattered over a wide range. The percentage of highmembrane potential cells increased as the subculture was continueduntil about 11 months and then decreased. The response of themembrane potential to electric stimulus, ionic composition,metabolic inhibitors, sugars and amino acids was characteristicof high (E_m=–{small tilde}–160 220 mV; H-cells)and low (E_m=–80{small tilde}–90 mV; L-cells) membranepotential cells. The membrane potential of H-cells was largelydepolarized by addition of CN^–, carbonium cyanide m-chlorophenylhydrazone,decyclohexylcarbodiimide, and triphenyltin chloride and transientlydepolarized by addition of glucose, galactose, mannose or sucrose,and D-alanine, L-alanine or Llysine, but the membrane potentialof L-cells was not. (Received December 3, 1982; Accepted March 16, 1983)     

钙对萝卜幼苗抗寒性及某些生理指标的影响

研究结果表明钙能够提高萝卜幼苗的抗寒性, 同时与抗寒性有关某些生理指标也受到影响。在低温胁迫后及恢复生长后处理组的萝卜幼苗的超氧物歧化酶、过氧化物酶、抗坏血酸过氧化物酶、过氧化氢酶的活性都比对照组高;在低温胁迫后处理组的脯氨酸及抗坏血酸的含量均比对照组高, 但处理组萝卜幼苗的丙二醛含量却比对照组低。表明钙对萝卜幼苗抗寒性的影响可能与其防止膜脂过氧化有关。     

ELECTROPHORESIS OF BACTERIA AS INFLUENCED BY HYDROGEN ION CONCENTRATION AND THE PRESENCE OF SODIUM AND CALCIUM SALTS

1. We have confirmed the results of earlier workers particularly of Northrop and De Kruif in regard to the following points: (a) the general tendency of the bacterial cell when suspended in distilled water near the zone of neutrality to move toward the anode of an electrical field; (b) the fact that the migration of bacterial cells in the electrical field is a function of the reaction of the menstruum. The curve obtained by plotting velocity of migration against pH passes through an isoelectric point at about pH 3.0, at greater acidity the direction of migration becomes reversed (toward the cathode) and in still more acid solution (pH = 1.0) again disappears; while at reactions less acid than pH 3.0 the velocity is toward the anode and increases with increasing alkalinity; (c) the fact that neutral salts depress the velocity of migration, calcium salts being much more effective than sodium salts of the same concentration. 2. We further find: (a) that on the extreme alkaline side of the curve of velocity of migration plotted against pH a maximum value is reached at about pH 10 with a fall at about pH 12.0 which in many experiments reaches an isopotential point; (b) that the depressing effect of salts is accompanied by a general shifting of the curve of migration velocity so that a maximum velocity (of course absolutely less than that manifest in the absence of salts) appears at about pH 7.0 and an abolition of velocity at pH 9.0 to 10.0; (c) that an apparent "antagonistic" effect is indicated between CaCl₂ and NaCl, the presence of a certain concentration of the latter salt diminishing to a slight but definite degree the depressing effect produced by the former; (d) that heat-killed bacterial cells exhibit essentially the same curve of migration velocity as that of the living cells; (e) that bacterial spores exhibit the same general curve of migration velocity as vegetative cells, although the actual velocity is apparently slightly less. 3. All of the observed phenomena appear to be in accord with the assumption that marked differences in dielectric constants did not appear under the conditions studied and if this assumption be granted the results are in accord with the fundamental postulates of the Donnan equilibrium as applied to the explanation of the origin of potential difference between a bacterial cell and its enveloping menstruum. It is possible but not at all certain that the phenomenon of antagonism may require the introduction of additional assumptions for its explanation. Professor Donnan and other investigators have clearly understood the importance of applying the concept of membrane equilibria in the elucidation of physiological phenomena. Our findings add to the numerous vindications favoring this view and emphasize the importance of further study of membrane equilibria in bacterial suspensions. We have pointed out that certain potential differences between bacteria and their menstrua are apparently associated with some of the phenomena of viability. Viability and potential differences may, however, under certain conditions vary quite independently as evidenced by the fact that normal rates of migration are demonstrable after the cells have been killed by heat. Thus, considerable caution must be exercised in relating the existence of these charges to the metabolism of the cell.     

Ca2+对小麦种根及其根毛生长发育的影响

低浓度的ＣａＣｌ２对小麦种根生长、根毛发生和生长无明显影响,高浓度的ＣａＣｌ２（０．１ｍｏｌ／Ｌ）对种根和根毛生长有抑制作用,但不影响根毛的发生。Ｃａ２＋专一性螯合剂ＥＧＴＡ抑制种根生长和根毛的发生及生长,添加一定浓度的外源ＣａＣｌ２,这种抑制作用可被消除。ＣａＭ抑制剂ＴＦＰ（三氟拉嗪）和ＣＰＺ（氯丙嗪）对种根生长、根毛发生和生长均有抑制作用,添加外源ＣａＭ可减弱或消除这种抑制作用。     

An Electrophysiological Study of Photomixotrophically Cultured Tobacco Cells

Membrane potential properties of photomixotrophically culturedgreen tobacco cells with chloroplasts were studied in comparisonwith white tobacco cells without chloroplasts. In the dark therewas almost no difference in their membrane potential properties.In the light some of the green cells showed a light-inducedpotential change (LPC), but other green cells did not, nor didthe white cells. Our results indicate that the green cells arecomposed of two kinds of cells, one that shows the LPC and onethat does not. (Received October 5, 1983; Accepted May 10, 1984)     

Photosynthetic Characteristics of Photoautotrophically Cultured Cells of Tobacco

The photosynthetic characteristics of photoautotrophically culturedcells of tobacco (Nicotiana tabacum cv. Samsun NN) as well asthose of photomixotrophically cultured cells and green leaveswere investigated. Analyses revealed that on a fresh weightbasis cultured tobacco cells had lower chlorophyll contentsthan cells of green leaves. The chlorophyll content per chloro-plast,however, was almost the same in both types of cell, and thechloroplast number per cell accounted for only small differencesin the cellular chlorophyll content. This indicates that thelarger cell volume of cultured cells is the main factor in thedifference in the chlorophyll content of these cells. Photosynthetic activity measurements also showed differencesin the chloroplasts of cultured and leaf cells. The maximumactivities of photosystem I and the Hill reaction for the culturedcells were about half those for leaf cells on a per unit chlorophyllbasis. Moreover, photo-autotrophic cells had relatively constantphotosystem I and Hill reaction activities during growth; whereas,on a fresh weight basis these activities in leaf cells reflecteddevelopmental changes in the chlorophyll content. Lithium dodecyl sulfate-polyacrylamide gel electrophoresis showedqualitatively similar thylakoid polypeptide compositions forcultured and leaf cells at all stages of growth even thoughthere were quantitative decreases in the contents of severalpolypeptides in the cultured green cells (especially in photomixotrophiccells) in comparison to the polypeptide contents of tobaccoleaves. We speculate that the lower photosynthetic activityof the cultured cells may be caused by this reduction in thecontents of certain thylakoid polypeptides. (Received November 14, 1988; Accepted June 19, 1989)