Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Regulation of T-type calcium channels by Rho-associated kinase

Nature Neuroscience volume 10, pages 854–860 (2007)Cite this article

Abstract

We investigated the regulation of T-type channels by Rho-associated kinase (ROCK). Activation of ROCK via the endogenous ligand lysophosphatidic acid (LPA) reversibly inhibited the peak current amplitudes of rat Cav3.1 and Cav3.3 channels without affecting the voltage dependence of activation or inactivation, whereas Cav3.2 currents showed depolarizing shifts in these parameters. LPA-induced inhibition of Cav3.1 was dependent on intracellular GTP, and was antagonized by treatment with ROCK and RhoA inhibitors, LPA receptor antagonists or GDPßS. Site-directed mutagenesis of the Cav3.1 α1 subunit revealed that the ROCK-mediated effects involve two distinct phosphorylation consensus sites in the domain II-III linker. ROCK activation by LPA reduced native T-type currents in Y79 retinoblastoma and in lateral habenular neurons, and upregulated native Cav3.2 current in dorsal root ganglion neurons. Our data suggest that ROCK is an important regulator of T-type calcium channels, with potentially far-reaching implications for multiple cell functions modulated by LPA.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Effect of LPA on Cav3.1 calcium channels.
Figure 2: Involvement of the RhoA-ROCK pathway in Cav3.1 inhibition by LPA.
Figure 3: Channel structural determinants of LPA modulation of Cav3.1.
Figure 4: Effect of LPA on Cav3.3 and Cav3.2 channel activity.
Figure 5: LPA regulation of native T-type currents.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Bertolesi, G.E. et al. The Ca2+ channel antagonists mibefradil and pimozide inhibit cell growth via different cytotoxic mechanisms. Mol. Pharmacol. 62, 210–219 (2002).

    Article  CAS  Google Scholar 

  2. Latour, I. et al. Expression of T-type calcium channel splice variants in human glioma. Glia 48, 112–119 (2004).

    Article  Google Scholar 

  3. Panner, A. et al. Is there a role for T-type Ca2+ channel in glioma cell proliferation? Cell Calcium 37, 105–119 (2005).

    Article  CAS  Google Scholar 

  4. Perez-Reyes, E. Molecular physiology of low-voltage-activated T-type calcium channels. Physiol. Rev. 83, 117–161 (2003).

    Article  CAS  Google Scholar 

  5. Rodman, D.M. et al. Low-voltage–activated (T-type) calcium channels control proliferation of human pulmonary artery myocytes. Circ. Res. 96, 864–872 (2005).

    Article  CAS  Google Scholar 

  6. Yunker, A.M. & McEnery, M.W. Immunological characterization of T-type voltage-dependent calcium channel CaV3.1 (α1G) and CaV3.3 (α1I) isoforms reveal differences in their localization, expression and neural development. J. Bioenerg. Biomembr. 35, 533–575 (2003).

    Article  CAS  Google Scholar 

  7. Bourinet, E. et al. Silencing of the Cav3.2 T-type calcium channel gene in sensory neurons demonstrates its major role in nociception. EMBO J. 24, 315–324 (2005).

    Article  CAS  Google Scholar 

  8. Mangoni, M.E. et al. Bradycardia and slowing of the atrioventricular conduction in mice lacking CaV3.1/α1G T-type calcium channels. Circ. Res. 98, 1422–1430 (2006).

    Article  CAS  Google Scholar 

  9. Gu, Y. et al. Osteoblasts derived from load-bearing bones of the rat express both L- and T-like voltage-operated calcium channels and mRNA for α1C, α1D and α1G subunits. Pflugers Arch. 438, 553–560 (1999).

    CAS  PubMed  Google Scholar 

  10. McRory, J.E. et al. Molecular and functional characterization of a family of rat brain T-type calcium channels. J. Biol. Chem. 276, 3999–4011 (2001).

    Article  CAS  Google Scholar 

  11. Kim, D. et al. Lack of the burst firing of thalamocortical relay neurons and resistance to absence seizures in mice lacking α1G T-type Ca2+ channels. Neuron 31, 35–45 (2001).

