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Binding of insulin to solubilized insulin receptor from human placenta. Evidence for a single class of noninteracting binding sites.

Binding of 125I-insulin to insulin receptor was studied at equilibrium using plasma membrane protein, from human placenta, solubilized in Triton X-100. Measured at 1 nM 125I-insulin, the amount of radioligand bound specifically was dependent upon pH, the optimal association at 4 degrees C occurring in the pH range 8.2-8.7. The Adair equation was employed for analysis of binding isotherms generated at several pH values for concentrations of radioligand varying from 0.01-8.0 nM. Association constants derived from these analyses showed the same pH dependence described above and were found to be independent of receptor concentration. Hill plots derived from these titrations consistently yielded Hill coefficients of 1 and Scatchard plots gave virtually straight lines. Close correspondence was found between theoretical analyses and observations under all experimental conditions and graphical methods employed. These results are consistent with a single class of noninteracting 125I-insulin binding sites on the solubilized insulin receptor.

Binding Sites↗

[Head shaking test and low-frequency rotation-acceleration test: comparison of sensitivity and specificity].

In studying the vestibular-oculomotor reflex (VOR) in patients with equilibrium disorders, both active and passive methods are used and at different stimulation frequencies, particularly to evaluate vestibular compensation. The present study compares the sensitivity and specificity of the low-frequency pendular test and the HST in normal subjects and in subjects suffering from various vestibular pathologies but showing no signs of spontaneous nystagmus, even under optimal evaluation conditions (infrared videonystagmoscopy). These spontaneous nystagmus-free subjects underwent a routine study including: case history, infrared videonystagmoscopy (HST with 2 Hz stimulation) and ENG recording (saccadic movement, smooth pursuit, OKN, VOR, VOR-fix during pendular stimulation at 0.05 Hz and VOR after Fitzgerald-Hallpike heat stimulation). On the basis of the results the subjects were classified as follows: normal (N): case history free of any equilibrium disorders and all tests negative (122 cases; 8.1%); peripheral vestibular pathology (P): case history of equilibrium disorders and labyrinthine predominance (LP) in excess of 20% upon caloric testing (716 cases; 47.6%); central pathology (C): case history of equilibrium disorders and at least 3 pathological results from among the following tests: saccadic movement, smooth pursuit, OKN, VOR-fix (226 cases; 15.0%); mixed pathology (M): with both signs of P and C (440 cases; 29.3%). The pendular test showed signs of directional dominance (DP) higher than 10% (normal limit) in 7 cases of N (5.7%), 308 P (43.0%), 33 C (14.6%) and 162 M (36.8%). DP was higher than 10% in 55.2% of the P and M cases with onset less than 1 month before, in 42.8% of those with onset within the year and in 37.2% of those with onset more than a year before. A pathological response to the HST was observed (characterized by a series of at least 3 nystagmus shakes after a maximum latency of 15 seconds) in 0 N (0.0%), 378 P (52.8%), 4 C (1.8%) and 247 M (56.1%). The nystagmus seen was nearly always monophasic (92.5%), biphasic nystagmus was only seen in only a few cases (7.5%); moreover it was predominantly horizontal in nature (94.9%) while it was vertical in only a few of the C cases (5.1%). The HST proved pathological in 46.5% of those pathologies with onset less than 1 month before, in 55.8% of those with onset within the year and in 54.4% of those with onset more than a year before. In 213 of the cases presenting pathological response to the HST there was agreement between VOR DP at the pendular test and the direction of the nystagmus evoked by the HST: 138 P (74.6%), 0 C (0.0%), 75 M (68.2%): moreover there was no agreement in 83 cases. When the DP was lower than 10% at the pendular test, the HST proved pathological in 213 P (52.2%), 3 C (1.6%), 150 M (53.9%). Both tests gave negative results in 112 N (91.8%), 163 P (22.8%), 186 C (82.3%), 123 M (27.9%). In cases of peripheral vestibular deficit (P and M) the sensitivity of the rotoacceleration test was 40.7%, specificity 88.5%. The sensitivity of the HST was 54.1%, specificity 98.8%. The sensitivity of the association using both tests was 75.2%, specificity 85.7%.

