Detection of total- and partial-body irradiation in a monkey model: a comparative study of chromosomal aberration, micronucleus and premature chromosome condensation assays.

PURPOSE: To investigate the efficacy of three cytogenetic methods (dicentrics, micronuclei (MN) and premature chromosome condensation (PCC) analysis) for assessment of the unirradiated fraction and the persistence of damage after total-body (TB) and partial-body (PB) irradiation of rhesus monkeys (Macaca mulatta). MATERIALS AND METHODS: Animals were exposed to X-rays (5 Gy), either TB or PB, with about 6% of marrow cells shielded. Blood samples were collected at different times after exposure, i.e. 1, 3 and 7 days, and cultures were set up for the different cytogenetic endpoints. In addition, blood count analysis was performed before and after irradiation. RESULTS: Blood count analysis was not suitable for discriminating between TB and PB exposure. By using Poisson or overdispersion distribution as the basis, it was not possible to distinguish TB from PB irradiation when dicentric chromosomes and MN were analysed. PCC analysis, in contrast, showed a Poisson distribution after TB exposure and overdispersion after PB exposure. Using the PCC assay, reliable dose estimates could be obtained up to 7 days after irradiation. CONCLUSIONS: For dicentrics and MN, shielding of 6% of bone marrow cells was found to be too small to estimate the unirradiated fraction accurately. The PCC technique was useful for dose assessment and the inhomogeneous exposure of 6% was detected within a short period of time after exposure.

PCDOSE: an interactive software system to calculate internal radiation dose on a personal computer.

An interactive, menu directed, software system to calculate committed dose equivalents for individuals with different physiques after inhalation, ingestion or injection of radionuclides has been developed. The calculations are based on ICRP 26/30 methods. The programs are written in PASCAL and can be implemented on a personal computer with a MS-DOS operating system and a hard disk with a storage capacity of at least 20 Mb. This paper describes the development and features of the system.

Models for the electronic processing of flow cytometric data at high particle rates.

In flow cytometry at high particle rates, complete processing of data is limited by the capacity of the flow cytometer electronics which are constrained by the waiting or cycling time of the processor. Four models of impulse processors were analyzed to study the influence of the waiting time and reset mechanisms on the input-output properties of commonly used electronic devices. The models contain a feedback loop to represent a waiting time and describe reset mechanisms to filter trains of consecutive pulses such as clumps and doublets. Discrete systems analytical tools have been used to derive formulas for the yield of a simple waiting time device, a doublet filter, a clump filter and a clump and doublet filter. Also, the response to a sudden onset of an input signal has been analyzed and is described. The reset mechanism is an important determinant of the capacity of a waiting time device depending on the impulse rate of the input signal.

Coincidence in high-speed flow cytometry: models and measurements.

In flow cytometry, the coincident arrival of particles becomes a major problem when high sample rates are required. For the development of our high-speed photodamage flow cytometer (ZAPPER), it was of importance to understand the behavior of cells at flow rates of around 50,000-250,000 event/s. We developed and compared two models that describe the relation between the real cell rate and the detectable single cell rate. Both the Computer Simulation model and the Input/Output Device model show distinct optima for the cell rate. The models were compared to measurements performed on the ZAPPER-prototype. Fits of the two models to the experimental data were excellent for cycle times of 4 and 15 microseconds and acceptable for a 2 microseconds cycle time. A third model (Mercer WB, Rev. Sci. Instr. 37:1515-1521,1966) could be fitted to the experimental data, after the proportionality constant k was adapted to the experimental data. At a yield of detectable single cells of 70%, the maximum cell rates are 180,000, 100,000, and 40,000 cells/s for cycle times of 2, 4, and 15 microseconds, respectively. Based on these results we can now select an optimal cell rate for analysis and sorting based on criteria such as accepted cell loss. In addition, the advantages of reducing the cycle time can now be evaluated with respect to the costs of that modification.

Confocal microscopy as a tool for the study of the intranuclear topography of chromosomes.

A scanning confocal microscope was used to investigate the spatial positions of specific regions within blood cell nuclei. These centromeric regions were fluorescently labelled by in-situ hybridization to suspended nuclei with a centromere-1-specific DNA probe. The 3-D image data sets, obtained by optical sectioning of the cells, were used to determine the spatial position of the centromeric regions in the nuclei by means of specially developed software. The centromeres were found to be localized near the nuclear boundary. This spatial pattern was tested against a random distribution model by means of the Kolmogorov-Smirnov test. The difference between the two patterns was at a P less than 0.01 significance level.

Spatial topography of a pericentromeric region (1q12) in hemopoietic cells studied by in situ hybridization and confocal microscopy.

