Alternative Splicing of a Short Cassette Exon in α1BGenerates Functionally Distinct N-Type Calcium Channels in Central and Peripheral Neurons

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Abstract
The N-type Ca channel α1Bsubunit is localized to synapses throughout the nervous system and couples excitation to release of neurotransmitters. In a previous study, two functionally distinct variants of the α1Bsubunit were identified, rnα1B-band rnα1B-d, that differ at two loci;four amino acids [SerPheMetGly (SFMG)] in IIIS3–S4 and two amino acids [GluThr (ET)] in IVS3–S4. These variants are reciprocally expressed in rat brain and sympathetic ganglia (Lin et al., 1997a). We now show that the slower activation kinetics of rnα1B-b(ΔSFMG/+ET) compared with rnα1B-d(+SFMG/ΔET) channels are fully accounted for by the insertion of ET in IVS3–S4 and not by the lack of SFMG in IIIS3–S4. We also show that the inactivation kinetics of these two variants are indistinguishable. Through genomic analysis we identify a six-base cassette exon that encodes the ET site and with ribonuclease protection assays demonstrate that the expression of this mini-exon is essentially restricted to α1BRNAs of peripheral neurons. We also show evidence for regulated alternative splicing of a six-base exon encoding NP in the IVS3–S4 linker of the closely related α1Agene and establish that residues NP can functionally substitute for ET in domain IVS3–S4 of α1B. The selective expression of functionally distinct Ca channel splice variants of α1Band α1Asubunits in different regions of the nervous system adds a new dimension of diversity to voltage-dependent Ca signaling in neurons that may be important for optimizing action potential-dependent transmitter release at different synapses.