A simplified procedure to determine the optimal rate of freezing biological systems

The effect of several cell-level parameters on the predicted optimal cooling rate B(opt) of an arbitrary biological system has been studied using a well-defined water transport model. An extensive investigation of the water transport model revealed three key cell level parameters: reference permeability of the membrane to water L(pg), apparent activation energy E(Lp), and the ratio of the available surface area for water transport to the initial volume of intracellular water (SA/WV). We defined B(opt) as the "highest" cooling rate at which a predefined percent of the initial water volume is trapped inside the cell (values ranging from 5% to 80%) at a predefined end temperature (values ranging from -5 degrees C to -40 degrees C). Irrespective of the choice of the percent of initial water volume trapped and the end temperature, an exact and linear relationship exists between L(pg), SA/WV, and B(opt0. However, a nonlinear and inverse relationship is found between E(Lp) and B(opt). Remarkably, for a variety of biological systems a comparison of the published experimentally determined values of B(opt) agreed quite closely with numerically predicted B(opt) values when the model assumed 5% of initial water is trapped inside the cell at a temperature of -15 degrees C. This close agreement between the experimental and model predicted optimal cooling rates is used to develop a generic optimal cooling rate chart and a generic optimal cooling rate equation that greatly simplifies the prediction of the optimal rate of freezing of biological systems.     

Reciprocal role of the inward currents ib, Na and i(f) in controlling and stabilizing pacemaker frequency of rabbit sino-atrial node cells.

Experiments and computations were done to clarify the role of the various inward currents in generating and modulating pacemaker frequency. Ionic currents in rabbit single isolated sino-atrial (SA) node cells were measured using the nystatin-permeabilized patch-clamp technique. The results were used to refine the Noble-DiFrancesco-Denyer model of spontaneous pacemaker activity of the SA node. This model was then used to show that the pacemaker frequency is relatively insensitive to the magnitude of the sodium-dependent inward background current ib, Na. This is because reducing ib, Na hyperpolarizes the cell and so activates more hyperpolarizing-activated current, i(f), whereas the converse occurs when ib, Na is increased. The result is that i(f) and ib, Na replace one another and so stabilize nodal pacemaker frequency.     

High resolution electrocardiography in healthy dogs: time domain parameters and comparison of the non-stationary (Wigner distribution) versus standard stationary frequency domain analysis methods

Fractionated heart activation can be detected as late potentials from surface recordings of signal-averaged electrocardiograms (SA ECG) which are considered as a marker of sustained ventricular tachycardia. For animal studies, reference values in time and frequency domain analyses are essentially missing. In the present study, we have established reference values in SA ECG time domain analysis and time-frequency representation of heart activation in healthy dogs. A group of 25 healthy mongrel dogs (body weight 12-15 kg) was investigated. Wigner distribution and our modification of Fast Fourier transform (FFT), gliding window FFT, was applied in SA ECG frequency domain analysis. Reference values in time domain SA ECG were established. Time and voltage criteria were adapted to short duration of heart cycle and fast voltage decrement of the QRS complex in dogs. Wigner distribution and gliding window FFT were applied in order to describe mean heart activation in the frequency domain. Contribution of higher frequencies (30-80 Hz) was detected by both frequency analysis methods in the second third of ventricular activation in healthy animals. Presented results could offer a basis for further experimental arrhythmologic studies.     

A patient risk model to determine the optimal output constancy check frequency for a radiotherapy machine

PurposeThe output constancy check, a basic quality control (QC) item for radiotherapy machines, is performed daily according to suggestions in technical reports by experienced experts. In this study, a patient risk model was built to determine the optimal frequency of an output constancy check for a specific radiotherapy machine.Methods and materialsThe method was based on the patient risk model and comprised three steps: 1) the power function graph was used to select a proper QC rule and the average number of QC measurements per QC rule evaluation. 2) The optimal QC frequency was determined by the minimum integer value of expected patients treated between QC measurements. 3) The individual control chart (I-Chart) was used to evaluate the effectiveness of the model. The model was implemented on the output constancy check of a Tomotherapy machine.ResultsThe QC rule with the limits set to the mean ± 3 standard deviations and 5 measurements per QC were selected according to the power function graph. The optimal frequency was observed every 21 patients. The I-Chart showed that the optimal frequency detected the machine failure earlier compared to the conventional daily frequency. The model could monitor whether Tomotherapy machine was in good condition and predicted the time to adjust the machine.ConclusionsThe optimal output constancy check frequency of a radiotherapy machine is determined by the number of patients, which uses patient risk model. The optimal frequency is superior to the conventional daily frequency in identifying machine failure earlier.     