Biophysical Phenomena↗

[The effect of the composition of food rations on the free amino acid content and the indices of the acid-base balance in the blood during aging].

In experiments on aging rats fed 5 different rations by protein, tryptophan, prevailing base or acid equivalents it was found that tryptophan and plant protein deficiency, high levels of acid equivalents affect serum amino acid composition, aggravates acidosis. Restriction of basic food substances intake in parallel with prevalence of base equivalents made amino acid pool optimal, increased base serum reserves and contributed to longer life-span.

Acid-Base Equilibrium↗

Osmotic properties of boar spermatozoa and their relevance to cryopreservation.

A series of six experiments was conducted to determine the fundamental cryobiological properties of boar spermatozoa to develop optimal approaches for cryopreserving this important cell type. In the first experiments, boar spermatozoa samples were diluted in various osmolalities of experimental solutions (185-900 mOsmol kg-1) to provide hypo-, iso-, and hyperosmotic conditions. Equilibrium cell volumes (Expts 1 and 2) were measured after exposure for 3 min and the change in cell volume was measured over time using an electronic particle counter (Expt 3). The isosmotic cell volume was found to be 26.3 +/- 0.39 microns 3 (mean +/- SEM; n = 5). Over this range of osmolalities, boar spermatozoa behaved as linear osmometers (a linear volume versus 1/osm plot, r2 = 0.99) with an osmotically inactive cell fraction of 67.4 +/- 4.5%. The rate of water permeability (Lp) was determined to be 1.03 +/- 0.05 microns min-1 atm-1, which was consistent within and among donors (P > 0.130). A second series of experiments was performed to determine the effect of temperature and osmolality on boar sperm motility (Expt 4), and the effect of osmolality on the integrity of the sperm plasma membrane and its temperature dependence. Plasma membrane integrity was measured before and after boar spermatozoa were returned to an isosmolality (Expt 6). Motility was not affected at 30 degrees C, relative to that at room temperature, but was significantly decreased (P < 0.05) at 8 degrees C and 0 degree C (yielding a relative reduction to 85% and 35% of original motility, respectively; n = 6). Sperm motility was not significantly decreased (P > 0.05) until the osmolality reached 210 mOsmol kg-1, at which time motility began to decrease from 95% to 10% of the original value at 90 mOsmol kg-1. The integrity of the plasma membrane of boar spermatozoa was found to be dependent on temperature, donor and osmolality, decreasing significantly (P < 0.05) below room temperature, and below 185 mOsmol kg-1 (P < 0.05). There was no significant difference (P > 0.10) in the integrity of the plasma membrane of the samples before and after returning to 290 mOsmol kg-1, indicating that osmotic damage occurs during the initial change from isosmotic to hyposmotic media. These osmotic characteristics could be used to determine optimal conditions for cryopreservation of boar spermatozoa.

Animals↗

Electrolyte profiles within the ileoanal pouch: measurement by an in vivo equilibrium dialysis technique.

BACKGROUND: Patients with an ileoanal pouch have high rates of fluid and electrolyte loss. These improve with pouch adaptation. There is limited information concerning secretion and absorption in the stable ileoanal pouch. A new method to measure and characterize electrolytes in the ileoanal pouch is described. METHODS: Following an in vitro study, nine patients with a stable ileoanal pouch had consecutive placement of dialysis bags consisting of a semi-permeable membrane containing 5 ml of 10% dextran in normal saline into the ileoanal pouch. These were left in place for 15, 30, 60, and 120 min. After determining that 60 min was the optimal timing for measurement of electrolyte concentrations, 12 normal volunteers underwent a similar in vivo dialysis study with dialysis bags withdrawn at 60 min. Sodium, chloride, potassium, phosphorus, calcium and magnesium concentrations in the dialysis bags were compared between the two groups. RESULTS: In the in vitro and in vivo studies, the measured electrolytes reached equilibrium within 60 min. Statistically significant differences between sodium concentrations (160.9 +/- 30.2 vs. 116.8 +/- 13.8 mmol/l, respectively) and phosphorus concentrations (6.8 +/- 5.2 vs. 1.8 +/- 0.7 mg/dl, respectively) at 60 min in ileoanal pouch patients and volunteers were found (p<0.001). There were no statistical differences in the other measured electrolytes between the two groups. CONCLUSION: An in vivo dialysis technique is described for measuring electrolyte concentrations within the ileoanal pouch. Differences in sodium and phosphate concentrations may reflect incomplete adaptation of the ileoanal pouch, and are a potential explanation for increased stool frequency in these patients.

