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PubMed1987

A Tight-Seal Whole Cell Study of the Voltage-Dependent Gating Mechanism of K-Channels of Protoplasmic Droplets of Chara corallina.

The biophysical properties of voltage-dependent K(+)-channels of protoplasmic droplets of Chara corallina Klein ex Willd., em, R.D.W. were investigated using the tight-seal whole cell method. Two potassium currents were observed in voltage-clamp mode and they can be used to explain the transient membrane potential time course observed in current-clamp mode. The K(+)-channels are identified by the effect of tetraethylammonium chloride which blocks both currents. A two-state, constant dipole moment model is used to fit the voltage-conductance curve. From this model the minimum equivalent gating charge involved in the gating mechanism of K(+)-channels of Chara can be estimated.

Journal Article↗
PubMed1987

Actin of chara giant internodal cells: a single isoform in the subcortical filament bundles and a larger, immunologically related protein in the chloroplasts.

Internodal cells of Chara corallina Klein ex. Wild have been studied to determine the number of actin isoforms they contain and whether actin occurs at locations in the cortical cytoplasm outside the filament bundles. A monoclonal antibody to chicken actin is specific for actin in numerous animal cells but binds to two Chara proteins after their separation by two-dimensional polyacrylamide gel electrophoresis. One protein resembles known actins in relative molecular mass (43,000-M(r)) and isoelectric point (5.5) while the other is distinctly different (58,000-M(r), isoelectric point = 4.8). Because it is indetectable in cells whose actin bundles have been extracted, the 43,000-M(r) protein is assigned to the bundles and concluded to be rare or absent in the remaining cortical cytoplasm. The 58,000-M(r) protein, in contrast, does not extract with the actin bundles. It was localized within the chloroplasts by immunofluorescence and by the dependence of proteolysis on the permeabilization of the chloroplast envelope.

Journal Article↗
PubMed1988

Degradation of Proteins Artificially Introduced into Vacuoles of Chara australis.

When an exogenous protein, bovine serum albumin, was introduced into the vacuole of a Chara australis internodal cell, it was degraded with time. This degradation proceeded only in the vacuole as far as could be observed by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Degradation was inhibited by protease inhibitors such as antipain and leupeptin. Endogenous proteins introduced into the vacuole were also degraded there. Furthermore, intravacuolar cytoplasmic drops, which were often formed by cell ligation, seemed to be degraded in the vacuole. However, bovine serum albumin degradation did not proceed when mixed with isolated vacuolar sap. These results show that the vacuole in the Chara internodal cell has the capacity to degrade cellular proteins, but that cytoplasmic support is needed for this degrading activity to be maintained.

Journal Article↗
PubMed1989

Measurement of the Cytoplasmic and Vacuolar Buffer Capacities in Chara corallina.

The cytoplasm and the vacuole were isolated from internodal cells of Chara corallina by using the intracellular perfusion technique, and their buffer capacities (beta(i)) were determined from the titration curves. The pH of the isolated vacuolar sap was 5.19 +/- 0.029 (mean +/- standard error). At this pH, beta(i) was minimal and amounted to 0.933 +/- 0.11 millimoles H(+)/pH unit/liter vacuolar sap. The pH of isolated cytoplasm was 7.22 +/- 0.028. beta(i) was minimal in this pH region and amounted to 14.2 +/- 0.80 millimoles H(+)/pH unit/liter cytoplasm. When 1% (volume/volume) Triton X-100 was added to the cytoplasmic solution to permeabilize the subcellular organelles, the cytoplasmic pH increased to 7.32 +/- 0.026, where beta(i) was 20.35 +/- 2.66 millimoles H(+)/pH unit/liter cytoplasm. This shows that alkaline subcellular compartments exist in the cytoplasm and also that the cytoplasmic pH before adding Triton X-100 may represent the cytosolic pH. These data indicate that the pH values of the cytoplasm and the vacuole are regulated at the values where the beta(i) values are minimal. This suggests that ATP- and inorganic pyrophosphate-dependent H(+) pumps in the plasma membrane and the tonoplast could efficiently regulate the pH of both cytoplasm and vacuole in Chara internodal cells.

Journal Article↗
PubMed1993

The protonema of Chara fragilis Desv.: regenerative formation, photomorphogenesis, and gravitropism.

