Neuronal voltage-activated calcium channels: on the roles of the alpha 1E and beta 3 subunits.

Many neurons of the central and peripheral nervous systems display multiple high voltage-activated (HVA) Ca2+ 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 alpha 1, subunits, in association with various beta subunits. However, there are large gaps in understanding how individual subunits contribute to Ca2+ channel diversity. Here we describe experiments to investigate the roles of alpha 1E and beta 3 subunits in mammalian neurons. The alpha 1E subunit is the leading candidate to account for the R-type channel, the least understood of the various types of high voltage-activated Ca2+ channels. Incubation with alpha 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 alpha 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 alpha 1 subunits support the high voltage-activated R-type current in cerebellar granule cells. We studied the role of the Ca2+ channel beta 3 subunit using a gene targeting strategy. In sympathetic beta 3-/- neurons, the L-type current was significantly reduced relative to wild type (wt). In addition, N-type Ca2+ channels made up a smaller proportion of the total Ca2+ 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 Ca2+ channels was described by two Boltzmann components with different voltage dependence. The absence of the beta 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 beta 3 subunit affects at least three distinct types of HVA Ca2+ channel, but no single type of channel is solely dependent on beta 3.