A simulation study of the dynamics of a driven filament in an Aristotelian fluid

We describe a method, based on techniques used in molecular dynamics, for simulating the inertialess dynamics of an elastic filament immersed in a fluid. The model is used to study the "one-armed swimmer". That is, a flexible appendage externally perturbed at one extremity. For small-amplitude motion our simulations confirm theoretical predictions that, for a filament of given length and stiffness, there is a driving frequency that is optimal for both speed and efficiency. However, we find that to calculate absolute values of the swimming speed we need to slightly modify existing theoretical approaches. For the more relevant case of large-amplitude motion we find that while the basic picture remains the same, the dependence of the swimming speed on both frequency and amplitude is substantially modified. For large-amplitudes we show that the one-armed swimmer is comparatively neither inefficient nor slow. This begs the question, why are there little or no one-armed swimmers in nature?     

Influence of milking frequency on mammary gland dynamics

The effects of milking frequency on milk production is a key question for the dairy industry. Milk production is related to the number of active alveoli in the mammary gland and movement between active and quiescent alveolar pools is influenced by the milking frequency. In this paper, we analyse a mechanistic model based on known biological inputs that describes the effect of milking frequency on the alveolar composition of the mammary gland. It is shown that the model can qualitatively reproduce the correct alveolar dynamics. We also investigate the model robustness and parameter sensitivity. Additionally, by making the plausible assumption that the senescence rate of alveoli is proportional to the number of quiescent alveoli present, we obtain an analytical solution requiring periodic resetting.     

Participation of the Breuer-Hering inflation reflex in regulation of respiration frequency in anaesthetized rats.

The respiration frequency and Breuer-Hering inflation reflex (BHIR) values were compared in anaesthetized rats after unilateral vagotomy. Nos significant differences were found in the decrease in respiration frequency after transection of the right of left vagus and there were no differences in BHIR values in correlation to conduction by the right or left vagus. With unilateral vagotomy and an inflation pressure of 20 cm H20 the BHIR values fell to about one fourth of the control values. Progressive cooling of the contralateral vagus led at 8-10 degrees C to a non-significant decrease in respiration frequency to 87% of the initial value; in this state the BHIR could not be elicited. Further cooling of the vagus to 0 degrees C reduced respiration frequency to the same values as in bilateral vagotomy. The discrepancy between the mild decrease in respiration frequency and simultaneous disappearance of the BHIR shows that changes in respiration frequency in anaesthetized rats are not directly correlated to the presence of the BHIR.     

Increased micronucleus frequency in exfoliated cells of the buccal mucosa in hairdressers

Hairdressers are exposed daily to chemical substances, such as dyes, chemical straighteners and curling chemicals, which can be absorbed, inhaled or possibly ingested. We analyzed the frequency of micronuclei (MNC) in exfoliated cells of the buccal mucosa of 50 hairdressers and 50 controls in Pelotas, RS, Brazil. An assessment was carried out on the incidence of MNC, binucleated cells (BNC), broken egg cells (BEC), budding cells (BC), and the sum of anomalies (SA), in 2000 cells per individual. The data were analyzed with SPSS, using the Mann-Whitney U-test, α = 0.05. The mean number of anomalies in hairdressers was 2.02 ± 3.60 MNC; 8.50 ± 5.07 BNC; 9.06 ± 3.83 BEC; 0.32 ± 0.62 BC, and 19.90 ± 9.61 SA; in controls it was 0.36 ± 1.06 MNC; 5.20 ± 4.73 BNC; 5.92 ± 2.67 BEC; 0.10 ± 0.36 BC, and 11.58 ± 6.67 SA; the differences for all parameters were significant. The non-occupational factors did not significantly influence the alterations. A significant increase of BEC (P = 0.003) was observed in the hairdressers and SA (P = 0.033) in females. The lowest income level influenced MNC (P = 0.044), and the habit of not smoking influenced SA (P = 0.020). We concluded that exposure to substances used by hairdressers is genotoxic for men.     

Unrestrained stochastic dynamics simulations of the UUCG tetraloop using an implicit solvation model.

Three unrestrained stochastic dynamics simulations have been carried out on the RNA hairpin GGAC[UUCG] GUCC, using the AMBER94 force field (Cornell et al., 1995. J. Am. Chem. Soc. 117:5179-5197) in MacroModel 5.5 (Mohamadi et al., 1990. J. Comp. Chem. 11:440-467) and either the GB/SA continuum solvation model (Still et al., 1990. J. Am. Chem. Soc. 112:6127-6129) or a linear distance-dependent dielectric (1/R) treatment. The linear distance-dependent treatment results in severe distortion of the nucleic acid structure, restriction of all hydroxyl dihedrals, and collapse of the counterion atmosphere over the course of a 5-ns simulation. An additional vacuum simulation without counterions shows somewhat improved behavior. In contrast, the two GB/SA simulations (1.149 and 3.060 ns in length) give average structures within 1.2 A of the initial NMR structure and in excellent agreement with results of an earlier explicit solvent simulation (Miller and Kollman, 1997. J. Mol. Biol. 270:436-450). In a 3-ns GB/SA simulation starting with the incorrect UUCG tetraloop structure (Cheong et al., 1990. Nature. 346:680-682), this loop conformation converts to the correct loop geometry (Allain and Varani, 1995. J. Mol. Biol. 250:333-353), suggesting enhanced sampling relative to the previous explicit solvent simulation. Thermodynamic effects of 2'-deoxyribose substitutions of loop nucleotides were experimentally determined and are found to correlate with the fraction of time the ribose 2'-OH is hydrogen bonded and the distribution of the hydroxyl dihedral is observed in the GB/SA simulations. The GB/SA simulations thus appear to faithfully represent structural features of the RNA without the computational expense of explicit solvent.     

Spatiotemporal frequency responses of cat retinal ganglion cells

Spatiotemporal frequency responses were measured at different levels of light adaptation for cat X and Y retinal ganglion cells. Stationary sinusoidal luminance gratings whose contrast was modulated sinusoidally in time or drifting gratings were used as stimuli. Under photopic illumination, when the spatial frequency was held constant at or above its optimum value, an X cell's responsivity was essentially constant as the temporal frequency was changed from 1.5 to 30 Hz. At lower temporal frequencies, responsivity rolled off gradually, and at higher ones it rolled off rapidly. In contrast, when the spatial frequency was held constant at a low value, an X cell's responsivity increased continuously with temporal frequency from a very low value at 0.1 Hz to substantial values at temporal frequencies higher than 30 Hz, from which responsivity rolled off again. Thus, 0 cycles X deg-1 became the optimal spatial frequency above 30 Hz. For Y cells under photopic illumination, the spatiotemporal interaction was even more complex. When the spatial frequency was held constant at or above its optimal value, the temporal frequency range over which responsivity was constant was shorter than that of X cells. At lower spatial frequencies, this range was not appreciably different. As for X cells, 0 cycles X deg-1 was the optimal spatial frequency above 30 Hz. Temporal resolution (defined as the high temporal frequency at which responsivity had fallen to 10 impulses X s-1) for a uniform field was approximately 95 Hz for X cells and approximately 120 Hz for Y cells under photopic illumination. Temporal resolution was lower at lower adaptation levels. The results were interpreted in terms of a Gaussian center-surround model. For X cells, the surround and center strengths were nearly equal at low and moderate temporal frequencies, but the surround strength exceeded the center strength above 30 Hz. Thus, the response to a spatially uniform stimulus at high temporal frequencies was dominated by the surround. In addition, at temporal frequencies above 30 Hz, the center radius increased.     

The frequency selectivity of information-processing channels in the tactile sensory system

The frequency selectivity of the P, NP I, and NP II channels of the four-channel model of mechanoreception for glabrous skin was measured psychophysically by an adaptation tuning curve procedure. The results substantially extend the frequency range over which the frequency selectivity of these channels is known and further confirm the hypothesis that the input stage of each of these channels consists of specific sensory nerve fibers and associated receptors. Specifically, the frequency characteristics of Pacinian nerve fibers, rapidly adapting (RA) nerve fibers, and slowly adapting Type II (SA II) nerve fibers were found to be the peripheral neurophysiological correlates of the P, NP I, and NP II channels, respectively. The finding that the tuning characteristic for a test stimulus of 250 Hz delivered through a small (0.008 cm2) contactor depended dramatically on the duration of the test stimulus whereas the detection threshold did not, provides new evidence in support of the hypothesis that separate NP II and P channels exist.     