    Article  CAS  Google Scholar 

  12. Chen, C.C. et al. Abnormal coronary function in mice deficient in α1H T-type Ca2+ channels. Science 302, 1416–1418 (2003).

    Article  CAS  Google Scholar 

  13. Moolenaar, W.H., Kranenburg, O., Postma, F.R. & Zondag, G.C. Lysophosphatidic acid: G protein signalling and cellular responses. Curr. Opin. Cell Biol. 9, 168–173 (1997).

    Article  CAS  Google Scholar 

  14. Moolenaar, W.H. Development of our current understanding of bioactive lysophospholipids. Ann. NY Acad. Sci. 905, 1–10 (2000).

    Article  CAS  Google Scholar 

  15. Contos, J.J. & Chun, J. The mouse lp(A3)/Edg7 lysophosphatidic acid receptor gene: genomic structure, chromosomal localization and expression pattern. Gene 267, 243–253 (2001).

    Article  CAS  Google Scholar 

  16. An, S., Bleu, T., Hallmark, O.G. & Goetzl, E.J. Characterization of a novel subtype of human G protein-coupled receptor for lysophosphatidic acid. J. Biol. Chem. 273, 7906–7910 (1998).

    Article  CAS  Google Scholar 

  17. Anliker, B. & Chun, J. Cell surface receptors in lysophospholipid signaling. J. Biol. Chem. 279, 20555–20558 (2004).

    Article  CAS  Google Scholar 

  18. Gardell, S.E., Dubin, A.E. & Chun, J. Emerging medicinal roles for lysophospholipid signaling. Trends Mol. Med. 12, 65–75 (2006).

    Article  CAS  Google Scholar 

  19. Lee, C-W., Rivera, R., Gardell, S., Dubin, A.E. & Chun, J. GPR92 as a new G12/13- and Gq-coupled lysophosphatidic acid receptor that increases cAMP, LPA5. J. Biol. Chem. 281, 23589–23597 (2006).

    Article  CAS  Google Scholar 

  20. Yanagida, K., Ishii, S., Hamano, F., Nogichi, K. & Shimizu, T. LPA4/p2y9/GPR23 mediates rho-dependent morphological changes in a rat neuronal cell line. J. Biol. Chem. 282, 5814–5824 (2007).

    Article  CAS  Google Scholar 

  21. Aznar, S. & Lacal, J.C. Rho signals to cell growth and apoptosis. Cancer Lett. 165, 1–10 (2001).

    Article  CAS  Google Scholar 

  22. Mueller, B.K., Mack, H. & Teusch, N. Rho kinase, a promising drug target for neurological disorders. Nat. Rev. Drug Discov. 4, 387–398 (2005).

    Article  CAS  Google Scholar 

  23. Riento, K. & Ridley, A.J. Rocks: multifunctional kinases in cell behaviour. Nat. Rev. Mol. Cell Biol. 4, 446–456 (2003).

    Article  CAS  Google Scholar 

  24. Shimokawa, H. & Takeshita, A. Rho-kinase is an important therapeutic target in cardiovascular medicine. Arterioscler. Thromb. Vasc. Biol. 25, 1767–1775 (2005).

    Article  CAS  Google Scholar 

  25. Dergham, P. et al. Rho signaling pathway targeted to promote spinal cord repair. J. Neurosci. 22, 6570–6577 (2002).

    Article  CAS  Google Scholar 

  26. Kawano, Y. et al. Phosphorylation of myosin-binding subunit (MBS) of myosin phosphatase by Rho-kinase in vivo. J. Cell Biol. 147, 1023–1038 (1999).

    Article  CAS  Google Scholar 

  27. Nakagawa, O. et al. ROCK-I and ROCK-II, two isoforms of Rho-associated coiled-coil forming protein serine/threonine kinase in mice. FEBS Lett. 392, 189–193 (1996).

    Article  CAS  Google Scholar 

  28. Matsui, T. et al. Rho-associated kinase, a novel serine/threonine kinase, as a putative target for small GTP binding protein Rho. EMBO J. 15, 2208–2216 (1996).

