Functional impact of alternative splicing of human T-type Cav3.3 calcium channels

Low-voltage-activated T-type (Cav3) Ca2+ channels produce low-threshold spikes that trigger burst firing in many neurons. The CACNA1I gene encodes the Cav3.3 isoform, which activates and inactivates much more slowly than the other Cav3 channels. These distinctive kinetic features, along with its brain-region-specific expression, suggest that Cav3.3 channels endow neurons with the ability to generate long-lasting bursts of firing. The human CACNA1I gene contains two regions of alternative splicing: variable inclusion of exon 9 and an alternative acceptor site within exon 33, which leads to deletion of 13 amino acids (Delta33). The goal of this study is to determine the functional consequences of these variations in the full-length channel. The cDNA encoding these regions were cloned using RT-PCR from human brain, and currents were recorded by whole cell patch clamp. Introduction of the Delta33 deletion slowed the rate of channel opening. Addition of exon 9 had little effect on kinetics, whereas its addition to Delta33 channels unexpectedly slowed both activation and inactivation kinetics. Modeling of neuronal firing showed that exon 9 or Delta33 alone reduced burst firing, whereas the combination enhanced firing. The major conclusions of this study are that the intracellular regions after repeats I and IV play a role in channel gating, that their effects are interdependent, suggesting a direct interaction, and that splice variation of Cav3.3 channels provides a mechanism for fine-tuning the latency and duration of low-threshold spikes.