Abstract
Neurotransmitter release from mammalian sensory neurons is controlled by CaV2.2 N-type calcium channels. N-type channels are a major target of neurotransmitters and drugs that inhibit calcium entry, transmitter release and nociception through their specific G protein–coupled receptors. G protein–coupled receptor inhibition of these channels is typically voltage-dependent and mediated by Gβγ, whereas N-type channels in sensory neurons are sensitive to a second G protein–coupled receptor pathway that inhibits the channel independent of voltage. Here we show that preferential inclusion in nociceptors of exon 37a in rat Cacna1b (encoding CaV2.2) creates, de novo, a C-terminal module that mediates voltage-independent inhibition. This inhibitory pathway requires tyrosine kinase activation but not Gβγ. A tyrosine encoded within exon 37a constitutes a critical part of a molecular switch controlling N-type current density and G protein–mediated voltage-independent inhibition. Our data define the molecular origins of voltage-independent inhibition of N-type channels in the pain pathway.
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Acknowledgements
We are grateful to S. Denome for technical assistance, K. Dunlap (Tufts University) for GABABR1a and GABABR2 cDNA clones, L. Devi (New York University) for the μ-opioid receptor cDNA clone and S.R. Ikeda (US National Institutes of Health) for the MAS-GRK-ct cDNA clone. This work was supported by US National Institutes of Health grants NS29967 and NS55251 (D.L.).
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All authors contributed to writing the manuscript. D.L. directed the project. J.R. performed experiments and analyses for all figures. A.J.C. originally identified the tyrosine kinase sites in e37a, contributed to the design and construction of the tyrosine mutants, and performed the sequence analysis in Figure 8a.
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Supplementary information
Supplementary Fig. 1
Time course of μ-opioid receptor? and GABAB receptor–mediated inhibition of CaV2.2 currents. (PDF 18 kb)
Supplementary Fig. 2
Voltage-independent but not voltage-dependent inhibition mediated by GABAB receptor activation is prevented by pp60c-src tyrosine kinase peptide inhibitor. (PDF 47 kb)
Supplementary Table 1
Averaged parameters estimated from Boltzmann linear fits of current-voltage relationships. (PDF 55 kb)
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Raingo, J., Castiglioni, A. & Lipscombe, D. Alternative splicing controls G protein–dependent inhibition of N-type calcium channels in nociceptors. Nat Neurosci 10, 285–292 (2007). https://doi.org/10.1038/nn1848
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DOI: https://doi.org/10.1038/nn1848
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