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. 1998 May 15;18(10):3521-8.
doi: 10.1523/JNEUROSCI.18-10-03521.1998.

Downregulation of transient K+ channels in dendrites of hippocampal CA1 pyramidal neurons by activation of PKA and PKC

D A Hoffman  1 , D Johnston
Affiliations

Downregulation of transient K+ channels in dendrites of hippocampal CA1 pyramidal neurons by activation of PKA and PKC

D A Hoffman et al. J Neurosci. .

Abstract

We have reported recently a high density of transient A-type K+ channels located in the distal dendrites of CA1 hippocampal pyramidal neurons and shown that these channels shape EPSPs, limit the back-propagation of action potentials, and prevent dendritic action potential initiation (). Because of the importance of these channels in dendritic signal propagation, their modulation by protein kinases would be of significant interest. We investigated the effects of activators of cAMP-dependent protein kinase (PKA) and the Ca2+-dependent phospholipid-sensitive protein kinase (PKC) on K+ channels in cell-attached patches from the distal dendrites of hippocampal CA1 pyramidal neurons. Inclusion of the membrane-permeant PKA activators 8-bromo-cAMP (8-br-cAMP) or forskolin in the dendritic patch pipette resulted in a depolarizing shift in the activation curve for the transient channels of approximately 15 mV. Activation of PKC by either of two phorbol esters also resulted in a 15 mV depolarizing shift of the activation curve. Neither PKA nor PKC activation affected the sustained or slowly inactivating component of the total outward current. This downregulation of transient K+ channels in the distal dendrites may be responsible for some of the frequently reported increases in cell excitability found after PKA and PKC activation. In support of this hypothesis, we found that activation of either PKA or PKC significantly increased the amplitude of back-propagating action potentials in distal dendrites.

