cyclic-gmp and gluconic-acid

Five mutants of Pseudomonas taiwanensis VLB120ΔCeGFP showed significant autoaggregation when growing on defined carbohydrates or gluconate, while they grew as suspended cells on complex medium and on organic acids like citrate and succinate. Surprisingly, the respective mutations affected very different genes, although all five strains exhibited the same behaviour of aggregate formation. To elucidate the mechanism of the aggregative behaviour, the microbial adhesion to hydrocarbons (MATH) assay and contact angle measurements were performed that pointed to an increased cell surface hydrophobicity. Moreover, investigations of the outer layer of the cell membrane revealed a reduced amount of O-specific polysaccharides in the lipopolysaccharide of the mutant cells. To determine the regulation of the aggregation, reverse transcription quantitative real-time PCR was performed and, irrespective of the mutation, the transcription of a gene encoding a putative phosphodiesterase, which is degrading the global second messenger cyclic diguanylate, was decreased or even deactivated in all mutants. In summary, it appears that the trophic autoaggregation was regulated via cyclic diguanylate and a link between the cellular cyclic diguanylate concentration and the lipopolysaccharide composition of P. taiwanensis VLB120ΔCeGFP is suggested.

Topics: Bacterial Adhesion; Carbohydrate Metabolism; Culture Media; Cyclic GMP; Gene Deletion; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Gluconates; Hydrophobic and Hydrophilic Interactions; Pseudomonas; Real-Time Polymerase Chain Reaction; Surface Properties

Mammalian vestibular organs have two types of hair cell, type I and type II, which differ morphologically and electrophysiologically. Type I hair cells alone express an outwardly rectifying current, I(K, L), which activates at relatively negative voltages. We used whole cell and patch configurations to study I(K,L) in hair cells isolated from the sensory epithelia of rat semicircular canals. I(K,L) was potassium selective, blocked by 4-aminopyridine, and permeable to internal cesium. It activated with sigmoidal kinetics and was half-maximally activated at -74.5 +/- 1.6 mV (n = 35; range -91 to -50 mV). It was a very large conductance (91 +/- 8 nS at -37 mV; 35 nS/pF for a cell of average size). Patch recordings from type I cells revealed a candidate ion channel with a conductance of 20-30 pS. Because I(K,L) was activated at the resting potential, the cells had low input resistances (R(m)): median 25 MOmega at -67 mV versus 1.3 GOmega for type II cells. Consequently, injected currents comparable to large transduction currents (300 pA) evoked small (

Topics: 4-Aminopyridine; 8-Bromo Cyclic Adenosine Monophosphate; Animals; Cyclic GMP; Gluconates; Guanylate Cyclase; Hair Cells, Vestibular; Membrane Potentials; Molsidomine; Nitric Oxide; Nitric Oxide Donors; Patch-Clamp Techniques; Penicillamine; Potassium; Potassium Channel Blockers; Rats; Rats, Long-Evans; Semicircular Canals; Signal Transduction

Inward rectifying K channel activity was observed in the luminal membrane of the marginal cell by use of the patch clamp technique. When pipette solution contained 150 mM KCl, the current reversed at 10.2 +/- 8.8 mV (N = 27). The channel current showed multiple subconductance levels and depolarization activated the channel very slowly (order of seconds). Current-voltage relationships in cell-attached mode showed inward rectification. The calculated slope conductance of inward and outward currents were 18 +/- 7 pS and 5.4 +/- 2.0 pS (N = 27), respectively. When pipette solution contained 75 mM NaCl and 75 mM KCl, the current reversed -7.0 +/- 7.6 mV (N = 6). When pipette solution contained NaCl or Na gluconate, the channel currents were outwardly directed within a range of -60 to 60 mV. These results suggested that the channel is selective to K+. The channel showed immediate run down after excision, and this rundown was not prevented by elimination of cytoplasmic Ca2+. The channel activity was not affected by an increase of intracellular cAMP and cGMP, an application of cytoplasmic catalytic subunit of PKA with ATP. Taking its density into consideration, the channel seems to contribute considerably to the K+ conductance of the luminal membrane of the marginal cell.

Topics: 1-Methyl-3-isobutylxanthine; Animals; Bucladesine; Calcium; Colforsin; Cyclic AMP; Cyclic GMP; Cytoplasm; Electric Stimulation; Electrophysiology; Gluconates; Guinea Pigs; Membrane Potentials; Patch-Clamp Techniques; Potassium Channels; Sodium Chloride; Stria Vascularis