PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion

  • Li Li (李 丽)
  • Lei Zhao (赵 磊)
  • Yang Wang (王 洋)
  • Ke-tao Ma (马克涛)
  • Wen-yan Shi (石文艳)
  • Ying-zi Wang (王英姿)
  • Jun-qiang Si (司军强)


The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1–1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1–1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001–1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca2+-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the “pre-synaptic inhibition” evoked by GABA, which may explain its role in pain and neurogenic inflammation.

Key words

peripheral nervous system substance P GABAA receptor protein kinase C dorsal root ganglion 


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  1. 1.
    Eccles JC. Presynaptic inhibition in the spinal cord. Progress Brain Res, 1964,12:65–91CrossRefGoogle Scholar
  2. 2.
    Chen QX, Stelzer A, Kay AR, et al. GABAA receptor function is regulated by phosphorylation in acutely dissociated guinea-pig hippocampal neurons. J Physiol, 1990,420:207–221CrossRefPubMedCentralPubMedGoogle Scholar
  3. 3.
    McMahon SB, Koltzenburg M. Novel classes of nociceptors: beyond Sherrington. Trends Neurosci, 1990,13(6): 199–201CrossRefPubMedGoogle Scholar
  4. 4.
    Si JQ, Li ZW, Hu HZ, et al. Inhibitory effect of baclofen on GABA-induced depolarization and GABA-activated current in primary sensory neurons. Neuroscience, 1997,81(3):821–827CrossRefPubMedGoogle Scholar
  5. 5.
    Si JQ, Zhang ZQ, Li CX, et al. Modulatory effect of substance P on GABA-activated currents from rat dorsal root ganglion. Acta Pharmacol Sin, 2004,25(5):623–629PubMedGoogle Scholar
  6. 6.
    Yamada K, Akasu T. Substance P suppresses GABAA receptor function via protein kinase C in primary sensory neurons of bullfrogs. J Physiol, 1996,496(Pt 2):439–449CrossRefPubMedCentralPubMedGoogle Scholar
  7. 7.
    Gyenes M, Wang Q, Gibbs TT, et al. Phosphorylation factors control neurotransmitter and neuromodulator actions at the gamma-aminobutyric acid type A receptor. Molecul Pharmacol, 1994,46(3):542–549Google Scholar
  8. 8.
    Velazquez KT, Mohammad H, Sweitzer SM. Protein kinase C in pain: involvement of multiple isoforms. Pharmacol Res, 2007,55(6):578–589CrossRefPubMedCentralPubMedGoogle Scholar
  9. 9.
    Hashimoto T, Ase K, Sawamura S, et al. Postnatal development of a brain-specific subspecies of protein kinase C in rat. J Neurosci, 1988,8(5):1678–1683PubMedGoogle Scholar
  10. 10.
    Huang FL, Young WS, 3rd, Yoshida Y, et al. Developmental expression of protein kinase C isozymes in rat cerebellum. Brain Res Dev Brain Res, 1990,52(1–2):121–130CrossRefPubMedGoogle Scholar
  11. 11.
    Mochly-Rosen D, Basbaum AI, Koshland DE, et al. Distinct cellular and regional localization of immunoreactive protein kinase C in rat brain. Proc Natl Acad Sci USA, 1987,84(13):4660–4664CrossRefPubMedCentralPubMedGoogle Scholar
  12. 12.
    Saito Y, Teshima R, Takagi K, et al. Activation of protein kinase C alpha enhances human growth hormone-binding protein release. Mol Cell Endocrinol, 1998,146(1–2):197–205CrossRefPubMedGoogle Scholar
  13. 13.
    Wetsel WC, Khan WA, Merchenthaler I, et al. Tissue and cellular distribution of the extended family of protein kinase C isoenzymes. J Cell Biol, 1992,117(1):121–133CrossRefPubMedGoogle Scholar
  14. 14.
    Aley KO, Messing RO, Mochly-Rosen D, et al. Chronic hypersensitivity for inflammatory nociceptor sensitization mediated by the epsilon isozyme of protein kinase C. J Neurosci, 2000,20(12):4680–4685PubMedGoogle Scholar