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Figures

Fig. 1.
Fig. 1.
Procedure for isolating the transient current in cell-attached patches. A, The total outward current from a dendritic patch located 320 μm from the soma for a 150 msec voltage step from −89 to +51 mV is shown. Voltage protocol is shownabove the trace. Here and in subsequent figures, traces are ensemble averages of 10–30 sweeps.B, A 50 msec prepulse (VPP) to −29 mV results in transient channel inactivation, leaving only the sustained component of the total outward current. C, Digital subtraction of the current remaining in B from that in A results in the isolated transient component shown.
Fig. 2.
Fig. 2.
Downregulation of transient channel activation by PKA activation. A, Representative example of isolated transient currents for the control condition and for that with 100 μm 8-br-cAMP included in the patch pipette, scaled to demonstrate their fraction of maximal conductance, for a step from −85 to −5 mV. Under control conditions, nearly 50% of the available channels are activated by this voltage step. With PKA activation, only ∼25% are activated. In the same patch, subsequent H7 application returned the fraction of channels activated back toward control levels.B, Steady-state activation and inactivation curves for control and 8-br-cAMP conditions and an activation curve for the condition with 8-br-cAMP in the pipette after bath application of 300 μm H7. Half-activation with 8-br-cAMP (V1/2 = +11 mV; n = 12) was shifted 13 mV to theright of control levels (V1/2 = −2 mV;n = 12). There was also a small (6 mV) shift in the inactivation curve with 8-br-cAMP (V1/2 = −50 mV and n = 8 vs V1/2 = −56 mV andn = 5 for controls). Bath application of H7 shifted the 8-br-cAMP curve back toward control levels (V1/2= +3.5 mV after 30 min in H7; n = 3).Inset, Time course of reversal of PKA effect by H7 application (top horizontal bar). The activation and inactivation curves under control conditions are nearly identical to those reported previously for transient K+ channels in distal dendrites (Hoffman et al., 1997). Curves (here and see Figs.3, 5, 6) are least-square fits of the data to Boltzmann functions (see Materials and Methods).
Fig. 3.
Fig. 3.
Downregulation of transient channel activation by PKC activation. A, Representative example of isolated transient currents for the control condition and for that with 10 μm PDA included in the patch pipette, scaled to demonstrate their fraction of maximal conductance, for a step from −85 to −5 mV. B, Steady-state activation and inactivation curves for control and two different phorbol ester conditions. Half-activation with PDA (V1/2 = +13 mV;n = 7) or PDBu (V1/2 = +12 mV;n = 4) was shifted ∼15 mV to theright of controls (V1/2 = −2 mV;n = 12). There was also a small (6 mV) shift in the inactivation curve with PDA (V1/2 = −50 mV andn = 6 vs V1/2 = −56 mV andn = 5 for controls). There was, however, no activation curve shift when the inactive phorbol ester 4-α-phorbol was included in the pipette (V1/2 = −2 mV;n = 8).
Fig. 4.
Fig. 4.
Both 8-br-cAMP and PDA alter the voltage dependency of inactivation for the transient channels. Time constants for inactivation are plotted versus membrane potential for control, 8-br-cAMP, and PDA. Time constants for control and drug conditions were similar for the smallest depolarization measured (−5 mV) but were significantly different at higher potentials, resulting in a shallower slope of the lines fit to the data. Slope equals 2.5 msec/10 mV for control and 1.9 msec/10 mV for both 8-br-cAMP and PDA conditions. Inset, Two traces, scaled to the same amplitude, for a step to +55 mV under control (bottom trace) and 8-br-cAMP (top trace) conditions fit by a single exponential (τ, 30.7 and 20.6 msec for control and 8-br-cAMP, respectively).
Fig. 5.
Fig. 5.
The effects of PKA and PKC activation are not additive. A, Representative example of isolated transient currents for the control condition and for that with 100 μm 8-br-cAMP plus 10 μm PDA included in the patch pipette, scaled to demonstrate their fraction of maximal conductance, for a step from −85 to −5 mV. B, Steady-state activation and inactivation curves for control and 8-br-cAMP plus PDA. Half-activation with 8-br-cAMP plus PDA (V1/2 = +8 mV; n = 5) was shifted 10 mV to the right of control levels (V1/2 = −2 mV; n = 7). There was also a small (5 mV) shift in the inactivation curve with 8-br-cAMP plus PDA (V1/2 = −51 mV and n = 3 vs V1/2 = −56 mV andn = 5 for controls).
Fig. 6.
Fig. 6.
The sustained current was unaffected by either 8-br-cAMP or PDA. Steady-state activation curves for the sustained component are plotted for control, 8-br-cAMP, and PDA. Half-activation with 8-br-cAMP (V1/2 = +7 mV; k = 10.5;n = 6) and PDA (V1/2 = +5 mV;k = 8.5; n = 8) was similar to that with controls (V1/2 = +2 mV;k = 8.5; n = 10). Note that the difference in V1/2 between control and 8-br-cAMP conditions was primarily caused by the small difference in fitted slope, although this slope change was not significant.
Fig. 7.
Fig. 7.
PKA and PKC activation increases the back-propagating action potential amplitude in the distal dendrites.A, Antidromically initiated action potentials before (pre) and after (+PDA) bath application of 10 μm PDA. In this recording 240 μm from the soma, application of PDA lead to a 62% increase in action potential amplitude (from 44 to 71 mV). B, Antidromically initiated action potentials before (pre) and after (post and +8-br-cAMP) bath application of 100 μm8-br-cAMP. Left, In a more proximal recording (150 μm from the soma), in which A-channel density is smaller, action potential amplitude was large to begin with, and 8-br-cAMP did not lead to an increase in amplitude. Right, In this distal recording (280 μm from the soma), in which action potential amplitude is attenuated because of high A-channel density, PKA activation increased the amplitude over 40% (from 33 to 47 mV). C, Summary data. Percent change in action potential amplitude and maximal rate of rise are plotted for five conditions: distal recordings after PDA application, proximal (150 μm) recordings after 8-br-cAMP application, mid (180–200 μm) recordings after 8-br-cAMP application, distal (250–320 μm) recordings after 8-br-cAMP application, and distal and mid recordings after 8-br-cAMP application with 300 μm H7 included in the external solution. Thenumber of cells for each group is inparentheses. Asterisks denote a significant percent increase over the controls (see Materials and Methods).
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