Irradiations of rabbit myofibrils with an ultraviolet microbeam. I. Effects of ultraviolet light on the myofibril components necessary for contraction

Glycerinated rabbit psoas myofibrils, F-actin, and myofibril ghosts were irradiated with ultraviolet light (UV) to investigate how UV blocks myofibril contraction. Myofibril contraction is most sensitive to 270- and 290-nm wavelength light. We irradiated I and A bands separately with 270- and 290-nm wavelength light using a UV microbeam and constructed dose-response curves for blocking sarcomere contraction. For both wavelengths, irradiations of A bands required less energy per area to block contraction than did irradiations of I bands, suggesting that the primary effects of both 270- and 290-nm wavelength light in stopping myofibril contraction are on myosin. We investigated whether the primary effect of UV in blocking I-band contraction is the depolymerization of actin by comparing the relative sensitivities of I-band contraction, F-actin depolymerization, and thin filament depolymerization to 270- and 290-nm light. We also compared the dose of UV required to depolymerize F-actin in solution with the dose needed to block I-band contraction and the dose required to alter thin filament structure in myofibril ghosts. The results confirm that UV blocks I-band contraction by depolymerizing actin. We discuss how the results might be relevant to the hypothesis that an actomyosin-based system is involved in chromosome movement.

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Action spectrum for changes in spindle fibre birefringence after ultraviolet microbeam irradiations of single chromosomal spindle fibres in crane-fly spermatocytes

Journal of Cell Science ◽

10.1242/jcs.62.1.1 ◽

1983 ◽

Vol 62 (1) ◽

pp. 1-25

Author(s):

P.J. Sillers ◽

A. Forer

Keyword(s):

Ultraviolet Light ◽

Action Spectrum ◽

The Other ◽

Spindle Fibre ◽

Chromosome Movement ◽

Ultraviolet Microbeam ◽

Two Peaks ◽

Wavelength Light ◽

Spindle Fibres ◽

In Cells

Single chromosomal spindle fibres in Nephrotoma suturalis (crane-fly) spermatocytes in metaphase and anaphase were irradiated with monochromatic ultraviolet light focussed to a 2 micrometer spot. In cells in both metaphase and anaphase either the birefringence of the irradiated spindle fibre was altered in the irradiated region, or there was no change, depending on the dose and wavelength of ultraviolet light used for the irradiation. When there was an area of reduced birefringence (ARB), it moved poleward regardless of whether the associated chromosome moved poleward. When cells were irradiated in early metaphase they remained in metaphase until the ARB reached the pole. In some cells irradiated in late metaphase the chromosomes began anaphase before the ARB reached the pole; in many such cells anaphase was abnormal in that all six half-bivalents separated at the start of anaphase but none moved polewards. In all cases the ARB moved poleward at the same speed as subsequent chromosome movement; that is, at about 0.8 micrometer/min. In cells irradiated in anaphase, spindle fibre birefringence was reduced independently of blockage of chromosome movement. Because birefringence and movement were altered independently there were four classes of results: (1) in some cases there was no effect on the movement of the chromosome associated with the irradiated spindle fibre and no effect on the birefringence of the irradiated spindle fibre. (2)In some cases, primarily with 260 nm wavelength light, there was no effect on the movement of the chromosome associated with the irradiated spindle fibre and there was an effect on the birefringence of the irradiated spindle fibre. (3) In some cases, primarily with 290 nm wavelength light, there was an effect on the movement of the chromosome associated with the irradiated spindle fibre and no effect on the birefringence of the irradiated spindle fibre. (4) In some cases, primarily with 270 nm and 280 nm wavelength light, there was an effect on the movement of the chromosomes associated with the irradiated spindle fibre and there was an effect on the birefringence of the irradiated spindle fibre. The action spectrum for reducing spindle fibre birefringence in crane-fly spermatocytes had two peaks, one at 260 nm and the other, less sensitive, at 280 nm. For irradiations at 270 nm, 280 nm and 290 nm, five to fifty times more energy was needed to reduce spindle fibre birefringence than to stop chromosome movement, but for irradiations at 260 nm five times less energy was needed to reduce spindle fibre birefringence than to stop chromosome movement. The action spectrum for reducing spindle fibre birefringence is quite different from that for stopping chromosome movement.

