Intracellular calcium buffering capacity in isolated squid axons

Changes in ionized calcium were studied in axons isolated from living squid by measuring absorbance of the Ca binding dye Arsenazo III using multiwavelength differential absorption spectroscopy. Absorption changes measured in situ were calibrated in vitro with media of ionic composition similar to axoplasm containing CaEGTA buffers. Calcium loads of 50-2,500 μmol/kg axoplasm were induced by microinjection, by stimulation in 112 mM Ca seawater, or by soaking in choline saline with 1-10 mM Ca. Over this range of calcium loading of intact axoplasm, the ionized calcium in the axoplasm rose about 0.6 nM/μM load. Similar loading in axons preteated with carbonyl cyanide 4- trifluoromethoxyphenylhydrazone (FCCP) to inhibit the mitochondrial proton gradient increased ionized calcium by 5-7 percent of the imposed load, i.e. 93-95 percent of the calcium load was buffered by a process insensitive to FCCP. This FCCP- insensitive buffer system was not saturated by the largest calcium loads imposed, indicating a capacity of at least several millimolar. Treatment of previously loaded axons with FCCP or apyrase plus cyanide produced rises in ionized calcium which could be correlated with the extent of the load. Analysis of results indicated that, whereas only 6 percent of the endogenous calcium in fresh axons is stored in the FCCP-sensitive (presumably mitochondrial) buffer system, about 30 percent of an imposed exogenous load in the range of 50-2,500 μM is taken up by this system.     

Growth advantage in stationary-phase (GASP) phenotype in long-term survival strains of Geobacter sulfurreducens

Geobacter sulfurreducens exists in the subsurface and has been identified in sites contaminated with radioactive metals, consistent with its ability to reduce metals under anaerobic conditions. The natural state of organisms in the environment is one that lacks access to high concentrations of nutrients, namely electron donors and terminal electron acceptors (TEAs). Most studies have investigated G. sulfurreducens under high-nutrient conditions or have enriched for it in environmental systems via acetate amendments. We replicated the starvation state through long-term batch culture of G. sulfurreducens, where both electron donor and TEA were scarce. The growth curve revealed lag, log, stationary, death, and survival phases using acetate as electron donor and either fumarate or iron(III) citrate as TEA. In survival phase, G. sulfurreducens persisted at a constant cell count for as long as 23 months without replenishment of growth medium. Geobacter sulfurreducens demonstrated an ability to acquire a growth advantage in stationary-phase phenotype (GASP), with strains derived from subpopulations from death- or survival phase being able to out-compete mid-log-phase populations when co-cultured. The molecular basis for GASP was not because of any detectable mutation in the rpoS gene (GSU1525) nor because of a mutation in a putative homolog to Escherichia coli lrp, GSU3370.     

Uptake of Trace Metals and Rare Earth Elements from Hornblende by a Soil Bacterium

Analysis of trace elements released from hornblende between pH 6.5 and 7.5 in the presence of Arthrobacter sp. shows that Fe, Ni, V, Mn, and, to a lesser extent, Co are preferentially released into solution relative to bacteria-free experiments. This enhanced release into solution could be due to contributions from the slightly lowered pH, the presence of low molecular weight organic acids (LMWOAs), or the presence of a catecholate siderophore in experiments with bacteria. The best explanation for enhanced metal release is siderophore complexation at the mineral surface followed by release to solution. However,the relative rates of metal release to solution in these experiments do not strictly follow the trend predicted by the relative ordering of metal hydrolysis, which might be predicted for siderophore-promoted dissolution. For some of these metals, release to solution is fast initially in biotic experiments, but concentrations in solution reach a steady state value or decrease with time as the bacteria cell numbers increase exponentially. Lack of enhanced release to solution for some metals and decreases in release rate with time for others may be explained by uptake into bacteria. Many of the metals predicted to strongly complex with siderophore (including Al, Ti, Fe, Cu) are heavily taken up into cellular material. The relative ordering of organic ligand-element complexation may therefore partially explain the relative ordering of uptake of trace metals and rare earth elements into cell material. Fractionation of heavy rare earth elements taken up into cellular material is also very strong, and increases from Ho to Lu. Strong fractionation in uptake of some elements by bacteria may create biological signatures either in the mineral substrate or in any mineral precipitates associated with the cellular material.