Adult↗

Osmotic characteristics of isolated human and canine pancreatic islets.

Cryopreservation protocols for pancreatic islets may be optimized by modeling the changes in volume that occur during cryoprotectant addition/dilution and cooling/warming. Modeling may be facilitated if it can be demonstrated that isolated islets behave as ideal osmometers where the equilibrium volume is a linear function of the reciprocal of the osmolality of the extracellular medium (the Boyle van't Hoff relationship). The present study was performed to test the hypotheses that: (i) human and canine islets exhibit a linear osmotic response during exposure to hypo-, iso-, and hyperosmotic solutions and to determine the human and canine islets osmotically inactive fraction (Vbp); (ii) to determine if the ionic properties of a concentrated electrolyte solution have an effect on the osmotic response; and (iii) to determine if the osmotic response of pancreatic islets varies between species (human and canine). Islets were isolated from four human and four canine pancreases using standard collagenase digestion and EuroFicoll purification. After 12 h incubation at 37 degrees C, islets (n = 4 from each isolation) were exposed to varying salt or sucrose concentrations (75-1200 mOsm/kg). Islets were held by a micropipette system attached to an inverted microscope, and solutions were exchanged while videotaping. An imaging system was used to determine the cross-sectional area of the islet which was then used to estimate the spherical volume. The results of linear regression analysis indicated that both human and canine islets respond ideally to osmotic conditions within a range of 150-1200 mOsm/kg in both the electrolyte (salt) and the nonelectrolyte (sucrose) solutions. The Vbp value (mean +/- SD) of the four donors was 0.513 +/- 0.068 for canine islets in sucrose solutions, 0.474 +/- 0.068 for canine islets in salt solutions, 0. 425 +/- 0.073 for human islets in sucrose solutions, and 0.39 +/- 0. 073 for human islets in salt solutions. There was a significant difference between human and canine islets (P < 0.05) but not between experimental solutes (NaCl or sucrose) (P < 0.05). These data may be used in conjunction with osmotic tolerance data to develop improved methods for the cryopreservation of isolated islets.

Animals↗

Electrokinetic migration of acidic drugs across a supported liquid membrane.

Electrokinetic cross membrane extraction of acidic drugs was demonstrated for the first time. The acidic drugs were extracted from an alkaline aqueous donor solution (300 microl), through a thin supported liquid membrane of 1-heptanol sustained in the pores of the wall of a porous hollow fiber, and into an aqueous alkaline acceptor solution (30 microl) present inside the lumen of the hollow fiber by the application of a d.c. electrical potential. The negative electrode was placed in the donor solution, and the positive electrode was placed in the acceptor solution. Optimal extractions were accomplished with 1-heptanol as the supported liquid membrane, with 50 V as the driving force, and with pH 12.0 in both the donor and acceptor solutions, respectively (NaOH). Equilibrium extraction conditions were obtained after 5 min of operation with the whole assembly agitated at 1200 rpm. Eleven different acidic drugs were extracted with recovery values between 8 and 100%, and initial data supported that electrokinetic cross membrane extraction provided repeatable data and linear response between original donor concentration and final acceptor concentration of the acidic model compounds.

Acids↗

A theoretical study of the structure of tricarbonatodioxouranate.

