The Viking mission: Implications for life on Mars

Summary The results of the Viking Biology experiments are best explained by non-biological phenomena: The interaction of the reagents with the materials comprising the regolith. Conditions of water activity, temperature, availability of carbon sources and others in most regions of the planet are too extreme for survival and growth of any known Earth microorganisms. Although the possibility persists that some very unusual form of life is somewhere on that planet the evidence is best interpreted as negative. Even though there is no evidence for current life on Mars, whether or not life ever originated there is not known.     

Glycerol permeability of human spermatozoa and its activation energy

Glycerol has commonly been employed as a cryoprotectant in cryopreservation of human spermatozoa. However, the addition of glycerol into the sperm before freezing and the removal of glycerol from the sperm after freezing and thawing result in anisotonic environments to the cells, which can cause cell injury. To define optimal procedures for the addition/removal of glycerol and to minimize the cell injury, one needs to know the kinetics of glycerol permeation across the sperm plasma membrane at different temperatures. For this, one has to determine the permeability coefficient of glycerol (Pg) and its activation energy (Ea). Values of Pg at different temperatures and at different glycerol concentrations were determined by measuring the time required for 50% spermolysis in hyperosmotic glycerol solutions which were hypotonic with respect to electrolytes. Value of the Ea was determined assuming an Arrhenius type temperature dependence of Pg. A dual fluorescent staining technique (propidium iodide and 6-carboxyfluoroscein diacetate) and flow cytometry were used to measure the spermolysis. The values of Pg in 0.5, 1.0, 1.5, and 2.0 M glycerol at 22 degrees C are 1.62, 1.88, 1.68, and 1.54 x 10(-3) cm/min, respectively. The values of Pg in 1 M glycerol at 0, 8, 22, and 30 degrees C are 0.33, 0.54, 1.88, and 2.60 x 10(-3) cm/min, respectively. The value of Ea is 11.76 kcal/mol.     

Disruptive medical patients. Forensically informed decision making.

Patients who disrupt medical care create problems for physicians. The risks are not entirely clinical. Although these patients may compromise sound clinical judgment, some are also litigious and express their dissatisfaction in legal or other forums. It then becomes necessary for treating physicians to be aware of the legal and ethical boundaries of their patient care responsibilities. Some disruptive patients are treated by setting limits, which is usually affirmed by health care agreements. A hospital review board may advise clinicians on these agreements and on the management of disruptive patients. If termination of the physician-patient relationship is considered, physicians must follow proper protocol. We examine these forensic considerations and place them in the context of malpractice. Communication, consultation, and documentation are the key elements in reducing liability.     

Color regulation in the archaebacterial phototaxis receptor phoborhodopsin (sensory rhodopsin II)

Phoborhodopsin, a repellent phototaxis receptor in Halobacterium halobium, exhibits vibrational fine structure, a feature that has not been identified for any other rhodopsin pigment at physiological temperatures. This conclusion follows form analysis of the absorption properties of the pigment in H. halobium membranes containing native retinal and an array of retinal analogues. The absorption spectrum of the native pigment has a maximum at 487 nm with a pronounced shoulder at 460 nm; however, the bandwidth is that expected for a single retinylidene species. Gaussian band-shape simulation with a spacing corresponding to the vibrational frequencies of polyene stretching modes reproduces the structured absorption spectra of native pigment as well as of analogue phoborhodopsin. Absorption shifts produced by a series of dihydroretinal and other retinal analogues strongly indicate that the dominant factor regulating the color of the pigment is planarization of the retinal ring with respect to the polyene chain.     

Pyrolytic Methylation-Gas Chromatography of Whole Bacterial Cells for Rapid Profiling of Cellular Fatty Acids

A novel, on-line derivatization technique has been developed which enables generation of fatty acid methyl ester (FAME) profiles from microorganisms by gas chromatography-mass spectrometry without the need for laborious and time-consuming sample preparation. Microgram amounts of bacterial cells are directly applied to a thin ferromagnetic filament and covered with a single drop of methanolic solution of tetramethylammonium hydroxide. After air drying, the filament is inserted into a special gas chromatograph inlet equipped with a high-frequency coil, thus enabling rapid inductive heating of the ferromagnetic filament. This so-called Curie-point heating technique is shown to produce patterns of bacterial FAMEs which are qualitatively and quantitatively nearly identical to those obtained from extracts of methylated lipids prepared by conventional sample pretreatment methods. Relatively minor differences involve the loss of hydroxy-substituted fatty acids by the pyrolytic approach as well as strongly enhanced signals of FAMEs derived from mycolic acids. This type of pyrolysis enables on-line derivatization and thermal extraction of volatile derivatives for analysis, whereas the residual components remain on a disposable probe (ferromagnetic wire) of a pyrolytic device. The reduced sample size (micrograms instead of milligrams) and the lack of sample preparation requirements open up the possibility of rapid microbiological identification of single colonies (thus overcoming the need for time-consuming subculturing) as well as analysis of FAME profiles directly from complex environmental samples.     