    Article  CAS  Google Scholar 

  29. Leung, T., Manser, E., Tan, L. & Lim, L. A novel serine/threonine kinase binding the Ras-related RhoA GTPase which translocates the kinase to peripheral membranes. J. Biol. Chem. 270, 29051–29054 (1995).

    Article  CAS  Google Scholar 

  30. Storey, N.M., O'Bryan, J.P. & Armstrong, D.L. Rac and Rho mediate opposing hormonal regulation of the ether-a-go-go-related potassium channel. Curr. Biol. 12, 27–33 (2002).

    Article  CAS  Google Scholar 

  31. Staruschenko, A. et al. Rho small GTPases activate the epithelial Na+ channel. J. Biol. Chem. 279, 49989–49994 (2004).

    Article  CAS  Google Scholar 

  32. Amano, M. et al. The COOH terminus of Rho-kinase negatively regulates rho-kinase activity. J. Biol. Chem. 274, 32418–32424 (1999).

    Article  CAS  Google Scholar 

  33. Shimizu, T. et al. Parallel coiled-coil association of the RhoA-binding domain in Rho-kinase. J. Biol. Chem. 278, 46046–46051 (2003).

    Article  CAS  Google Scholar 

  34. Wolfe, J.T., Wang, H., Howard, J., Garrison, J.C. & Barrett, P.Q. T-type calcium channel regulation by specific G-protein βγ subunits. Nature 424, 209–213 (2003).

    Article  CAS  Google Scholar 

  35. Welsby, P.J. et al. A mechanism for the direct regulation of T-type calcium channels by Ca2+/calmodulin-dependent kinase II. J. Neurosci. 23, 10116–10121 (2003).

    Article  CAS  Google Scholar 

  36. Park, J.Y. et al. Activation of protein kinase C augments T-type Ca2+ channel activity without changing channel surface density. J. Physiol. (Lond.) 577, 513–523 (2006).

    Article  CAS  Google Scholar 

  37. Hirooka, K. et al. T-Type calcium channel α1G and α1H subunits in human retinoblastoma cells and their loss after differentiation. J. Neurophysiol. 88, 196–205 (2002).

    Article  CAS  Google Scholar 

  38. Wilcox, K.S., Gutnick, M.J. & Christoph, G.R. Electrophysiological properties of neurons in the lateral habenula nucleus: an in vitro study. J. Neurophysiol. 59, 212–225 (1988).

    Article  CAS  Google Scholar 

  39. Huguenard, J.R., Gutnick, M.J. & Prince, D.A. Transient Ca2+ currents in neurons isolated from rat lateral habenula. J. Neurophysiol. 70, 158–166 (1993).

    Article  CAS  Google Scholar 

  40. McKay, B.E. et al. Ca(v)3 T-type calcium channel isoforms differentially distribute to somatic and dendritic compartments in rat central neurons. Eur. J. Neurosci. 24, 2581–2594 (2006).

    Article  Google Scholar 

  41. Todorovic, S.M., Jevtovic-Todorovic, V., Mennerick, S., Perez-Reyes, E. & Zorumski, C.F. Ca(v)3.2 channel is a molecular substrate for inhibition of T-type calcium currents in rat sensory neurons by nitrous oxide. Mol. Pharmacol. 60, 603–610 (2001).

    CAS  PubMed  Google Scholar 

  42. Davies, S.P., Reddy, H., Caivano, M. & Cohen, P. Specificity and mechanism of action of some commonly used protein kinase inhibitors. Biochem. J. 351, 95–105 (2000).

    Article  CAS  Google Scholar 

  43. Inoue, M. et al. Initiation of neuropathic pain requires lysophosphatidic acid receptor signaling. Nat. Med. 10, 712–718 (2004).

    Article  CAS  Google Scholar 

  44. Nelson, M.T., Joksovic, P.M., Perez-Reyes, E. & Todorovic, S.M. The endogenous redox agent L-cysteine induces T-type Ca2+ channel–dependent sensitization of a novel subpopulation of rat peripheral nociceptors. J. Neurosci. 25, 8766–8775 (2005).

    Article  CAS  Google Scholar 

  45. Ahmed, Z. et al. Disinhibition of neurotrophin-induced dorsal root ganglion cell neurite outgrowth on CNS myelin by siRNA-mediated knockdown of NgR, p75NTR and Rho-A. Mol. Cell. Neurosci. 28, 509–523 (2005).