The results of a study on the ground states of tricarbonato complexes of dioxouranate using multiconfigurational second-order perturbation theory (CASSCF/CASPT2) are presented. The equilibrium geometries of the complexes corresponding to uranium in the formal oxidation states VI and V, [UO(2)(CO(3))(3)](4)(-) and [UO(2)(CO(3))(3)],(5)(-) have been fully optimized in D(3)(h)() symmetry at second-order perturbation theory (MBPT2) level of theory in the presence of an aqueous environment modeled by a reaction field Hamiltonian with a spherical cavity. The uranyl fragment has also been optimized at CASSCF/CASPT2, to obtain an estimate of the MBPT2 error. Finally, the effect of distorting the D(3)(h)() symmetry to C(3) has been investigated. This study shows that only minor geometrical rearrangements occur in the one-electron reduction of [UO(2)(CO(3))(3)](4)(-) to [UO(2)(CO(3))(3)],(5)(-) confirming the reversibility of this reduction.

Journal Article↗

Miniature device for aqueous and non-aqueous solubility measurements during drug discovery.

PURPOSE: A miniature device was developed for the measurement of aqueous and non-aqueous equilibrium solubility during drug discovery. The solubility values obtained using the miniature device were compared to those obtained using the conventional shake-flask method. METHODS: The aqueous solubility of six structurally diverse compounds, the solubility of carbamazepine in various cosolvent systems, and the pH-solubility profile of saquinavir were determined using the miniature device. The device contains a multichannel cartridge pump and a Tygon tubing that is mounted on the pump with two ends linked by a syringe filter. The drug slurry was filled into the tubing and circulated inside, continually passing through the syringe filter. At the end of the experiment, the filtrate was collected and analyzed directly by High-Pressure Liquid Chromatography (HPLC). The solubility was also determined by the shake-flask method. RESULTS: The solubility values determined by the miniature device were in good agreement with those measured by the conventional shake-flask method. CONCLUSIONS: The miniature device provides a unique way of testing aqueous and non-aqueous equilibrium solubility in a microscale setting. With approximately 1 mg of compound, it is possible to determine the entire pH-solubility profile. The device is useful for solubility screening during lead optimization and candidate selection in early drug discovery, when compound supply is limited. It can also be used for screening solubility in non-aqueous systems to select vehicles for preclinical in vivo studies.

Anti-HIV Agents↗

Some properties of acid and alkaline phosphates from boar sperm plasma membranes.

Boar sperm plasma membranes were purified by differential and sucrose density equilibrium centrifugation and were found to yield a single band at a density of 1.14 g/cm3. Both alkaline and acid phosphatase activities were enriched in this fraction. The alkaline phosphatase activity was optimal in 100 mM tris (hydroxymethyl) methylamine (Tris)-NaHCO3 at pH 9.9 with 0.05% Triton X-100 and 1 mM MgCl2. This activity was inhibited by ethylenediaminetetraacetic acid (EDTA), cadmium, zinc or heating at 60 degrees C for 30 min. Also, L-homoarginine caused approximately 70% inhibition and L-phenylalanine or L-leucine caused about 10 to 20% inhibition. Acid phosphatase activity was optimal in 100 mM sodium acetate at pH 5.1 with 0.05% Triton. Sodium dodecyl sulfate, potassium fluoride (KF) or sulfhydryl reagents inhibited the activity, while EDTA or heating at 60 degrees C had no effect. These data for enzymes from boar sperm plasma membranes can be used for future work on the quantitation of the enzymes, distinguishing these two phosphatases from other phosphohydrolases, purification of the enzymes and for comparison to phosphatases in other tissues.

Acid Phosphatase↗

Universality of the off-equilibrium response function in the kinetic Ising chain.

The off-equilibrium response function chi(t,t(w)) and autocorrelation function C(t,t(w)) of an Ising chain with spin-exchange dynamics are studied numerically and compared with the same quantities in the case of spin-flip dynamics. It is found that, even though these quantities are different in the two cases, the parametric plot of chi(t,t(w)) versus C(t,t(w)) is the same. While this result could be expected in higher dimensionality, where chi(C) is related to the equilibrium state, it is far from trivial in the one-dimensional case where this relation does not hold. The origin of the universality of chi(C) is traced back to the optimization of domains position with respect to the perturbing external field. This mechanism is investigated resorting to models with a single domain moving in a random environment.