When exposed to constant white light for four weeks, isolated nodes of Chara fragilis Desv. regenerated side branches, rhizoids, and multicellular protonemata, the latter being similar to those germinated from oospores. When kept in darkness the nodes developed protonemata exclusively. These were single-celled, colourless, and tip-growing and, with the light microscope, they looked like rhizoids. Upon exposure to blue light, but not to red or far-red, the growth rates of the protonemata rapidly declined, the cell apices swelled, and the nucleus migrated acropetally. Within 24 h the cells went through the first of a series of divisions resulting in the formation of multicellular protonemata. When returned to darkness after a blue light pulse of 5 h the cell divisions proceeded normally, but the protonemata showed etiolated growth. While growth of the internode was drastically promoted, the development of the multicellular apex and the lateral initial were suppressed. Both uni- and multicellular etiolating protonemata showed negative gravitropism but were phototropically, insensitive. It is argued that the single-celled protonema is an organ specialized for the penetration of mud covering the nodes or oospores of Chara and thus serves to search for light, comparable to etiolated hypocotyls and stems in seedlings of higher plants.

Cell Division↗
PubMed1976

Cytoplasmic streaming in Chara corallina studied by laser light scattering.

An apparatus is described by means of which the power versus frequency spectrum of the photomultiplier current can be obtained for laser light scattered by streaming cytoplasm in the algal cell Chara corallina. A Doppler peak is noted in the spectrum which is abolished when cytoplasmic streaming is arrested by electrical stimulation. For 5 cells of Chara, this simple laser-Doppler velocimeter gave streaming velocities (46-7 mum s-1, S.D. +/- 4-8 at 20 degrees C) similar to those obtained for the same cells using the light microscope (44-3 mum s-1, S.D. +/- 5-3 at 20 degrees C). A narrow distribution of streaming velocities is indicated. The technique described provides a rapid, quantitative assay of the in vivo rheological properties of cytoplasm.

Chlorophyta↗
PubMed1990

The Vividness of Visual Imagery Questionnaire: commentary on the Marks-Chara debate.

The purpose of this article is to comment on a recent debate between Chara and Marks over the construct validity of the Vividness of Visual Imagery Questionnaire (VVIQ). Following a description of their positions, it is argued that, although some of Chara's logical criticisms merit attention, much of his empirical work is methodologically unsound. However, in view of the nature of construct validity itself, it is also suggested that Marks ought to specify more clearly his concept of imagery vividness so critical experimental predictions can be made.

Arousal↗
PubMed2001

Through pore diameter in the cell wall of Chara corallina.

Determination of pore size of the cell wall of Chara corallina has been made by using the polyethylene glycol (PEG) series as the hydrophilic probing molecules. In these experiments, the polydispersity of commercial preparation of PEGs was allowed for. The mass share (gamma(p)) of polyethylene glycol preparation fractions penetrating through the pores was determined using a cellular 'ghost', i.e. fragments of internodal cell walls filled with a 25% solution of non-penetrating PEG 6000 and tied up at the ends. In water, such a 'ghost' developed a hydrostatic pressure close to the cell turgor which persisted for several days. The determination of gamma(p), for polydisperse polyethylene glycols with different average molecular mass (M) was calculated from the degree of pressure restoration after water was replaced by a 5-10% polymer solution. Pressure was recorded using a dynamometer, which measures, in the quasi-isometric mode, the force necessary for the partial compression of the 'ghost' in its small fragment. By utilizing the data on the distribution of PEG 1000, 1450, 2000, and 3350 fractions over molecular mass (M), it was found that gamma(p), for these polyethylene glycols corresponded to the upper limit of ML=800-1100 D (hydrodynamic radius of molecules, r(h)=0.85-1.05 nm). Thus, the effective diameter of the pores in the cell wall of Chara did not exceed 2.1 nm.

Biomechanical Phenomena↗
PubMed1991

The density of the cell sap and endoplasm of Nitellopsis and Chara.

We measured the densities of the cell sap, endoplasm and cell wall of Nitellopsis obtusa and Chara corallina using interference microscopy, refractometry, immersion refractometry, equilibrium sedimentation and chemical microanalysis techniques. These values are important for the determination of many rheological properties of the cytoplasm as well as for understanding buoyancy regulation, dispersal mechanisms and how cells respond to gravity. The average densities of the cell sap, endoplasm and cell wall are 1,006.9, 1,016.7 and 1,371 kg m-3 for Nitellopsis and 1,005.0, 1,013.9, and 1,355.3 kg m-3 for Chara.

Cell Wall↗
PubMed1993

Structural polarity in the Chara rhizoid: a reevaluation.

