The Ca2α1 EF-hand F helix tyrosine, a highly conserved locus for GPCR inhibition of Ca2 channels.

The sensory neuron of Aplysia californica participates in several forms of presynaptic plasticity including homosynaptic depression, heterosynaptic depression, facilitation and the reversal of depression. The calcium channel triggering neurotransmitter release at most synapses is Ca2, consisting of the pore forming α1 subunit (Ca2α1), and auxiliary Caβ, and Caα2δ subunits. To determine the role of the Ca2 channel in presynaptic plasticity in Aplysia, we cloned Aplysia Ca2α1, Caβ, and Caα2δ and over-expressed the proteins in Aplysia sensory neurons (SN). We show expression of exogenous Ca2α1 in the neurites of cultured Aplysia SN. One proposed mechanism for heterosynaptic depression in Aplysia is through inhibition of Ca2. Here, we demonstrate that heterosynaptic depression of the Ca2 calcium current is inhibited when a channel with a Y-F mutation at the conserved Src phosphorylation site is expressed, showing the strong conservation of this mechanism over evolution. We also show that the Y-F mutation reduces heterosynaptic inhibition of neurotransmitter release, highlighting the physiological importance of this mechanism for the regulation of synaptic efficacy. These results also demonstrate our ability to replace endogenous Ca2 channels with recombinant channels allowing future examination of the structure function relationship of Ca2 in the regulation of transmitter release in this system.