Journal Article↗

Multilane simulations of traffic phases.

The optimal velocity model, as modified by the author, is used in simulations of traffic on a dual-lane highway and a single-lane highway with an on-ramp. The equilibrium solutions of the modified model cover a two-dimensional region of flow-density space beneath the fundamental-diagram curve, rather than just lying on the curve as in the original model. Thus it satisfies a requirement of the three-phase model of Kerner [Phys. Rev. Lett. 81, 3797 (2002)]. Synchronization of velocity across dual lanes due to frequent lane changes is observed in free flow. True synchronized flow, as determined by the region of density-flow space it occupies, is obtained in on-ramp simulations with typical driver reaction times. A gradual change to the formation of a jam is observed for increasing delay times.

Journal Article↗

The influence of water on protease-catalyzed peptide synthesis in acetonitrile/water mixtures.

Protease-catalyzed peptide synthesis in acetonitrile/water mixtures, containing 0-90% water, was investigated. alpha-Chymotrypsin, as well as thermolysin, were deposited on solid supports, prior to exposure to the reaction media. Peptide syntheses were performed using both a kinetically controlled process (chymotrypsin) and an equilibrium-controlled synthesis (thermolysin). The activity of chymotrypsin decreased at low water contents. However, at low water contents (1-10%) hydrolytic side reactions were suppressed and high yields of dipeptides were obtained. Optimal water content for the thermolysin-catalyzed reaction was 4-8%. The dipeptides produced were fully soluble in the reaction mixtures. High operational stability for alpha-chymotrypsin was obtained during 216 h of reaction.

Acetonitriles↗

Dependence of the Calvin cycle activity on kinetic parameters for the interaction of non-equilibrium cycle enzymes with their substrates.

Kinetic model studies and control analyses of the Calvin photosynthesis cycle have been performed to characterize the dependence of the cycle activity on maximum velocities and Km values for the interaction of the non-equilibrium cycle enzymes and ATP synthetase with their substrates under conditions of light and carbon dioxide saturation. The results show that Km values have no major influence on the cycle activity at optimal concentrations of external orthophosphate. The maximum cycle activity is controlled mainly by the catalytic capacities of ATP synthetase and sedoheptulose-bisphosphatase, and is close to the maximum cycle flux that can be supported by these two enzymes.

Chloroplasts↗

5-Fluorouracil derivatives with serum protein binding potencies.

To develop an optimal delivery system for 5-fluorouracil (5FU) using serum protein as a drug carrier, a series of its benzyl derivatives was synthesized. Then their binding to the serum protein was investigated by equilibrium dialysis. The benzyl derivatives of 5FU were strongly bound to rat plasma protein or human serum albumin. The bound percentage increased with increasing hydrophobicity. It was suggested that the benzyl derivative of 5FU existed in the blood as a complex with serum albumin and circulated for a long time as a polymeric drug does.

Animals↗

Biomechanical analysis of movement strategies in human forward trunk bending. II. Experimental study.

The large mass of the human upper trunk, its elevated position during erect stance, and the small area limited by the size of the feet, stress the importance of equilibrium control during trunk movements. The objective of the present study was to perform a biomechanical analysis of fast forward trunk movements in order to understand the coordination between movement and posture. The analysis is based on a comparison between experimentally observed bending and hypothetical "optimal bending" performed on an infinitely narrow support, as presented in a companion paper. The experimental data were obtained from 16 subjects who performed fast forward bending while standing on a wide platform or on a narrow beam. The analysis is performed by decomposition of the movement into three dynamically independent components, each representing a movement along one of the three eigenvectors of the motion equation. The eigenmovements are termed "hip", "ankle", and "knee" eigenmovements, according to the dominant joint. The experimentally observed movement is characterized mainly by the hip and ankle eigenmovements, whereas the knee eigenmovement is negligible. Similarly to the "optimal bending" the ankle eigenmovement starts earlier and lasts longer than the hip eigenmovement. An early forward acceleration of the center of gravity in the ankle eigenmovement is caused by anticipatory changes in the ankle joint torque. This clarifies the role of the early tibialis anterior burst and/or soleus inhibition usually observed in electromyographic recordings during forward bending. The results suggest that the hip and the ankle eigenmovements can be treated as independently controlled motion units aimed at functionally different behavioral goals: the bending per se and postural adjustment. It is proposed that the central nervous system has to control these motion units sequentially in order to perform the movement and maintain equilibrium. It is also suggested that the hip and ankle eigenmovements can be regarded as a biomechanical background for the hip and ankle strategies introduced by Horak and Nashner (1986) on the basis of electromyographic recordings and kinematic patterns in response to postural perturbations.