    Article  CAS  Google Scholar 

  46. Arimura, N. et al. Phosphorylation of collapsin response mediator protein-2 by Rho-kinase. Evidence for two separate signaling pathways for growth cone collapse. J. Biol. Chem. 275, 23973–23980 (2000).

    Article  CAS  Google Scholar 

  47. Sah, V.P. et al. Cardiac-specific overexpression of RhoA results in sinus and atrioventricular nodal dysfunction and contractile failure. J. Clin. Invest. 103, 1627–1634 (1999).

    Article  CAS  Google Scholar 

  48. Hirdes, W., Horowitz, L.F. & Hille, B. Muscarinic modulation of erg potassium current. J. Physiol. (Lond.) 559, 67–84 (2004).

    Article  CAS  Google Scholar 

  49. Altier, C. et al. Opioid, cheating on its receptors, exacerbates pain. Nat. Neurosci. 9, 31–40 (2006).

    Article  CAS  Google Scholar 

  50. Molineux, M.L. et al. Specific T-type calcium channel isoforms are associated with distinct burst phenotypes in deep cerebellar nuclear neurons. Proc. Natl. Acad. Sci. USA 103, 5555–5560 (2006).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank T. Snutch and J. McRory for providing wild-type Cav3 cDNAs, and M. Molineux for early recording tests in vitro. This work was supported by grants from the Canadian Institutes for Health Research. R.W.T. and G.W.Z. are Scientists of the Alberta Heritage Foundation for Medical Research (AHFMR). G.W.Z. is a Canada Research Chair. D.V. holds postdoctoral fellowships from the AHFMR and the Heart and Stroke Foundation of Canada.

Author information

Authors and Affiliations

  1. Departments of Physiology & Biophysics, Hotchkiss Brain Institute, University of Calgary, 3330 Hospital Drive N.W., Calgary, T2N 4N1, Canada

    Mircea Iftinca, Jawed Hamid, Lina Chen, Diego Varela, Reza Tadayonnejad, Christophe Altier, Ray W Turner & Gerald W Zamponi

  2. Cell Biology & Anatomy, University of Calgary, 3330 Hospital Drive N.W., Calgary, T2N 4N1, Canada

    Mircea Iftinca, Jawed Hamid, Lina Chen, Diego Varela, Reza Tadayonnejad, Christophe Altier, Ray W Turner & Gerald W Zamponi

  3. Pharmacology & Therapeutics, University of Calgary, 3330 Hospital Drive N.W., Calgary, T2N 4N1, Canada

    Mircea Iftinca, Jawed Hamid, Lina Chen, Diego Varela, Christophe Altier & Gerald W Zamponi

Authors
  1. Mircea Iftinca
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jawed Hamid
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Lina Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Diego Varela
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Reza Tadayonnejad
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Christophe Altier
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ray W Turner
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Gerald W Zamponi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Gerald W Zamponi.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Expression of endogenous LPA receptors in tsA-201 cells as assayed by RT-PCR analysis (PDF 40 kb)

Supplementary Fig. 2

Analysis of Rho kinase phosphorylation sites on the Cav3.1 channel. (PDF 35 kb)

Supplementary Fig. 3

Average peak current amplitudes obtained from Cav1.2, Cav2.1 and Cav2.2 calcium channels under control conditions, and after 15-min incubation with LPA with or without pretreatment by Fasudil. (PDF 50 kb)

Supplementary Table 1

Lack of direct effects of inhibitors on half-activation potentials and peak current amplitude of rat Cav3.1 T-type calcium channels. (PDF 52 kb)

Supplementary Table 2

The effects of LPA (10 μM) and Fasudil (10 μM) on paramters of tonic spike discharge in LHb neurons in vitro (PDF 56 kb)

Supplementary Methods (PDF 77 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Iftinca, M., Hamid, J., Chen, L. et al. Regulation of T-type calcium channels by Rho-associated kinase. Nat Neurosci 10, 854–860 (2007). https://doi.org/10.1038/nn1921

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn1921

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing