The Drosophila cacts2 mutation reduces presynaptic Ca2+ entry and defines an important element in Cav2.1 channel inactivation

Voltage‐gated Ca2+ channels in nerve terminals open in response to action potentials and admit Ca2+, the trigger for neurotransmitter release. The cacophony gene encodes the primary presynaptic voltage‐gated Ca2+ channel in Drosophila motor‐nerve terminals. The cacts2 mutant allele of cacophony is associated with paralysis and reduced neurotransmission at non‐permissive temperatures but the basis for the neurotransmission deficit has not been established. The cacts2 mutation occurs in the cytoplasmic carboxyl tail of the α1‐subunit, not within the pore‐forming trans‐membrane domains, making it difficult to predict the mutation's impact. We applied a Ca2+‐imaging technique at motor‐nerve terminals of mutant larvae to test the hypothesis that the neurotransmission deficit is a result of impaired Ca2+ entry. Presynaptic Ca2+ signals evoked by single and multiple action potentials showed a temperature‐dependent reduction. The amplitude of the reduction was sufficient to account for the neurotransmission deficit, indicating that the site of the cacts2 mutation plays a role in Ca2+ channel activity. As the mutation occurs in a motif conserved in mammalian high‐voltage‐activated Ca2+ channels, we used a heterologous expression system to probe the effect of this mutation on channel function. The mutation was introduced into rat Cav2.1 channels expressed in human embryonic kidney cells. Patch‐clamp analysis of mutant channels at the physiological temperature of 37 °C showed much faster inactivation rates than for wild‐type channels, demonstrating that the integrity of this motif is critical for normal Cav2.1 channel inactivation.