Biomedical subjects

U-P Hansen

Publications and source records attributed to U-P Hansen.

7 recordsLinked to original sources

PubMed2006

Strengths and limits of Beta distributions as a means of reconstructing the true single-channel current in patch clamp time series with fast gating.

Single-channel current seems to be one of the most obvious characteristics of ion transport. But in some cases, its determination is more complex than anticipated at first glance. Problems arise from fast gating in time series of patch-clamp current, which can lead to a reduced apparent (measured) single-channel current. Reduction is caused by undetected averaging over closed and open intervals in the anti-aliasing filter. Here it is shown that fitting the measured amplitude histograms by Beta distributions is an efficient tool of reconstructing the true current level from measured data. This approach becomes even more powerful when it is applied to amplitude distributions-per-level. Simulated time series are employed to show that the error sum is a good guideline for finding the correct current level. Furthermore, they show that a Markov model smaller than the one used for gating analysis can be used for current determination (mostly O-C, i.e., open-closed). This increases the reliability of the Beta fit. The knowledge of the true current level is not only important for the understanding of the biophysical properties of the channel. It is also a prerequisite for the correct determination of the rate constants of gating. The approach is applied to measured data. The examples reveal the limits of the analysis imposed by the signal-to-noise ratio and the shape of the amplitude distribution. One application shows that the negative slope of the I-V curve of the human MaxiK channel expressed in HEK293 cells is caused by fast gating.

Algorithms↗

PubMed2005

Effect of CO2 supply on formation of reactive oxygen species in Arabidopsis thaliana.

Light-induced generation of reactive oxygen species (ROS) in 2-week-old leaves of Arabidopsis thaliana was studied by means of the ROS-sensitive dyes nitroblue tetrazolium (NBT) and 5-(and-6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (DCF-DA). Superposition of pictures of chlorophyll fluorescence and DCF fluorescence indicated that the origin of ROS was in the chloroplasts. Experiments were done with zero, 0.1, or 10 mM NaHCO3 in the infiltration medium. Energy quenching in photosystem II was higher under low CO2 concentrations as measured by chlorophyll fluorescence. DCF fluorescence showed that CO2 deficiency led to an increase of ROS generation. In contrast, the photosystem II inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea reduced the light-induced increase of DCF fluorescence. This indicates that ROS production does not primarily result from over-reduction of photosystem II as caused by impeding electron flow in the electron transfer chain. More likely, it is an effect of diverting electron flux normally aimed at carboxylation in the Calvin cycle to other sinks more prone to the generation of toxic radicals. There was no significant effect of salicyl hydroxamate (a blocker of the alternative oxidase), showing that the mitochondrial electron transfer chain seems to play a minor role as already indicated by the superposition of chlorophyll and DCF fluorescence.

Arabidopsis↗

PubMed2005

Light-induced decrease in DCF fluorescence of wheat leaves in the presence of salicyl hydroxamate.

5-(and-6)-Carboxy-2',7'-dichlorodihydrofluorescein diacetate (DCF-DA), a permeative indicator of oxidative stress, was loaded into dissected leaves of wheat in order to monitor the temporal development of reactive oxygen species. DCF fluorescence was found to be constant under dark conditions. Upon loading the leaves with salicyl hydroxamate, a blocker of the alternative oxidase, DCF fluorescence linearly increased in the dark. This indicates a function of alternative oxidase in preventing reactive oxygen radicals in the mitochondria. Upon illumination, the DCF signal decreased within 5 min. As illuminated chloroplasts would increase the load of reactive oxygen species, the observed decrease cannot be assigned to the production of reactive oxygen species in the chloroplasts. Three different putative mechanisms are considered which all assign an important role to light-induced delivery of NAD(P)H: (1) direct quenching of DCF fluorescence by light-generated NAD(P)H, (2) light-stimulated activation of scavenging enzymes, or (3) redirection of mitochondrial electron fluxes as caused by the delivery of excess redox equivalents (NADH) from the chloroplasts.

Chloroplasts↗

PubMed2003

A reduced model of the fluorescence from the cyanobacterial photosynthetic apparatus designed for the in situ detection of cyanobacteria.

