Acute stress inhibits food intake and alters ghrelin signaling in the brain of tilapia (Oreochromis mossambicus).

This study investigated the effect of an acute stress on food intake and on the expression of neuropeptide Y (NPY), corticotropin-releasing hormone (CRH), and ghrelin and its receptors, growth hormone secretagogue receptors (GHSRs) in the tilapia (Oreochromis mossambicus). Food intake was significantly (P < 0.01) reduced in fish after a 30-min crowding and handling stress. In a second group of animals exposed to the same 30-min stressor, tissue samples were collected immediately after the stressor to determine changes in the neuroendocrine regulators of food intake. Although CRH and NPY are considered the major mediators of appetite during stress, both mRNA levels were unaltered in the telencephalon/pre-optic area and in the hypothalamic/optic tectum. Interestingly, there was an elevation in the ghrelin transcript (P < 0.05) in the telencephalon/pre-optic area and elevation of its functional receptor (GHSR1a-LR) (P < 0.001) in the hypothalamic/optic tectum. Elevation of GHSR-LR heteronuclear RNA (P < 0.01) in the telencephalon/pre-optic area and suppression in the hypothalamic/optic tectum (P < 0.001) suggest rapid control of the ghrelin regulatory system in response to acute stress. These results suggest that ghrelin signaling is altered during acute stress. It is not clear if these changes result in altered feeding behavior because no changes in CRH or NPY mRNA expression were observed or if ghrelin is playing a role in regulating overall metabolic changes after acute stress.

An efficient disk based data structure for rapid searching of quantitative two-dimensional gel databases.

Fast access of two-dimensional (2-D) gel quantitative databases is important for rapid searching for protein differences between sets of 2-D gels from an experiment. The GELLAB-II system organizes corresponding spots from the gels in the database into reference or "Rspot" sets. These Rspot numeric names index fixed regions in the paged composite gel database file. This is adequate for an existing database, but has several problems. (i) Building the initial database requires guessing how much disk space to pre-allocate for each corresponding spot (i.e. spots from different gels). If it ever runs out of pre-allocated space during this process, it must expand the size of each corresponding set of spots copying the old database data into the new in-place on the disk. (ii) When adding new gels or editing the database, if a new spot is created, the system may also go into this expansion mode. The time spent and wasted disk space can be appreciable--depending on the size of the database (order of 100 gel database). (iii) Because each set of corresponding spots is the same size, we waste space in most spot sets since they do not require the additional space a few spot sets require which contain additional fragmented spots. We present a new low-level disk object-based structure and algorithm, paged indexed buckets (PIB), which optimizes disk space usage while having similar retrieval speed to the original method.

A fast spot segmentation algorithm for two-dimensional gel electrophoresis analysis.

An important issue in the automation of two-dimensional gel electrophoresis image analysis is the detection and quantification of protein spots. A spot segmentation algorithm must detect, define the extent of, and measure the integrated density of spots under a wide variety of actual gel image conditions. Besides these functions, the algorithm must be memory efficient to be able to process very large gel images and do this in a reasonable amount of computation time on low-cost computers, such as workstations and personal computers. We have developed a fast spot segmentation algorithm, extending the GELLAB-II segmenter, which extracts spots in a single raster scanning pass through the gel image. The performance analysis of the algorithm will be given in the paper as well as a discussion of the algorithm.

Splitting merged spots in two-dimensional polyacrylamide gel electrophoresis gel images.

We describe a heuristic computer algorithm using boundary analysis for improving spot finding and spot quantitation of large saturated or near-saturated spots in two-dimensional polyacrylamide electrophoresis gels. This spot quantitation is done using spot segmentation, which consists of spot finding and subsequent quantification steps. Occasionally, clusters of large saturated spots may become merged during spot finding. To correct this, the merged spots must be cut apart before quantitation. It is generally obvious from viewing the merged spot's border where they should be cut--at opposing saddlepoints (concavities in the boundary). The algorithm uses an analysis of the missegmented spot's boundary when a saturated spot is detected. If a near-saturated spot is larger than a given size, the spot segmenter program attempts to merge saturated fragments. When merging occurs, the segmenter program analyses the boundary to see if the spot should be split. The new algorithm first finds all robust concavities and then tries to match complementary ones. These paired concavities are then used to guide cutting of the missegmented spot into two or more separate spot regions. Finally, control is returned to the segmenter program to reprocess the data as a set of smaller separated spots.