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Analysis of chromosome movement in crane fly spermatocytes by ultraviolet microbeam irradiation of individual chromosomal spindle fibres. I. General results

Canadian Journal of Biochemistry ◽

10.1139/o81-107 ◽

1981 ◽

Vol 59 (9) ◽

pp. 770-776 ◽

Cited By ~ 7

Author(s):

Peggy J. Sillers ◽

Arthur Forer

Keyword(s):

Ultraviolet Light ◽

Monochromatic Light ◽

The Other ◽

Spindle Fibre ◽

Chromosome Movement ◽

Opposite Pole ◽

Other Hand ◽

Ultraviolet Microbeam ◽

The Difference ◽

Spindle Fibres

Single chromosomal spindle fibres in anaphase Nephrotoma ferruginea (crane fly) spermatocytes were irradiated with monochromatic ultraviolet light focussed to a 4-μm spot by means of an ultraviolet microbeam apparatus. The movement of the half-bivalent associated with the irradiated spindle fibre was either unaffected or the half-bivalent stopped moving; i.e., the effect was all-or-none. When the half-bivalent associated with the irradiated spindle fibre did stop moving, the partner half-bivalent moving towards the opposite pole (i.e., the half-bivalent with which the first half-bivalent was previously paired) also stopped moving: all other half-bivalents moved normally. In over 90% of the 69 cases the movements of the two half-bivalents were only temporarily blocked; when movement resumed both half-bivalents resumed movement at the same time, after stoppage times ranging from 2 min to more than 15 min. In a few cases the half-bivalents never resumed poleward motion.When half-bivalents that had stopped movement finally resumed movement they often did not reach the poles; i.e., they "lagged" and remained separate from the other chromosomes. This result occurred only in spermatocytes of N. ferruginea. In spermatocytes of N. suturalis or N. abbreviata, on the other hand, the stopped half-bivalents did not lag but always reached the poles.Half-bivalent pairs that stopped moving in N. ferruginea spermatocytes did so for shorter times than did those previously reported (after irradiation of chromosomal spindle fibres) in N. suturalis spermatocytes. We suggest that the difference is due to our use of monochromatic ultraviolet light as opposed to the previous use of heterochromatic ultraviolet light. We assume that different wavelengths of monochromatic light produce different effects, that any given monochromatic irradiation produces only one effect (albeit different effects at different wavelengths), but that heterochromatic irradiations can produce multiple effects.Irradiation of the interzone (between separating half-bivalents) had no effect on the chromosome-to-pole movements of the half-bivalents. Therefore the stoppage of movement of half-bivalent pairs is specific for irradiation of chromosomal spindle fibres. On the other hand, irradiation of the interzone often blocked pole-to-pole elongation.

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Irradiations of rabbit myofibrils with an ultraviolet microbeam. II. Phalloidin protects actin in solution but not in myofibrils from depolymerization by ultraviolet light

Biochemistry and Cell Biology ◽

10.1139/o87-047 ◽

1987 ◽

Vol 65 (4) ◽

pp. 376-385 ◽

Cited By ~ 12

Author(s):

Paula Wilson ◽

Esther Fuller ◽

Arthur Forer

Keyword(s):