The Chara rhizoid is a useful model system to study gravitropism since all phases of gravitropism occur in a single cell. Despite years of study, a complete description of the distinctive ultrastructure of Chara rhizoids is not available. Therefore, in this paper, we reevaluate the ultrastructural features of vertically grown rhizoids, which have a structural polarity consisting of seven distinct zones. We also characterize the apical vesicles and the cell wall in these rhizoids by using antibodies against pectic polysaccharides. These studies demonstrate that the cell wall consists of two pectinaceous domains and that a distinct population of apical vesicles contain methyl esterified pectin.

Antibodies, Monoclonal↗
PubMed1997

The effect of the external medium on the gravity-induced polarity of cytoplasmic streaming in Chara corallina (Characeae).

Gravity induces a polarity of cytoplasmic streaming in vertical internodal cells of Chara such that the downwardly directed stream moves faster than the upwardly directed stream. In order to determine whether the statolith theory (in which intracellular sedimenting particles are responsible for gravity sensing) or the gravitational pressure theory (in which the entire protoplast acts as the gravity sensor) best explain the gravity response in Chara internodal cells, we controlled the physical properties of the external medium, including density and osmolarity, with impermeant solutes and examined the effect on the polarity of cytoplasmic streaming. As the density of the external medium is increased, the polarity of cytoplasmic streaming decreases and finally disappears when the density of the external medium is equal to that of the cell (1015 kg/m3). A further increase in the density of the external medium causes a reversal of the gravity response. These results are consistent with the gravitational pressure theory of gravity sensing since the buoyancy of the protoplast is dependent on the difference between the density of the protoplast and the external medium, and are inconsistent with the statolith theory since the buoyancy of intracellular particles are unaffected by changes in the external medium.

Biophysical Phenomena↗
PubMed1979

[Proof of the subapical differential growth of the flanks in the Chara rhizoid during graviresponse].

The displacement of resin spheres attached to the tip of the gravitropically bending Chara rhizoid was measured by means of time-lapse photography. The relative growth of the flanks which at the beginning of stimulation were opposite to each other, was calculated during the earliest response time. The physically upper subapical flank section continues growing after stimulation at a decreasing rate. The growth of the opposite lower flank section is inhibited immediately after stimulation by the position of the statoliths. As soon as the statoliths are displaced in a basal direction, the growth of this section is transiently promoted. The gravitropic downward bending of the Chara rhizoid is by bowing, not by bulging.

Chlorophyta↗
PubMed2001

[Effect of avermectns on Ca(2+)-dependent chloride currents in plasmalemma of Chara corallina cells].

A natural complex of avermectins, aversectin C, and a component of this complex, avermectin A1, were shown to change the conductivity of Ca(2+)-dependent chloride channels of plasmalemma of Chara corallina cells by acting only from the outer side of the cellular membrane. Low concentrations of aversectin C and avermectin A1 increased the chloride current: K1/2 = 3.5 x 10(-5) mg/ml for the whole complex and K1/2 = 2.1 x 10(-3) mg/ml for A1. Relatively high concentrations of the compounds suppressed the chloride current: K1/2 = 2.2 x 10(-3) mg/ml for aversectin C and K1/2 = 4.2 x 10(-6) mg/ml for A1. The Hill coefficients for the interaction of avermectin A1 with the corresponding targets for stimulation and suppression of the chloride current were 2.8 and 2.5 respectively. Bicuculine, a non-specific inhibitor of the GABA alpha-receptors, did not influence stimulation of chloride currents caused by action of low concentrations of avermectins, but at the same time blocked suppression of the chloride currents associated with the action of high doses of avermectins. Avermectins A2, B1 (abamectin), B2 and 22,23-dihydroavermectin B1 (vermectin) in the concentration range studied, did not affect the chloride currents of Chara corallina cells.

Calcium↗
PubMed2004

Moyse Charas, Francesco Redi, the Viper and the Royal Society of London.

We have described the 17th century dispute on the nature of the poison of the viper and its possible use as an antidote which arose between published in Britain at that time. We have also seen the reciprocal interest of Charas and members of the Royal Society in each other's work, and how Charas settled in England for a few years at the time of discrimination against the Huguenots in France. We prefer to leave any conclusions from this story to individual readers. However the dispute seems to be part of a transition from an animate view of the world to one where empirical methods do not appear to need to invoke 'spirits'. The story also seems to illustrate the limitation of experiments there can still be disagreement. If there is a the feeling that the subject is of interest, we can broaden it to cover scientific exchanges and connections between British and French Scholars in the field of pharmaceutical chemistry in the 17th and 18th centuries. At that time a number of pharmacists and scientists moved or corresponded between the two countries influencing one another. That was when modern chemistry gradually evolved out of traditional origins.

