Clamped modes

Figure 13.13. Patch-Clamp Modes. The patch-clamp technique for monitoring channel activity is highly versatile. A high-resistance seal (gigaseal) is formed between the pipette and a small patch of plasma membrane. This configuration is called cell attached. The breaking of the membrane patch by increased suction produces a low-resistance pathway between the pipette and interior of the cell. The activity of the channels in the entire plasma membrane can be monitored in this whole-cell mode. To prepare a membrane in the excised-patch mode, the pipette is pulled away from the cell. A piece of plasma membrane with its cytosolic side now facing the medium is monitored by the patch pipette.

Current clamp mode. In this mode the current is kept constant by the patch-clamp amplifier while voltage is measured, allowing recordings of the membrane potential. 

Under these space-clamped conditions the action potential is referred to as a membrane action potential. With the switch shown in Fig. 5a in the voltage-clamp position, the membrane potential is under control. The problem was to switch from zero current to voltage-clamp mode quickly at various times during the membrane action potential. 

This is a true single-channel patch clamp mode allowing the measurement of singlechannel currents with the added benefit of making it possible to change the medium to which the intracellular surface of the membrane is exposed. Thus, the inside-out configuration is particularly valuable when studying the influence of intracellular molecules on ion channel function, like calcium or ATP. 

The average of single-channel conductances was calculated from at least 150 channel events, which were measured under the voltage-clamp mode at the membrane potential of -1-50 mV in 1 M salt solution containing 2.5 mM HEPES (pH7.2)... 

The time-dependence of the force is determined by the protocol applied in the actual application of DFS. One common way to perform the experiments or simulations is the force-ramp mode, in which the applied force increases with a constant velocity, F t) = where k denotes the force constant of the pulling device. The other protocol, called force-clamp mode, crmsists in the application of a constant external force, F(t) = Fext- In the force-ramp case, one finds the logarithmic dependence of the mean rupture force and v quoted above. The simple model appears to work quite well for small pulling velocities but fails if one pulls fast. In this simation, more detailed calculations of the rupture force distributions via the computation of the mean first passage time in model free-energy landscapes give more reliable results [104]. 

If the reversible dynamics of bond rupture is considered in the force clamp mode, one can exploit an analogy to the treatment of single molecule fluorescence to treat the statistical properties of the transition events [108, 109]. If one considers two states, A (closed) and B (open) with rates k, = k(A —> B) and = (B A), the equilibrium constant is given by = A b/ a- Due to the strong exponential dependence of the kinetic rates on the external force, in the Bell model given by a( ) = and k F) = one can vary Kover a broad range. This... 

To overcome some of the problems associated with the variable frequency control of resonant converters, they are operated at a fixed frequency (Kit Sum, 1988 Bhat, 1988). A number of configurations and control methods for fixed-frequency operation are available in the literature (Bhat (1988) gives a list of papers). One of the most popular methods of control is the phase-shift control (also called clamped mode or PWM operation) method. Figure 10.89(b) illustrates the clamped mode fixed-frequency operation of the modified SRC shown in Fig. 10.89(a). The load power control is achieved by changing the phase-shift angle

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This is a true single-channel patch clamp mode allowing the measurement of single-channel currents with the... 

Figure 2 Hypoxia inhibits K+ channels in several 02-sensitive tissues, (a) Whole-ceU patch-clamp experiments show K" " current inhibition in response to hy poxia in rat resistance PASMCs. The complex I ETC blocker rotenone and the Kv blocker 4-aminopyridine (4-AP) also inhibit K" current, (b) Using the current clamp mode, resistance PASMCs depolarize in response to hypoxia. (From Ref. 29.) (c,d,e) Hypoxia inhibits outward K" " current in isolated cells from the NEB, CB, and PC-12 cells. (From Refs. 39,105,106.) Inset Proposed mechanism for HPy featuring the effector portion of the pathway.

The action of phosphonic acid substituted calix[4]arenes (701)-(704) on solvent-containing planar bilayer membranes made of cholesterol and egg phosphatidylcholine (egg PC) or synthetic 18-carbon-tail phospholipid DOPC have been investigated in a voltage-clamp mode. A steady-state voltage-dependent transmembrane current has been achieved only after addition of the compound (702) from the side of the membrane the positive potential has been applied to. This current exhibited anion selectivity passing more chloride at negative potentials applied from the side of the membrane to which calix[4]arene (702) has been introduced. The kinetics and temperature-dependence determined for calix[4]arene (702)-mediated ionic transport suggested a carrier mode of facilitated diffusion. 

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