Characterization of outward K(+) currents in isolated smooth muscle cells from sheep urethra.

The perforated-patch technique was used to measure membrane currents in smooth muscle cells from sheep urethra. Depolarizing pulses evoked large transient outward currents and several components of sustained current. The transient current and a component of sustained current were blocked by iberiotoxin, penitrem A, and nifedipine but were unaffected by apamin or 4-aminopyridine, suggesting that they were mediated by large-conductance Ca(2+)-activated K(+) (BK) channels. When the BK current was blocked by exposure to penitrem A (100 nM) and Ca(2+)-free bath solution, there remained a voltage-sensitive K(+) current that was moderately sensitive to blockade with tetraethylammonium (TEA; half-maximal effective dose = 3.0 +/- 0.8 mM) but not 4-aminopyridine. Penitrem A (100 nM) increased the spike amplitude and plateau potential in slow waves evoked in single cells, whereas addition of TEA (10 mM) further increased the plateau potential and duration. In conclusion, both Ca(2+)-activated and voltage-dependent K(+) currents were found in urethral myocytes. Both of these currents are capable of contributing to the slow wave in these cells, suggesting that they are likely to influence urethral tone under certain conditions.

Hyperpolarisation-activated inward current in isolated sheep mesenteric lymphatic smooth muscle.

1. Freshly isolated sheep lymphatic smooth muscle cells were studied using the perforated patch-clamp technique. Hyperpolarisation with constant-current pulses caused a time-dependent rectification evident as a depolarising 'sag' followed by an anode-break overshoot at the end of the pulse. Both sag and overshoot were blocked with 1 mM Cs+. 2. Cells were voltage clamped at -30 mV and stepped to -120 mV in 10 mV steps of 2 s duration. Steps negative to -60 mV evoked a slowly activating, non-inactivating inward current which increased in size and rate of activation with increasing hyperpolarisation. 3. The slowly activating current was reduced in Na+-free bathing solution but enhanced when the extracellular K+ concentration was increased to 60 mM. The current was significantly reduced by 1 mM Cs+ and 1 microM ZD7288 but not by 1.8 mM Ba2+. 4. The steady-state activation curve of the underlying conductance showed a threshold at -50 mV and half-maximal activation at -81 mV. Neither threshold nor half-maximal activation was significantly affected by increasing the external K+ concentration to 60 mM. 5. The frequency of spontaneous contractions and fluid propulsion in isolated cannulated segments of sheep mesenteric lymphatics were decreased by 1 mM Cs+ and by 1 microM ZD7288. 6. We conclude that sheep lymphatics have a hyperpolarisation-activated inward current similar to the If seen in sinoatrial node cells of the heart. Blockade of this current slows spontaneous pumping in intact lymphatic vessels suggesting that it is important in normal pacemaking.

Effect of purinergic blockers on outward current in isolated smooth muscle cells of the sheep bladder

Freshly dispersed cells from sheep urinary bladder were voltage clamped using the whole cell and inside-out patch-clamp technique. Cibacron and Basilen blue increased outward current in a dose-dependent manner with a half-maximal response at 10(-5) M. Suramin, in concentrations to 10(-3) M, had no such effect. The Cibacron blue response was abolished in Ca2+-free physiological salt solution, suggesting that it was acting on a Ca2+-dependent current. Similarly, the Cibacron blue-sensitive current was significantly attenuated by charybdotoxin. Cibacron blue did not modulate inward current nor were its effects modified by caffeine or heparin, suggesting that its effect on outward current was not secondary to an increase in intracellular Ca2+. Application of 10(-4) M Cibacron blue to the inside membrane of excised patches caused a rapid increase in open probability of a large-conductance (300 pS) K+ channel. These results suggest that Cibacron blue is a potent activator of a Ca2+-dependent outward current in bladder smooth muscle cells in addition to its action as a purinergic blocker.

A novel radiation-resistant Deinobacter sp. isolated from irradiated pork

A red-pigmented, radiation-resistant, Gram-negative, rod-shaped bacterium isolated from irradiated pork is described. The D,, values in buffer solution and on pork mince are 3.45 and 5.05 kGy respectively. The strain has been identified as a Deinobacter species