A fluorescent in situ hybridization procedure with a chromosome 1-specific (1q12) repetitive satellite DNA probe was used to label the 1q12 regions of the chromosomes 1 in spherical and polymorphic hemopoietic cell nuclei. The entire procedure was performed in suspension to preserve nuclear morphology. The result was studied by three-dimensional analysis, as provided by a scanning laser confocal microscope. The 1q12 regions of chromosome 1 were measured to be closely associated with the nuclear envelope in isolated nuclei of unstimulated diploid human lymphocytes. The relative positions to each other in the periphery of these spherical nuclei could not be distinguished from a random distribution pattern. In the diploid and tetraploid polymorphic nuclei of cells of the promyelocytic leukemia cell line HL60 these pericentromeric sequences were also associated with the nuclear surface.

Radiosensitivity of cells in recurrent experimental tumours and the effectiveness of tumour retreatment.

Two experimental tumour models, a rat rhabdomyosarcoma (R-1) and a rat urether carcinoma (RUC-2) have been employed to evaluate the X-ray sensitivity of tumours recurrent after primary treatments with various doses of X-rays and to correlate changes in volume responses with the cellular radiosensitivity. The responsiveness of R-1 tumours, assessed from the volume reduction as a function of the time after treatment, was less for recurrent tumours, but their growth delay was slightly increased, while the X-ray sensitivity of the tumour cells, assessed by cell survival, was equal to that of the controls. For RUC-2 tumours, however, the reduction in volume after irradiation of the recurrent tumour was larger than after primary treatment, the growth delay was increased, but cell survival curves were not significantly different from those of the controls. It is concluded that differences in volume responses between untreated tumours and recurrent tumours are largely determined by a tumour bed effect (TBE) and that changes in cellular radiosensitivity in these tumours do not play a significant part.

Electric and magnetic fields of the brain computed by way of a discrete systems analytical approach: theory and validation.

It is shown how the stationary volume conduction phenomena in the brain, namely the electric and magnetic fields can be described in discrete terms. The volume conductor is sampled in space by introducing a sampling distance corresponding to the uncertainty in the measurements. In this way, a three-dimensional lattice is needed with equidistantly spaced nodes. The electric and magnetic properties of such a lattice are assumed to be equivalent to that of brain and other tissues. The electric and magnetic potential fields are calculated for each node as the output of a linear feedback system which has the impressed currents as the input. By way of the feedback loop the reflection phenomena at the boundaries between media of different conductivity can be taken into account. This discrete formalism has been implemented in a software system. To demonstrate the validity and accuracy of this system a number of analytically tractable problem in volume conduction has been evaluated.

The double dipole model of theta rhythm generation: simulation of laminar field potential profiles in dorsal hippocampus of the rat.

A set of compartmental models of CA1 pyramidal, granular and polymorph cells of the dorsal hippocampus have been used to simulate membrane potentials generated by synaptic activation at various levels along these cells. From the membrane potential distributions the field potentials in dorsal CA1 and the dorsal blade of the dentate area have been simulated using a model based on volume conduction theory. Field potential profiles similar to laminar profiles, found experimentally in the dorsal hippocampus during theta rhythm, could only be simulated by assuming (almost) simultaneous synaptic excitation of the 3 cell types at given sites. The results lead to 2 alternative models for the simultaneous excitation of CA1 pyramidal cells and dentate granular cells during theta rhythm. Other electrophysiological evidence favours the model in which the two neuronal populations are activated distally near the fissure.

Relative contributions of intracortical and thalamo-cortical processes in the generation of alpha rhythms, revealed by partial coherence analysis.

The thalamo-cortical relationships of alpha rhythms have been analysed in dogs using partial coherence function analysis. The objective was to clarify how far the large intracortical coherence commonly recorded between different cortical sites could depend on a common thalamic site. It was found that the alpha rhythms of the LGN influenced only moderately the coherence between cortical alpha rhythms whereas that of the pulvinar had much more influence, at least in relation to some cortical areas. Significant phase shifts between thalamus and cortex were also measured. The results demonstrate that there are thalamo-cortical and cortico-cortical components which interact in the generation of cortical alpha rhythms.

A computer system for the analysis and synthesis of field potentials.

It is shown how field potentials due to neuronal membrane processes can be computed by means of a digital computer system. The essence of the method consists in evaluating a discrete expression for the field potential as a function of membrane potentials. This expression is considered as a three-dimensional convolution. In the computer system this convolution is performed by multiplication in the spatial Fourier domain. Introduction of a density function provides the possibility of evaluating in an easy way the field potential of a population of neurons. To demonstrate the applicability of the system, field potentials of different membrane potential configurations in the brain have been computed.