The effect of acclimation on mating frequency and mating competitiveness in the Queensland fruit fly, Dacus tryoni, in optimal and cool mating regimes

Mating frequency in groups of Dacus tryoni which had been either warm-acclimated or cold-acclimated were compared in temperature regimes ranging from just above mating-threshold to optimal. Cold-acclimation appeared to suppress initial mating ability of mature insects of both sexes to an extent which depended upon the acclimation regime used. The most favourable cold-acclimation regime produced flies which in certain circumstances were able to mate at an initial frequency similar to that of warm-acclimated flies. In no mating regime was initial mating significantly more frequent in any cold-acclimated group than it was in any warm-acclimated group. In most cases warm-acclimated flies in a given regime mated at high frequency for 1–2 days, whereas the cold-acclimated flies mated at low frequency for a greater number of days. In all cases, cold-acclimated flies accumulated a similar or significantly lower total number of matings than warm-acclimated groups. In experiments in which both warm-acclimated and cold-acclimated males competed for cold-acclimated females, the warm-acclimated males always out-competed the cold-acclimated males in two mild (near optimal) regimes. In a relatively harsh (near torpor threshold) regime, there was no significant difference in the competitive abilities of cold-acclimated and warm-acclimated males. The relevance of these results to possible acclimation procedures used in control campaigns involving release of sterile males is discussed.     

Temperature dependence of the low frequency dynamics of myoglobin. Measurement of the vibrational frequency distribution by inelastic neutron scattering.

Inelastic neutron scattering spectra of myoglobin hydrated to 0.33 g water (D2O)/g protein have been measured in the low frequency range (1-150 cm-1) at various temperatures between 100 and 350 K. The spectra at low temperatures show a well-resolved maximum in the incoherent dynamic structure factor Sinc(q, omega) at approximately 25 cm-1 and no elastic broadening. This maximum becomes gradually less distinct above 180 K due to the increasing amplitude of quasielastic scattering which extends out to 30 cm-1. The vibrational frequency distribution derived independently at 100 and 180 K are very similar, suggesting harmonic behavior at these temperatures. This result has been used to separate the vibrational motion from the quasielastic motion at temperatures above 180 K. The form of the density of states of myoglobin is discussed in relation to that of other amorphous systems, to theoretical calculations of low frequency modes in proteins, and to previous observations by electron-spin relaxation of fractal-like spectral properties of proteins. The onset of quasielastic scattering above 180 K is indicative of a dynamic transition of the system and correlates with an anomalous increase in the atomic mean-squared displacements observed by M?ssbauer spectroscopy (Parak, F., E. W. Knapp, and D. Kucheida. 1982. J. Mol. Biol. 161: 177-194.) and inelastic neutron scattering (Doster, W., S. Cusack, and W. Petry, 1989. Nature [Lond.]. 337: 754-756.) Similar behavior is observed for a hydrated powder of lysozyme suggesting that the low frequency dynamics of globular proteins have common features.     

A general program to estimate gene-frequencies by the method of maximum-likelihood.

The program estimates the optimal gene-frequencies from observed values of phenotypes by the method of maximum-likelihood. The program does not make assumptions about a specific blood-group-system. This information has to be supplied by input cards. A straightforward iteration process is used for obtaining initial estimates for the gene-frequencies in question. Then an iterative application of the Newton-Raphson method is used to solve the maximum-likelihood equations. The program yields the gene-frequencies and their standard deviations, expected values of the phenotypes, expected values of the genotypes. Finally the chi-square between observed frequencies and (optimal) expected values of phenotypes is computed.     

Seven-helix bundles: molecular modeling via restrained molecular dynamics.

Simulated annealing via restrained molecular dynamics (SA/MD) has been used to model compact bundles of seven approximately (anti)parallel alpha-helices. Seven such helix bundles occur, e.g., in bacteriorhodopsin, in rhodopsin, and in the channel-forming N-terminal domain of Bacillus thuringiensis delta-endotoxin. Two classes of model are considered: (a) those consisting of seven Ala20 peptide chains; and (b) those containing a single polypeptide chain, made up of seven Ala20 helices linked by GlyN interhelix loops (where N = 5 or 10). Three different starting C alpha templates for SA/MD are used, in which the seven helices are arranged (a) on a left-handed circular template, (b) on a bacteriorhodopsin-like template, or (c) on a zig-zag template. The ensembles of models generated by SA/MD are analyzed in terms of their geometry and energetics, and the most stable structures from each ensemble are examined in greater detail. Structures resembling bacteriorhodopsin and structures resembling delta-endotoxin are both represented among the most stable structures. delta-Endotoxin-like structures arise from both circular and bacteriorhodopsin-like C alpha templates. A third helix-packing mode occurs several times among the stable structures, regardless of the C alpha template and of the presence or absence of interhelix loops. It is characterized by a "4 + 1" core, in which four helices form a distorted left-handed supercoil around a central, buried helix. The remaining two helices pack onto the outside of the core. This packing mode is comparable with that proposed for rhodopsin on the basis of two-dimensional electron crystallographic and sequence analysis studies.     