Ultraviolet Light ◽

Staining Pattern ◽

Apparent Contradiction ◽

Thin Filaments ◽

Rhodamine Phalloidin ◽

Fluorescence Image ◽

Rabbit Psoas ◽

Ultraviolet Microbeam ◽

Presence And Absence ◽

Over Time

We tested whether phalloidin protects actin in myofibrils from depolymerization by ultraviolet light (UV). I bands in glycerinated rabbit psoas myofibrils were irradiated with a UV microbeam in the presence and absence of phalloidin. We used the retention of contractility of the irradiated I band as the assay for protection of actin by phalloidin, since previous experiments indicated that UV blocks contraction of an irradiated I band by depolymerizing the thin filaments. The I bands of myofibrils incubated in phalloidin were as sensitive to UV as control I bands, indicating that phalloidin did not protect the thin filaments. However, phalloidin did protect F-actin in solution from depolymerization by UV. This apparent contradiction between F-actin in myofibrils and F-actin in solution was resolved by observing unirradiated myofibrils that were stained with rhodamine–phalloidin. It was found that phalloidin does not bind uniformly to the thin filaments, though as the fluorescence image is observed over time the staining pattern changes until it does appear to bind uniformly. We conclude that phalloidin does not protect F-actin in myofibrils from depolymerization by UV because it does not bind uniformly to the filaments.

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Dose-response Curves in the Fibrin Plate Assay Fibrinolytic Activity of Proteases

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647705 ◽

1974 ◽

Vol 32 (02/03) ◽

pp. 356-365 ◽

Cited By ~ 12

Author(s):

F Haverkate ◽

D. W Traas

Keyword(s):

Dose Response ◽

Fibrinolytic Activity ◽

Enzyme Concentration ◽

Response Curves ◽

Agar Gel ◽

Porcine Tissue ◽

Plate Assay ◽

Fibrin Plate ◽

Different Types ◽

Dose Response Curves

SummaryIn the fibrin plate assay different types of relationships between the dose of applied proteolytic enzyme and the response have been previously reported. This study was undertaken to determine whether a generally valid relationship might exist.Trypsin, chymotrypsin, papain, the plasminogen activator urokinase and all of the microbial proteases investigated, including brinase gave a linear relationship between the logarithm of the enzyme concentration and the diameter of the circular lysed zone. A similar linearity of dose-response curves has frequently been found by investigators who used enzyme plate assays with substrates different from fibrin incorporated in an agar gel. Consequently, it seems that this linearity of dose-response curves is generally valid for the fibrin plate assay as well as for other enzyme plate bioassays.Both human plasmin and porcine tissue activator of plasminogen showed deviations from linearity of semi-logarithmic dose-response curves in the fibrin plate assay.

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Factor VII Clotting Assay: Influence of Different Thromboplastins and Factor VII-Deficient Plasmas

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647476 ◽

1991 ◽

Vol 65 (02) ◽

pp. 160-164 ◽

Cited By ~ 27

Author(s):

Marina Poggio ◽

Armando Tripodi ◽

Guglielmo Mariani ◽

Pier Mannuccio Mannucci ◽

Keyword(s):

Heart Disease ◽

Ischemic Heart Disease ◽

Dose Response ◽

Response Curve ◽

Ischemic Heart ◽

Factor Vii ◽

Response Curves ◽

Assay Sensitivity ◽

Clinical Laboratories ◽

Coagulant Activity

SummaryBeing a putative predictor of ischemic heart disease, the measurement of factor VII (FVTI) coagulant activity will be presumably requested to clinical laboratories with increasing frequency. To assess the influence on FVII assays of different thromboplastins and FVII-deficient plasmas we compared performances of all possible combinations of 5 thromboplastins and 6 deficient plasmas. The reproducibility of the clotting times of the dose-response curves for human and rabbit thromboplastins were acceptable (CV lower than 7%), whereas bovine thromboplastin had a higher CV. Reproducibility was very similar for all deficient plasmas when they were used in combination with a given thromboplastin. Responsiveness of the dose-response curve did not depend on the deficient plasma but rather on the thromboplastin: one rabbit thromboplastin was the least responsive, the bovine thromboplastin the most responsive, the human and the remaining two rabbit thromboplastins had intermediate responsiveness. Assay sensitivity to cold-activated FVII varied according to the thromboplastin: the bovine thromboplastin was the most sensitive, the human thromboplastin the least sensitive, of the three rabbit thromboplastins two were relatively sensitive, one was almost insensitive. In conclusion, our results indicate that thromboplastin rather than deficient plasma is the crucial factor in the standardization of FVII assay.

