Imaging in five dimensions: time-dependent membrane potentials in individual mitochondria.

Because of its importance in the chemiosmotic theory, mitochondrial membrane potential has been the object of many investigations. Significantly, however, quantitative data on how energy transduction might be regulated or perturbed by the physiological state of the cell has only been gathered via indirect studies on isolated mitochondrial suspensions; quantitative studies on individual mitochondria in situ have not been possible because of their small size, their intrinsic motility, and the absence of appropriate analytical reagents. In this article, we combine techniques for rapid, high resolution, quantitative three-dimensional imaging microscopy and mathematical modeling to determine accurate distributions of a potentiometric fluorescent probe between the cytosol and individual mitochondria inside a living cell. Analysis of this distribution via the Nernst equation permits assignment of potentials to each of the imaged mitochondrial membranes. The mitochondrial membrane potentials are distributed over a narrow range centered at -150 mV within the neurites of differentiated neuroblastoma cells. We find that the membrane potential of a single mitochondrion is generally remarkably stable over times of 40-80 s, but significant fluctuations can occasionally be seen. The motility of individual mitochondria is not directly correlated to membrane potential, but mitochondria do become immobile after prolonged treatment with respiratory inhibitors or uncouplers. Thus, three spatial dimensions, a key physiological parameter, and their changes over time are all quantitated for objects at the resolution limit of light microscopy. The methods described may be readily extended to permit investigations of how mitochondrial function is integrated with other processes in the intact cell.     

Effects of estrogen deprivation on human benign prostatic hyperplasia

Sex steroids are thought to play an essential role in the pathogenesis of human benign prostatic hyperplasia (BPH). Since recent studies in animal models and in men have shown that estrogens might be causally linked to the onset and maintenance of BPH, we examined the effect of 1-methyl-androsta-1,4-diene-3,17-dione (Atamestane), a newly developed aromatase inhibitor, in men with BPH. In an open multicenter study 49 men (mean age 70.1 years, range 55 to 84) with obstructive BPH were treated with atamestane (3 × 200 mg/day) for 3 months. Of the 49 patients 44 completed the treatment period; the other patients discontinued the study for reasons unrelated to treatment. With treatment BPH-related symptoms such as daytime voiding frequency, nycturia, peak flow and residual urine improved considerably; however, these parameters did not reach statistical significance. The mean prostatic volume decreased significantly from 74.2 ± 31.7 to 64.0 ± 31 ml (mean ± SD). Serum estrogen levels decreased markedly during treatment. In addition intraprostatic estrogen concentration decreased with treatment as compared to estrogen levels in hyperplastic prostates from untreated patients. The following conclusions can be drawn from this study: first, estrogens appear to have an important supportive role in established BPH, and second, estrogen deprivation improved BPH-related symptoms and reduced significantly prostatic volume.     

Growth and Osmotic Adjustment of Cultured Suspension Cells from Alternanthera philoxeroides (Mart.) Griseb After an Abrupt Increase in Salinity

We have developed a cell suspension culture from alligator weed(Alternanthera philoxeroides [Mart.] Griseb), a C₃ member ofthe Amaranthaceae. Intact plants of alligator weed can growat 400 mol m^–3 NaCl. Growth of alligator weed suspensionswas compared to growth of tobacco (Nicotiana tabacum L. cv.Wisconsin 38) suspensions after subculture to 200 mol m^–3NaCl. Fresh weight and cell density of salt-treated alligatorweed suspensions more than doubled by 7 d after subculture,but the fresh weight of salt-treated tobacco suspensions didnot double during the 21 d experiment. Correspondingly, cellviability dropped from about 90% to 77% in alligator weed andto 41% in tobacco, at 1 d after subculture to 200 mol m^–3NaCl. The symplastic volume of alligator weed cells declined36% by 2 h after subculture to 200 mol m^–3 NaCl, but cellcontents became iso-osmotic with the media at this point. Between2 h and 6 h there was a further decrease in osmotic potential,an increase in turgor potential and a partial recovery of symplasticvolume. Turgor potential was similar to that in control cellsby 24 h, indicating significant osmotic adjustment. Turgor potentialsremained similar in both treatments from 24 h through 21 d butthe average symplastic volume of salt-treated cells was 11 %less than in control cells. Therefore, alligator weed suspensioncells exhibit a rapid recovery of water balance and cell growthafter an abrupt and substantial increase in salinity. Key words: Cell culture, growth, osmotic adjustment, salinity, turgor potential