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The Drosophila cac ts2 mutation reduces presynaptic Ca 2+ entry and defines an important element in Ca v 2.1 channel inactivation
Author(s) -
Macleod G. T.,
Chen L.,
Karunanithi S.,
Peloquin J. B.,
Atwood H. L.,
McRory J. E.,
Zamponi G. W.,
Charlton M. P.
Publication year - 2006
Publication title -
european journal of neuroscience
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.346
H-Index - 206
eISSN - 1460-9568
pISSN - 0953-816X
DOI - 10.1111/j.1460-9568.2006.04873.x
Subject(s) - neurotransmission , mutant , patch clamp , mutation , microbiology and biotechnology , biophysics , biology , voltage dependent calcium channel , ion channel , neurotransmitter , chemistry , neuroscience , electrophysiology , biochemistry , calcium , receptor , gene , organic chemistry
Voltage‐gated Ca 2+ channels in nerve terminals open in response to action potentials and admit Ca 2+ , the trigger for neurotransmitter release. The cacophony gene encodes the primary presynaptic voltage‐gated Ca 2+ channel in Drosophila motor‐nerve terminals. The cac ts2 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 cac ts2 mutation occurs in the cytoplasmic carboxyl tail of the α 1 ‐subunit, not within the pore‐forming trans‐membrane domains, making it difficult to predict the mutation's impact. We applied a Ca 2+ ‐imaging technique at motor‐nerve terminals of mutant larvae to test the hypothesis that the neurotransmission deficit is a result of impaired Ca 2+ entry. Presynaptic Ca 2+ 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 cac ts2 mutation plays a role in Ca 2+ channel activity. As the mutation occurs in a motif conserved in mammalian high‐voltage‐activated Ca 2+ channels, we used a heterologous expression system to probe the effect of this mutation on channel function. The mutation was introduced into rat Ca v 2.1 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 Ca v 2.1 channel inactivation.