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

GT Macleod, L Chen, S. Karunanithi, JB Peloquin, HL Atwood, JE McRory, GW Zamponi, MP Charlton

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Abstract

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 1-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.
Original languageEnglish
Pages (from-to)3230-3244
Number of pages15
JournalEuropean Journal of Neuroscience
Volume23
Issue number12
DOIs
Publication statusPublished - Jun 2006

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    Macleod, GT., Chen, L., Karunanithi, S., Peloquin, JB., Atwood, HL., McRory, JE., Zamponi, GW., & Charlton, MP. (2006). The Drosophila cacts2 mutation reduces presynaptic Ca2+ entry and defines an important element in Cav2.1 channel inactivation. European Journal of Neuroscience, 23(12), 3230-3244. https://doi.org/10.1111/j.1460-9568.2006.04873.x