Adult↗

Ropivacaine 7.5 mg/ml for elective caesarean section. A clinical and pharmacokinetic comparison of 150 mg and 187.5 mg.

BACKGROUND: The new, long-acting local anaesthetic ropivacaine has shown less systemic toxicity than bupivacaine and a concentration of 7.5 mg/ml can therefore be used for epidural anaesthesia in Caesarean section. The present pilot study was undertaken to find indications for an optimal dosage by comparing the clinical effects, quality of anaesthesia and pharmacokinetics of ropivacaine 150 mg (lower dose = LD) vs 187.5 mg (higher dose = HD) for women undergoing elective Caesarean section under epidural anaesthesia. METHODS: Sixteen full-term women scheduled for elective Caesarean section in two equal-sized consecutive groups received 20 or 25 ml ropivacaine epidurally in this non-randomised, open study. Study parameters included sensory and motor blockade, circulatory response, intraoperative pain and discomfort, neonatal evaluation and pharmacokinetic determinations. RESULTS: Block height varied between T5 and T2 in the LD group, whereas the HD group produced 4 unnecessarily high blocks (C8 in 3 women and C7 in 1 woman). Surgical anaesthesia was excellent in both groups. Circulatory stability was pronounced in the LD group (no ephedrine given), while 4 women required ephedrine in the HD group. Neonatal outcome as judged by Apgar scores; umbilical blood gas determinations and NACS scores were excellent in both groups. The plasma concentration-time profiles indicated linearity in the concentration range studied, with similar clearance values to those reported previously. Placental drug equilibrium was rapid; however, the foetal drug exposure depended on intrauterine exposure time. CONCLUSIONS: 20-25 ml ropivacaine 7.5 mg/ml produced very satisfactory conditions for elective Caesarean section under epidural anaesthesia. In this small population, 150 mg ropivacaine seemed optimal, while 187.5 mg produced unnecessarily extended block height in 50% of the women.

Adult↗

Effect of Herbivory and Plant Species Replacement on Primary Production.

Grazing optimization occurs when herbivory increases primary production at low grazing intensities. In the case of simple plant-herbivore interactions, such an effect can result from recycling of a limiting nutrient. However, in more complex cases, herbivory can also lead to species replacement in plant communities, which in turn alters how primary production is affected by herbivory. Here we explore this issue using a model of a limiting nutrient cycle in an ecosystem with two plant species. We show that two major plant traits determine primary production at equilibrium: plant recycling efficiency (i.e., the fraction of the plant nutrient stock that stays within the ecosystem until it is returned to the nutrient pool in mineral form) and plant ability to deplete the soil mineral nutrient pool through consumption of this resource. In cases where sufficient time has occurred, grazing optimization requires that herbivory improve nutrient conservation in the system sufficiently. This condition sets a minimum threshold for herbivore nutrient recycling efficiency, the fraction of nutrient consumed by herbivores that is recycled within the ecosystem to the mineral nutrient pool. This threshold changes with plant community composition and herbivore preference and is, therefore, strongly affected by plant species replacement. The quantitative effects of these processes on grazing optimization are determined by both the recycling efficiencies and depletion abilities of the plant species. However, grazing optimization remains qualitatively possible even with plant species replacement.

boreal forests↗