Fluorometric determination of the chlorophyll (Chl) content of cyanobacteria is impeded by the unique structure of their photosynthetic apparatus, i.e., the phycobilisomes (PBSs) in the light-harvesting antennae. The problems are caused by the variations in the ratio of the pigment PC to Chl a resulting from adaptation to varying environmental conditions. In order to include cyanobacteria in fluorometric analysis of algae, a simplified energy distribution model describing energy pathways in the cyanobacterial photosynthetic apparatus was conceptualized. Two sets of mathematical equations were derived from this model and tested. Fluorescence of cyanobacteria was measured with a new fluorometer at seven excitation wavelength ranges and at three detection channels (650, 685 and 720 nm) in vivo. By employing a new fit procedure, we were able to correct for variations in the cyanobacterial fluorescence excitation spectra and to account for other phytoplankton signals. The effect of energy-state transitions on the PC fluorescence emission of PBSs was documented. The additional use of the PC fluorescence signal in combination with our recently developed mathematical approach for phytoplankton analysis based on Chl fluorescence spectroscopy allows a more detailed study of cyanobacteria and other phytoplankton in vivo and in situ.

Cyanobacteria↗

PubMed2002

A new level detector for ion channel analysis.

The algorithm proposed here for automatic level detection in noisy time series of patch-clamp current is based on the detection of jump-free sections in the time series. The detector moves along the time series and uses a chi(2) test for the detection of jumps. When a jump is detected, the mean value, the variance and the length of the preceding jump-free section are stored. A Student's t-test was employed for the assignment of detected jump-free sections to discrete levels of the Markov model and for rejection of all sections with multiple assignments. The choice of the two significance levels is based on a 3-D diagram displaying the average number of detected levels from several time series vs. the significance levels of jump detection and of level assignment. The correct one is selected out of several plateaus with integer number of levels by means of the criterion of minimum scatter or other plausibility considerations. The test has been applied to simulated data obtained from a 2-state model and a 5-state aggregated Markov model, and the influences of SNR and of gating frequency are shown. Finally, the performance of the level detector is compared with a fit-by-eye and with a fit of the amplitude histogram by a sum of gaussians. At high noise, the fit of amplitude histograms failed, whereas the other two approaches were about equal.

Algorithms↗

PubMed2002

A chi(2) test for model determination and sublevel detection in ion channel analysis.

A chi(2) test is proposed that provides a means of discriminating between different Markov models used for the description of a measured (patch clamp) time series. It is based on a test statistic constructed from the measured and the predicted number of transitions between the current levels. With a certain probability, this test statistic is below a threshold if the model with a reduced number of degrees of freedom is compatible with the data. A second criterion is provided by the dependence of the test statistic on the number of data points. For data generated by the alternative model it increases linearly. The applicability of this test for verifying and rejecting models is illustrated by means of time series generated by two distinct channels with different conductances and by time series generated by one channel with two conductance levels. For noisy data, a noise correction is proposed, which eliminates noise-induced false jumps that would interfere with the test. It is shown that the test can also be extended to aggregated Markov models.

Chi-Square Distribution↗

PubMed2002

A fluorometric method for the differentiation of algal populations in vivo and in situ.

Fingerprints of excitation spectra of chlorophyll (Chl) fluorescence can be used to differentiate 'spectral groups' of microalgae in vivo and in situ in, for example, vertical profiles within a few seconds. The investigated spectral groups of algae (green group, Chlorophyta; blue, Cyanobacteria; brown, Heterokontophyta, Haptophyta, Dinophyta; mixed, Cryptophyta) are each characterised by a specific composition of photosynthetic antenna pigments and, consequently, by a specific excitation spectrum of the Chl fluorescence. Particularly relevant are Chl a, Chl c, phycocyanobilin, phycoerythrobilin, fucoxanthin and peridinin. A laboratory-based instrument and a submersible instrument were constructed containing light-emitting diodes to excite Chl fluorescence in five distinct wavelength ranges. Norm spectra were determined for the four spectral algal groups (several species per group). Using these norm spectra and the actual five-point excitation spectrum of a water sample, a separate estimate of the respective Chl concentration is rapidly obtained for each algal group. The results of dilution experiments are presented. In vivo and in situ measurements are compared with results obtained by HPLC analysis. Depth profiles of the distribution of spectral algal groups taken over a time period of few seconds are shown. The method for algae differentiation described here opens up new research areas, monitoring and supervision tasks related to photosynthetic primary production in aquatic environments.

Journal Article↗