Effect of DNA gyrase inhibitors on gene expression of the cysteine regulon

Summary Nalidixic acid inhibits the expression of those cysteine genes which are regulated by the cysB product, it has no affect, however, on the constitutively expressed cysE gene. The expression of cysteine genes in a strain carrying a mutation in the nalA locus is resistant to this drug. Novobiocin affects the expression of cysteine genes similarly to nalidixic acid. The effect of nalidixic acid on the expression of genes in a cysteine constitutive mutant was studied.     

Relative contributions of the fraction of unfrozen water and of salt concentration to the survival of slowly frozen human erythrocytes.

As suspensions of cells freeze, the electrolytes and other solutes in the external solution concentrate progressively, and the cells undergo osmotic dehydration if cooling is slow. The progressive concentration of solute comes about as increasing amounts of pure ice precipitate out of solution and cause the liquid-filled channels in which the cells are sequestered to dwindle in size. The consensus has been that slow freezing injury is related to the composition of the solution in these channels and not to the amount of residual liquid. The purpose of the research reported here was to test this assumption on human erythrocytes. Ordinarily, solute concentration and the amount of liquid in the unfrozen channels are inversely coupled. To vary them independently, one must vary the initial solute concentration. Two solutes were used here: NaCl and the permeating protective additive glycerol. To vary the total initial solute concentration while holding the mass ratio of glycerol to NaCl constant, we had to allow the NaCl tonicity to depart from isotonic. Specifically, human red cells were suspended in solutions with weight ratios of glycerol to NaCl of either 5.42 or 11.26, where the concentrations of NaCl were 0.6, 0.75, 1.0, 2.0, 3.0, or 4.0 times isotonic. Samples were then frozen to various subzero temperatures, which were chosen to produce various molalities of NaCl (0.24-3.30) while holding the fraction of unfrozen water constant, or conversely to produce various unfrozen fractions (0.03-0.5) while holding the molality of salt constant. (Not all combinations of these values were possible). The following general findings emerged: (a) few cells survived the freezing of greater than 90% of the extracellular water regardless of the salt concentration in the residual unfrozen portion. (b) When the fraction of frozen water was less than 75% the majority of the cells survived even when the salt concentration in the unfrozen portion exceeded 2 molal. (c) Salt concentration affected survival significantly only when the frozen fraction lay between 75 and 90%. To find a major effect on survival of the fraction of water that remains unfrozen was unexpected. It may require major modifications in how cryobiologists view solution-effect injury and its prevention.     

Permeability of the 17-day fetal rat pancreas to glycerol and dimethylsulfoxide

Experimentally induced diabetes in rats can be reversed by the transplantation of several fresh or frozen-thawed fetal pancreases. An important question to both the mechanistic and practical aspects of cryobiology is the role played by the permeation of protective additives during freezing, thawing, and subsequent dilution. Answers require knowledge of the kinetics of permeation of the specific additive into the cell or tissue. In this paper, we report isotopic measurements of the rate of permeation of 2 M glycerol and 1 and 2 M dimethylsulfoxide (Me₂SO) into 17-day fetal pancreases at 0 and 22 °C. In Me₂SO, equilibrium was achieved in about 10–15 min at 0 °C and in less than 10 min at 22 °C. In glycerol, equilibrium was attained in about 60 min at 22 °C; but at 0 °C permeation was only 65% complete after 180 min. In general, Me₂SO permeated 10–30 times more rapidly than glycerol at 0 °C, and glycerol permeated about 10 times more rapidly at 22 than at 0 °C.The kinetics of permeation were more characteristic of a two-compartment than a single-compartment system. In all probability, the two compartments are the intercellular space and the intracellular space. The permeability data suggest that each compartment occupies about half the total volume.