31
766 ± 242 fS, with 2214 ± 986 (std., n=6) channels per rod (figure 3.3 C2).
A small activation delay, which is sometimes seen as an early minimum in HCN cur-
rents, is visible in figure 2.2 ЗА and 3B. It appears to be more evident in recordings of cell
attached HCN currents. This activation delay has been observed in HCN currents recorded
from Purkinje fibers [38], salamander rods [54], and expressed HCN channels [77]. One
explanation for this delay is that HCN channels have many different conformational states,
and must transition to a lower conducting state shortly after a voltage pulse before reaching
full activation.
While a larger current can be recorded from a whole cell as opposed to a cell-attached
patch, the cell-attached technique of recording currents has several advantages, including
less membrane noise, avoidance of cell internal dialysis, and the ability to localize the dis-
tribution of HCN currents on the cell membrane (see Methods). Therefore, to confirm our
whole-cell NSFA results, we also estimated the single channel conductance of HCN channels
with cell-attached patches, using special pipette solutions to increase the HCN driving force
(see Solutions in Materials Methods). The increased extracellular potassium level raised the
reversal potential of HCN channels without affecting the gating kinetics of ∕⅛ [108], increas-
ing the driving force of Ih currents. These cell attached patches in rods gave estimates of a
single channel conductance of 663 ± 71 fS, with 155 ± 42 (std., n=3) channels present in
each patch (figure 3.3 Cl). From the single channel conductance we estimate the number
of channels per cell by dividing the whole cell conductance (μ = 1.4 ± .89 nS) by the single
channel conductance, which predicts 2111 ± 1342 channels per cell. This result is similar