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OVARIAN ASCORBIC ACID DEPLETION TEST FOR LUTEINIZING HORMONE

Acta Endocrinologica ◽

10.1530/acta.0.0560619 ◽

1967 ◽

Vol 56 (4) ◽

pp. 619-625 ◽

Cited By ~ 1

Author(s):

Hans Jacob Koed ◽

Christian Hamburger

Keyword(s):

Ascorbic Acid ◽

Luteinizing Hormone ◽

Dose Response ◽

Response Curve ◽

Dose Response Curve ◽

Human Origin ◽

Response Curves ◽

Significant Difference ◽

The Mean ◽

Different Levels

ABSTRACT Comparison of the dose-response curves for LH of ovine origin (NIH-LH-S8) and of human origin (IRP-HMG-2) using the OAAD test showed a small, though statistically significant difference, the dose-response curve for LH of human origin being a little flatter. Two standard curves for ovine LH obtained with 14 months' interval, were parallel but at different levels of ovarian ascorbic acid. When the mean ascorbic acid depletions were calculated as percentages of the control levels, the two curves for NIH-LH-S8 were identical. The use of standards of human origin in the OAAD test for LH activity of human preparations is recommended.

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BIOASSAY OF THYROID HORMONE USING HYPOTHYROID TADPOLES

Acta Endocrinologica ◽

10.1530/acta.0.0410143 ◽

1962 ◽

Vol 41 (1) ◽

pp. 143-153 ◽

Cited By ~ 2

Author(s):

U. Henriques

Keyword(s):

Thyroid Hormone ◽

Dose Response ◽

Developmental Rate ◽

Response Curves ◽

Sodium Salts ◽

Dose Response Curves

ABSTRACT A bioassay of thyroid hormone has been developed using Xenopus larvae made hypothyroid by the administration of thiourea. Only tadpoles of uniform developmental rate were used. Thiourea was given just before the metamorphotic climax in concentrations that produced neoteni in an early metamorphotic stage. During maintained thiourea neotoni, 1-thyroxine and 1-triiodothyronine were added as sodium salts to the water for three days and at the end of one week the stage of metamorphosis produced was determined. In this way identical dose-response curves were obtained for the two compounds. No qualitative differences between their effects were noted except that triiodothyronine seemed more toxic than thyroxine in equivalent doses. Triiodothyronine was found to be 7–12 times as active as thyroxine.

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ÜBER DIE ANTAGONISTISCHE WIRKUNG VON TRIJODTHYRONIN UND PROGESTERON AUF DEN PREDNISOLONEFFEKT AM EPIPHYSENKNORPEL (PRÜFUNG DER REISSFESTIGKEIT)

Acta Endocrinologica ◽

10.1530/acta.0.0420029 ◽

1963 ◽

Vol 42 (1) ◽

pp. 29-38 ◽

Cited By ~ 12

Author(s):

L. Ther ◽

H. Schramm ◽

G. Vogel

Keyword(s):

Tensile Strength ◽

Dose Response ◽

Single Injection ◽

Relative Effectiveness ◽

Prednisolone Acetate ◽

Response Curves ◽

Deoxycorticosterone Acetate ◽

Weight Range ◽

Diuretic Action ◽

Maximal Load

ABSTRACT Longitudinal tension exerted on a killed rat's femur results always in rupture on the same place viz: the distal epiphyseal line. The power required to produce this effect can be measured with a simple dynamometer. The tensile strength increases with advancing age and during exsiccation (thirst). In untreated rats (weight range 100–120 g) this value amounts to about 1.5–1.8 kg. Tensile strength of the femoral epiphyseal line may be influenced by hormones. Corticosteroids such as prednisolone acetate and cortisol acetate increase the level of maximal load to more than 2.0 kg. In general the effect of a single injection reaches a climax after 48 hours. Dose-response-curves can be established and relative effectiveness may be gathered therefrom. Deoxycorticosterone acetate likewise increases tensile strength in spite of the fact that contrary to prednisolone exhibiting diuretic action this compound has a water retaining effect. Triiodothyronine and progesterone when given alone produce a slight decrease in tensile strength. The effect of prednisolone when given simultaneously with one of these two compounds is antagonized: this phenomenon is more pronounced in the case of triiodothyronine.

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