Abstract
Neurokinin (NK) 1 receptors and CaV2.3 calcium channels are both expressed in nociceptive neurons, and mice lacking either protein display altered responses to noxious stimuli. Here, we examined modulation of CaV2.3 through NK1 receptors expressed in human embryonic kidney 293 cells. We find that NK1 receptors generate complex modulation of CaV2.3. In particular, weak activation of these receptors evokes mainly stimulation of CaV2.3, whereas strong receptor activation elicits profound inhibition that overlaps with channel stimulation. Unlike R-type channels encoded by CaV2.3, L-type (CaV1.3), N-type (CaV2.2), and P/Q-type (CaV2.1) channels are inhibited, but not stimulated, through NK1 receptors. Pharmacological experiments show that protein kinase C (PKC) mediates stimulation of CaV2.3 through NK1 receptors. The signaling mechanisms underlying inhibition were explored by expressing proteins that buffer either Gαq/11 (regulator of G protein signaling protein 3T and carboxyl-terminal region of phospholipase C-β1) or Gβγ subunits (transducin and the carboxyl-terminal region of bovine G-protein-coupled receptor kinase). A fast component of inhibition was attenuated by buffering Gβγ, whereas a slow component of inhibition was reduced by buffering Gαq/11. When both Gβγ and Gαq/11 were simultaneously buffered in the same cells, inhibition was virtually eliminated, but receptor activation still triggered substantial stimulation of CaV2.3. We also report that NK1 receptors accelerate the inactivation kinetics of CaV2.3 currents. Altogether, our results indicate that NK1 receptors modulate CaV2.3 using three different signaling mechanisms: a fast inhibition mediated by Gβγ, a slow inhibition mediated by Gαq/11, and a slow stimulation mediated by PKC. This new information concerning R-type calcium channels and NK1 receptors may help in understanding nociception, synaptic plasticity, and other physiological processes.
Footnotes
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This work was supported by Consejo Nacional de Ciencia y Tecnologia grant 39865-Q and UASLP grant C06-FAI-03-4.7 (to U.M.) and by Muscular Dystrophy Association grant MDA3663 and the Utah Agricultural Experimental Station Project UTA00581 (to B.A.A.).
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Portions of this work were previously presented at the 2003 and 2006 annual meetings of the Biophysical Society (Biophys J Suppl84:537a; Biophys J Suppl90:396a).
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U.M. and A.T. contributed equally to this work.
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ABBREVIATIONS: NK, neurokinin; HEK, human embryonic kidney; PKC, protein kinase C; EGFP, enhanced green fluorescent protein; MAS-GRK3ct, carboxyl-terminal region of G protein-coupled receptor kinase 3; Gαt, rod transducin; RGS3T, regulator of G protein signaling protein 3T; RGS2, regulator of G protein signaling protein 2; PLCβ1ct, carboxyl-terminal region (T903-L1216) of phospholipase C-β1; C, cell capacitance; RS, series access resistance; NKA, neurokinin A; PTX, pertussis toxin; CTX, cholera toxin; Bis, bisindolylmaleimide; I-V, current-voltage; PP2, 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine.
- Received July 5, 2006.
- Accepted October 18, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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