Animals↗
PubMed1989

Determination of the inorganic pyrophosphate level and its subcellular localization in Chara corallina.

In order to determine the concentration of pyrophosphate (PPi) and its subcellular distribution in Chara corallina, a new method to concentrate PPi from cell extracts was developed. PPi was extracted and concentrated as Ca2P2O7 under alkaline conditions. The amount of PPi in the precipitate was measured using an enzyme system containing pyrophosphate:fructose-6-phosphate 1-phosphotransferase (EC 2.7.1.90) coupled to NADH oxidation in the presence of [ethylene-bis(oxyethylenenitrilo)]tetraacetic acid. The subcellular localization of PPi and inorganic phosphate (Pi) was studied using the intracellular perfusion technique. The relative volumes of the cytoplasm (6.4%) and the vacuole (93.6%) were determined by perfusing Lucifer Yellow CH into the vacuole and by assuming that the Lucifer Yellow CH dead space represented the cytoplasmic volume. The volume of the chloroplast layer was determined microscopically, and it was found that it occupied 10% of the Chara cytoplasm. PPi was present predominantly in the cytosol at a level of 193 microM, while it existed in the vacuole at a level of only 2.20 microM and less than 1 microM in chloroplasts. By contrast, Pi was distributed almost equally in the cytosol (12.0 mM), chloroplasts (16.2 mM), and the vacuole (6.70 mM). The electrochemical potential gradient across the tonoplast for H+ (delta mu H+ = -11.6 to -18.0 KJ/mol) was nearly equal to the free energy release from the hydrolysis of PPi in cytoplasm (delta Gpp = -18.9 KJ/mol), indicating that the H+-translocating inorganic pyrophosphatase can work as a H+ pump in C. corallina.

Chemical Precipitation↗
PubMed1989

The cell walls of Chara aspera Willd. (Charophyta) vegetative cells.

The ultrastructure of the vegetative cell walls of the charophyte Chara aspera Willd was studied with TEM. Thallus cells, rhizoid bulbil and rhizoidal node cells were investigated. The internodal cells transverse walls contained plasmodesmata. The longitudinal walls of the internodal cells were uniform, fibrillar, with two thin structurally distinct layers with different structure facing the cytoplasm. The outermost layers of internodal, cortical and rhizoid bulbil cells were composed of randomly orientated fibrils. The longitudinal walls of the cortical cells were helicoidal in structure. In the rhizoid bulbil cell walls, six different layers could be distinguished, but their occurrence seemed to depend on the fixation, staining and cutting procedures. A middle lamella and osmophilic deposits were found in the wall between rhizoidal node cells. The cytoplasmic structure of the internodal and cortical cells was not found to differ from other species of Chara. Charasomes were observed only in cortical cells.

Cell Wall↗
PubMed1988

Electrical tolerance of the Chara corallina plasmalemma.

[Ca2+]o controls at least three distinct electrical phenomena in the Chara plasmalemma. 1). Increase of the electrical tolerance. 2). Decrease of the G without a marked change of the Gp/Gd ratio. 3). Elevation of the threshold for excitation (data not shown). On the other hand, [Mg2+]o shows quite opposite effects. Thus, optimum [Ca2+]o and [Mg2+]o are expected in vivo. Voltage dependencies of various conductances in the Chara plasmalemma are schematically shown in Fig. 4.

Biological Transport, Active↗
PubMed1994

DNA endoreduplication, RNA and protein synthesis during growth and development of the antheridial basal cell in Chara vulgaris L.

Cytophotometric measurements of nuclear DNA contents and morphometric analyses indicate that the level of endopolyploidy plays an important role in determining the maximum size, transcriptional and translational activity that the antheridial basal cell attains during successive stages of spermatogenesis in Chara vulgaris. During the proliferative period of antheridial development, the metabolic activity of basal cell, expressed as the total incorporation of radioactive uridine and leucine was found to increase gradually with the increasing DNA C-values, yet both the synthesis of RNA and then the synthesis of proteins become reduced at the stage preceding spermiogenesis. In accordance with some earlier data [2], the obtained results seem to support the hypothesis that regulatory mechanisms of symplasmic connections between the antheridium and a thallus participate in the regulation of morphogenesis of the male sex organs in Chara.

Autoradiography↗