Many neurons of the central and peripheral nervous systems display
multiple high voltage-activated (HVA) Ca
2+ currents, often classified
as L-, N-, P-, Q, and R-type. The heterogeneous properties of
these channels have been attributed to diversity in their pore-forming
1, subunits, in association with various ß subunits.
However, there are large gaps in understanding how individual
subunits contribute to Ca
2+ channel diversity. Here we describe
experiments to investigate the roles of
1E and ß
3 subunits in mammalian neurons. The
1E subunit is the leading
candidate to account for the R-type channel, the least understood
of the various types of high voltage-activated Ca
2+ channels.
Incubation with
1E antisense oligonucleotide caused a 53% decrease
in the peak R-type current density, while no significant changes
in the current expression were seen in sense oligonucleotide-treated
cells. The specificity of the
1E antisense oligonucleotides
was supported by the lack of change in the amplitude of P/Q
current. These results upheld the hypothesis that members of
the E class of
1 subunits support the high voltage-activated
R-type current in cerebellar granule cells. We studied the role
of the Ca
2+ channel ß
3 subunit using a gene targeting
strategy. In sympathetic ß
3-/- neurons, the L-type
current was significantly reduced relative to wild type (wt).
In addition, N-type Ca
2+ channels made up a smaller proportion
of the total Ca
2+ current than in wt due to a lower N-type current
density in a group of neurons with small total currents. Voltage-dependent
activation of P/Q-type Ca
2+ channels was described by two Boltzmann
components with different voltage dependence. The absence of
the ß
3 subunit was associated with a shift in the
more depolarized component of the activation along the voltage
axis toward more negative potentials. The overall conclusion
is that deletion of the ß
3 subunit affects at least
three distinct types of HVA Ca
2+ channel, but no single type
of channel is solely dependent on ß
3.
1Eand ß3 Subunits