Comparison of the Bio Image Visage 2000 and the GELLAB-II two-dimensional electrophoresis image analysis systems.

To compare the Visage 2000 analysis system (Bio Image, Ann Arbor, MI, USA) with the GELLAB-II analysis system (National Cancer Institute, Frederick, MD, USA), we used each to perform image analysis of the same 29 silver-stained two-dimensional electrophoresis (2DE) gel image files from a study of urinary proteins in metal recovery plant workers who had confirmed body burdens of cadmium. Visage, aided by interactive analysis, detected an average of 890 +/- 177.6 spots per gel, or a total of 25,800 spots, whereas GELLAB-II detected 1971 +/- 198.5 spots per gel, or a total of 57,160 (a 222% increase over the Visage system), without operator intervention. Visage automatically quantified 52.5% (13,556) of the spots; 47.2% (12,173), consisting mostly of larger spots, had to be quantified interactively with an image editor, and 0.3% (71) were not quantified. GELLAB-II automatically quantified all detected spots. After we interactively assigned the maximum allowed number of landmarks (30 for Visage and 52 for GELLAB-II), we found that Visage matched 657 +/- 211.2 spots per gel, and GELLAB-II matched all detected spots and also extrapolated an average of 1269 virtual spots per gel. Plots of densities from the two systems on selected spots showed excellent agreement, and both systems showed high correlation between their measurements of the beta-2-microglobulin spot densities and an independent radioimmunoassay quantification of the original urine samples. By comparing the regression of the densities of all spots with urinary cadmium (UCD) levels, we found that several of the same detected spots from each system were highly correlated. The densities of four acidic proteins with relative molecular weights of approximately 112,000 Da (as quantified by GELLAB-II but not by Visage) were highly correlated with UCD concentrations. These proteins are new candidate biomarkers of cadmium toxicity. We compared the estimated labor costs of using each system to analyse a hypothetical 20-sample (60 gels) 2DE study and found that GELLAB-II was six times less expensive to use than Visage, primarily because of the operator time required to do interactive error correction with the Visage system.

Study into the ability of patients with impaired lung function to use breath alcohol testing devices.

A subject who fails to provide an adequate breath sample for a breath alcohol measuring device under the provisions of Road Traffic Act 1988 may be charged with refusing to supply a sample unless the police officer believes the person is genuinely unable to do so. Subjects who are confronted with this situation may approach their general practitioner or chest physician for advice on whether they are medically able to provide adequate breath samples to satisfy the breath testing devices. There is currently no guidance available for medical practitioners concerning respiratory performance or lung function which will impair the use of such breath testing devices. This paper describes experiments with human volunteers suffering from respiratory illnesses and their ability to provide adequate breath samples to satisfy the requirements of the breath alcohol testing devices used in Great Britain. It was found that the most suitable parameters for determining whether a subject was capable of using a breath alcohol testing device were spirometry measurements of Forced Expired Volume in one second (FEV1) and Forced Vital Capacity (FVC). In this study subjects with a FEV1 less than 2.0 litres and a FVC less than 2.6 litres were generally unable to use all the devices.

The effect of salbutamol on breath alcohol testing in asthmatics.

Subjects suffering from asthma can occasionally experience difficulty in providing adequate breath samples for evidential breath alcohol testing devices and may therefore resort to the use of bronchodilators such as salbutamol to improve their respiration. Experiments showed that although salbutamol caused bronchodilation it did not affect breath alcohol levels of asthmatics who have been drinking. The blood:breath alcohol ratios obtained from asthmatics were within the normally recorded range before and after use of salbutamol. We conclude that the use of salbutamol by asthmatics does not affect the reliability of measurements made by evidential breath alcohol testing devices.