Physiological modulation of inactivation in L-type Ca2+ channels: one switch

Abstract

The relative contributions of voltage- and Ca(2+)-dependent mechanisms of inactivation to the decay of L-type Ca(2+) channel currents (I(CaL)) is an old story to which recent results have given an unexpected twist. In cardiac myocytes voltage-dependent inactivation (VDI) was thought to be slow and Ca(2+)-dependent inactivation (CDI) resulting from Ca(2+) influx and Ca(2+)-induced Ca(2+)-release (CICR) from the sarcoplasmic reticulum provided an automatic negative feedback mechanism to limit Ca(2+) entry and the contribution of I(CaL) to the cardiac action potential. Physiological modulation of I(CaL) by Beta-adrenergic and muscarinic agonists then involved essentially more or less of the same by enhancing or reducing Ca(2+) channel activity, Ca(2+) influx, sarcoplasmic reticulum load and thus CDI. Recent results on the other hand place VDI at the centre of the regulation of I(CaL). Under basal conditions it has been found that depolarization increases the probability that an ion channel will show rapid VDI. This is prevented by Beta-adrenergic stimulation. Evidence also suggests that a channel which shows rapid VDI inactivates before CDI can become effective. Therefore the contributions of VDI and CDI to the decay of I(CaL) are determined by the turning on, by depolarization, and the turning off, by phosphorylation, of the mechanism of rapid VDI. The physiological implications of these ideas are that under basal conditions the contribution of I(CaL) to the action potential will be determined largely by voltage and by Ca(2+) following Beta-adrenergic stimulation.

Figure 1. Inactivation of ICaL under basal conditions

A, normalized I_CaL carried by Ca²⁺ and Ba²⁺ in one ventricular myocyte. At negative voltages (left) Ca²⁺ current inactivates more rapidly than Ba²⁺ current. At positive voltages (right) the initial decay of Ba²⁺ current is as fast as that of Ca²⁺ current, only later does Ca²⁺ current show more inactivation. B, the contribution of Ca²⁺ to the inactivation of I_CaL evaluated by the ratio of decay recorded with Ca²⁺ relative to that recorded with Ba²⁺, 20 ms (R20, left) and 200 ms (R200, right) after activation. At both times the contribution of Ca²⁺ declines with depolarization though there is clearly more of an effect of Ca²⁺ at positive voltages after 200 ms than after 20 ms. The figure has been redrawn from Findlay (2002a).

Figure 2. The development of rapid VDI under basal conditions

A, the time course of the development of VDI was recorded in the absence of Ca²⁺, and in the absence of ion flux through the channels, between voltages which evoke minimal (–30 mV) and maximal (+10 mV) inactivation. The lines connecting data points represent the fitting of either a single (–30 and –20 mV) or a double (–10, 0 and +10 mV) exponential function to the data. B, the proportions of fast (open columns), slow (hatched columns) and no (filled columns) VDI which were recorded at different membrane potentials show that as the amount of current which showed fast VDI increased, that which showed no VDI declined. C, the relative contributions of Ca²⁺ and voltage to the decay of I_CaL were assessed by measuring the time course of development of inactivation at +10 mV in the presence (circles) and the absence (squares) of extracellular Ca²⁺. The first recorded total inactivation, being the sum of VDI and CDI. The second recorded only VDI since no ion flux through I_CaL occurred at this voltage under these conditions (see Findlay, 2002b for further details). It is clear that Ca²⁺ influx does little to increase the initial rapid phase of the development of inactivation. The results shown here were conducted in the presence of ryanodine. A and B have been redrawn from Findlay (2002d) and C from Findlay (2002b).

Figure 3. Inactivation of ICaL following β-adrenergic stimulation

A, normalized I_CaL carried by Ca²⁺ and Ba²⁺ in one ventricular myocyte. At negative (left) and positive (right) voltages Ca²⁺ current inactivates more rapidly than Ba²⁺ current. B, the contribution of Ca²⁺ to the inactivation of I_CaL 20 ms (R20, left) and 200 ms (R200, right) after activation is clearly sustained at all membrane voltages. The figure has been redrawn from Findlay (2002a).

Figure 4. Inhibition of rapid VDI by β-adrenergic stimulation

A, dose-dependent reduction of the availability–voltage relationship of I_CaL by Isoprenaline (Iso). Experiments were conducted in the absence of extracellular Ca²⁺ with a double-pulse voltage-clamp protocol with 1000 ms duration prepulse voltage steps. B, the effects of isoprenaline upon the amount of I_CaL which showed fast VDI (filled columns) and no inactivation (open columns). These experiments were conducted in the absence of extracellular Ca²⁺ and Findlay (2002c) should be consulted for further details. C, the relative contributions of Ca²⁺ and voltage to the decay of I_CaL were assessed as described in Fig. 2C in the presence (circles) and the absence (squares) of extracellular Ca²⁺ in the presence of 100 nm isoprenaline. It is clear that Ca²⁺ influx dramatically increases the rate of development of inactivation, and comparison with Fig. 2C reveals that this is due to the suppression of fast VDI. These experiments were conducted in the presence of ryanodine. A and B have been redrawn from Findlay (2002c) and C from Findlay (2002b).