The frequency selectivity of information-processing channels in the tactile sensory system

The frequency selectivity of the P, NP I, and NP II channels of the four-channel model of mechanoreception for glabrous skin was measured psychophysically by an adaptation tuning curve procedure. The results substantially extend the frequency range over which the frequency selectivity of these channels is known and further confirm the hypothesis that the input stage of each of these channels consists of specific sensory nerve fibers and associated receptors. Specifically, the frequency characteristics of Pacinian nerve fibers, rapidly adapting (RA) nerve fibers, and slowly adapting Type II (SA II) nerve fibers were found to be the peripheral neurophysiological correlates of the P, NP I, and NP II channels, respectively. The finding that the tuning characteristic for a test stimulus of 250 Hz delivered through a small (0.008 cm²) contactor depended dramatically on the duration of the test stimulus whereas the detection threshold did not, provides new evidence in support of the hypothesis that separate NP II and P channels exist.     

Low frequency dynamics of chymotrypsin by neutron spectroscopy

The low frequency vibrational modes of enzymes have large amplitudes of vibration which should be related to conformational changes that occur during enzyme action. In the present paper we present inelastic neutron scattering measurements for -chymotrypsin that show a peak in the low frequency spectrum. This peak is well defined at 77 K. Gaussian fits yield values of 0.93±0.05, 0.86±0.04, 0.81±0.05, and 0.87±0.06 THz for the peak position at wave vector transfers (Q) of 1.00, 1.40, 1.85, and 3.00 Å^-1, respectively. The full widths at half maximum are all greater than the resolution (0.2 THz) by at least a factor of two. At 298 K a weak peak at about 0.6 THz was observed for Q values of 1.0, 1.4 and 1.85 Å^-1. The data are interpreted in terms of the allowed oscillations of a large globular protein treated as an elastic sphere. Assuming a Raman active oscillation at 0.9 THz it is shown that a peak in the neutron scattering response at 0.6 THz may arise from a rotational shear mode of the chymotrypsin molecule.     

Relationship between the tension-time area and the frequency of stimulation in motor units of the rat medial gastrocnemius muscle.

The tension-time area is an estimation of the work performed by contracting motor units. The relationship between tension and frequency of stimulation and between tension-time area and frequency have been studied on 148 single motor units of the rat medial gastrocnemius muscle, under isometric conditions. Motor units were classified as fast fatigable (FF), fast resistant to fatigue (FR) or slow (S). Trains of stimuli of increasing frequency and constant duration were used. For all motor units a half of the maximum tetanic tension corresponded to lower frequencies compared to frequencies at a half of the maximum tension-time area. Moreover, the slopes of tension-frequency and area-frequency curves (change of tension or area per 1 Hz rise in frequency) were higher for slow than for fast motor units. The tension-time area per one pulse was calculated for different frequencies of stimulation. For slow units the maximum area per pulse corresponded to significantly lower frequencies than for fast ones, especially of FF type. However, for all three types of motor units this optimal frequency corresponded to sub-fused tetani with a tension of about 75% of the maximum tension, and with the fusion index slightly over 0.90. The absolute values of the maximum tension-time area per pulse revealed that in one contraction within the tetanus, slow units are generating greater work than FR units. The work performed by FF units is nearly two times larger than for S units, although the tension of slow units is over eight times lower. The presented results reveal that the contraction of slow motor units is much more effective than was suggested based on their low tension.     

Estimation of gene frequency and heterozygosity from pooled samples

Pooling of DNA samples can significantly reduce the effort of population studies with DNA markers. I present a statistical model and numerical method for estimating gene frequency when pooled DNA is assayed for the presence/absence of alleles. Analytical and Monte‐Carlo methods examined estimation variance and bias, and hence optimal pool size, under a triangular allele frequency distribution. For gene frequency of rarer alleles, the optimal number of pooled individuals is approximately the inverse of the gene frequency. For heterozygosity, the optimal pool is approximately half the allele number; this results in pools containing, on average, 60% of possible alleles.     

Optimization of batch and fed-batch bioreactors using simulated annealing

In order for biobased industrial products to compete economically with petroleum-derived products, significant reduction in their processing cost is necessary. Since most bioprocesses are operated in batch or fed-batch mode, their optimization involves theoretical and computational challenges. Simulated annealing (SA), a stochastic optimization algorithm, is used in this study to solve a number of challenging optimization problems related to the design and operation of bioreactors. Two well-known case studies are considered in which the robustness and efficiency of the SA algorithm is demonstrated. More specifically, in the first case study it is shown that the global optimal solution located by SA achieves significant improved productivity when compared with the results of previous investigations. In the second case study a realistic objective function is considered where the economic performance of a bioprocess is optimized. SA exhibits impeccable performance and robustness and was able to locate the global optimal solution irrespective of the initial point selected.