3. TI was closely associated with a phasic increase in force ('aftercontraction'). Like TI, the aftercontraction was evoked by a preceding action potential or by the break of a strong depolarizing pulse.4. TI and the aftercontraction displayed similar wave forms although peak current preceded peak force by 50-100 msec. Both transients were enhanced by increasing the strength or duration of the preceding depolarization pulse. Both events were slowed as the potential level following the pulse was displaced in the negative direction.5. TI and the aftercontraction could be evoked in the absence of cardiotonic steroids by strongly elevating Ca0.6. Additional experiments were carried out to test the hypothesis that TI reflects an influx of Ca2+ ions. Mn inhibited TI but the development and removal of the inhibition lagged far behind the effects on the slow inward current.7. TI could be suppressed and eventually inverted by varying the membrane potential in the positive direction. The inversion potential averaged -5 mV and was not consistent with a Ca-specific pathway. The aftercontraction was more closely related to the phasic conductance change underlying the current than to the current flow itself.8. The results are consistent with the idea that an oscillatory release of Ca from an intracellular store is the primary event underlying both the aftercontraction and the conductance change which generates TI. Digitalis intoxication or very high Cao may promote such events by elevating intracellular Ca levels.

SUMMARY1. Voltage clamp experiments studied the ionic basis of the strophanthidininduced transient inward current (TI) in cardiac Purkinje fibres.2. The reversal potential of TI (Erev) was determined in the presence of various bathing solutions. Erev averaged -5 mV in the standard modified Tyrode solution (Kass, Lederer, Tsien & Weingart, 1978). Erev was displaced toward more negative potentials when the external Na concentration (Na0) was reduced by replacement of NaCl with Tris Cl, choline Cl or sucrose.3. 'A sudden reduction of Nao evoked a temporary increase in TI, followed after a few minutes by a sustained diminution. The initial increase was closely paralleled by an enhanced aftercontraction and could be explained by an indirect effect of Nao on internal Ca. The subsequent fall in TI amplitude could be accounted for by the reduced driving force, E -Erev. (1-8 mM).5. These results are consistent with an increase in membrane permeability to Na and perhaps K.6. TI was not directly affected by TTX, which blocks excitatory Na channels, or by Cs, which inhibits inwardly rectifying K channels. TI may be distinguished from the slow inward current by its kinetic, pharmacological and ionic properties.7. TI might be carried by a pre-existing ionic pathway such as the 'leak' channel which provides inward current underlying normal pace-maker depolarization. Another possibility is that TI reflects Ca extrusion by an electrogenic Ca-Na exchange.

A clone of human HeLa cells stably transfected with mouse connexin40 DNA was used to examine gap junctions. Two separate cells were brought into physical contact with each other ("induced cell pair") to allow insertion of gap junction channels and, hence, formation of a gap junction. The intercellular current flow was measured with a dual voltage-clamp method. This approach enabled us to study the electrical properties of gap junction channels (cell pairs with a single channel) and gap junctions (cell pairs with many channels). We found that single channels exhibited multiple conductances, a main state (gamma j(main state)), several substates (gamma j(substates)), a residual state (gamma j (residual state)), and a closed state (gamma j(closed state)). The gamma j(main state) was 198 pS, and gamma j(residual state) was 36 pS (temperature, 36-37 degrees C; pipette solution, potassium aspartate). Both properties were insensitive to transjunctional voltage, Vj. The transitions between the closed state and an open state (i.e., residual state, substate, or main state) were slow (15-45 ms); those between the residual state and a substate or the main state were fast (1-2 ms). Under steady-state conditions, the open channel probability, Po, decreased in a sigmoidal manner from 1 to 0 (Boltzmann fit: Vj,o = -44 mV; z = 6). The temperature coefficient, Q10, for gamma j(main state) and gamma j(residual state) was 1.2 and 1.3, respectively (p < 0.001; range 15-40 degrees C). This difference suggests interactions between ions and channel structure in case of gamma j(residual state). In cell pairs with many channels, the gap junction conductance at steady state, gj, exhibited a bell-shaped dependency from Vj (Boltzmann fit, negative Vj, Vj,o = -45 mV, gj(min) = 0.24; positive Vj, Vj,o = 49 mV, gj(min) = 0.26; z = 6). We conclude that each channel is controlled by two types of gates, a fast one responsible for Vj gating and involving transitions between open states (i.e., residual state, substates, main state), and a slow one involving transitions between the closed state and an open state.

Abstract-Gap junctions formed between transfected cells expressing connexin (Cx) 40 and Cx43 (Cx43-RIN, Cx40-HeLa, and Cx43-HeLa) revealed a relationship, g j ϭf(V j ), at steady state, that is typified by a nonsymmetrical behavior similar to that previously reported for other heterotypic channels (gap junction conductance [g j ]; transjunctional voltage [V j ]). The unitary conductance of the channels was sensitive to the polarity of V j . A main state conductance of 61 pS was found when the Cx43 cell was stepped positively or the Cx40 cell negatively (V j ϭ70 mV); the reverse polarities yielded a conductance of 100 pS. These heterotypic channels were permeable to carboxyfluorescein. In addition, two other heterotypic forms are illustrated to demonstrate that endogenous Cx45 expression cannot explain the results. The demonstration of heterotypic Cx40 -Cx43 channels may have implications for the propagation of the electrical impulse in heart. For example, they may contribute to the slowing of the impulse propagation through the junctions between Purkinje fibers and ventricular muscle.