  15. 15.
    Igwe OJ, Chronwall BM. Hyperalgesia induced by peripheral inflammation is mediated by protein kinase C betaII isozyme in the rat spinal cord. Neuroscience, 2001,104(3):875–890CrossRefPubMedGoogle Scholar
  16. 16.
    Ma KT, Si JQ, Zhang ZQ, et al. Modulatory effect of CCK-8S on GABA-induced depolarization from rat dorsal root ganglion. Brain Res, 2006,1121(1):66–75CrossRefPubMedGoogle Scholar
  17. 17.
    Baker MD. Protein kinase C mediates up-regulation of tetrodotoxin-resistant, persistent Na+ current in rat and mouse sensory neurones. J Physiol, 2005,567(Pt 3):851–867CrossRefPubMedCentralPubMedGoogle Scholar
  18. 18.
    Yang L, Liu G, Zakharov SI, et al. Ser1928 is a common site for Cav1.2 phosphorylation by protein kinase C isoforms. J Biol Chem, 2005,280(1):207–214CrossRefPubMedGoogle Scholar
  19. 19.
    Otsuka M, Yoshioka K. Neurotransmitter functions of mammalian tachykinins. Physiol Rev, 1993,73(2):229–308PubMedGoogle Scholar
  20. 20.
    Akasu T, Ishimatsu M, Yamada K. Tachykinins cause inward current through NK1 receptors in bullfrog sensory neurons. Brain Res, 1996,713(1–2):160–167CrossRefPubMedGoogle Scholar
  21. 21.
    Dray A, Pinnock RD. Effects of substance P on adult rat sensory ganglion neurones in vitro. Neurosci Lett, 1982,33(1):61–66CrossRefPubMedGoogle Scholar
  22. 22.
    Hu HZ, Li ZW, Si JQ. Evidence for the existence of substance P autoreceptor in the membrane of rat dorsal root ganglion neurons. Neuroscience, 1997,77(2):535–541CrossRefPubMedGoogle Scholar
  23. 23.
    Ishimatsu M. Substance P produces an inward current by suppressing voltage-dependent and -independent K+ currents in bullfrog primary afferent neurons. Neurosci Res, 1994,19(1):9–20CrossRefPubMedGoogle Scholar
  24. 24.
    Si JQ, Li ZW. Effect of substance P on the somatic membrane of rat DRG neurons. Sheng Li Xue Bao (Chinese), 1996,48(1):8–14Google Scholar
  25. 25.
    Desarmenien M, Feltz P, Occhipinti G, et al. Coexistence of GABAA and GABAB receptors on A delta and C primary afferents. Br J Pharmacol, 1984,81(2):327–333CrossRefPubMedCentralPubMedGoogle Scholar
  26. 26.
    Zhou Y, Zhou ZS, Zhao ZQ. PKC regulates capsaicin-induced currents of dorsal root ganglion neurons in rats. Neuropharmacology, 2001,41(5):601–608CrossRefPubMedGoogle Scholar
  27. 27.
    Frayer SM, Barber LA, Vasko MR. Activation of protein kinase C enhances peptide release from rat spinal cord slices. Neurosci Lett, 1999,265(1):17–20CrossRefPubMedGoogle Scholar
  28. 28.
    Barber LA, Vasko MR. Activation of protein kinase C augments peptide release from rat sensory neurons. J Neurochem, 1996,67(1):72–80CrossRefPubMedGoogle Scholar
  29. 29.
    Malcangio M, Fernandes K, Tomlinson DR. NMDA receptor activation modulates evoked release of substance P from rat spinal cord. Br J Pharmacol, 1998,125(8): 1625–1626CrossRefPubMedCentralPubMedGoogle Scholar
  30. 30.
    Qin X, Wan Y, Wang X. CCL2 and CXCL1 trigger calcitonin gene-related peptide release by exciting primary nociceptive neurons. J Neurosci Res, 2005,82(1):51–62CrossRefPubMedGoogle Scholar
  31. 31.
    Cheng HJ, Ma KT, Li L, et al. Differential expression of alpha-adrenoceptor subtypes in rat dorsal root ganglion after chronic constriction injury. J Huazhong Univ Sci Technolog Med Sci, 2014,34(3):322–329CrossRefPubMedGoogle Scholar
  32. 32.
    McCarthy PW, Lawson SN. Cell type and conduction velocity of rat primary sensory neurons with substance P-like immunoreactivity. Neuroscience, 1989,28(3):745–753CrossRefPubMedGoogle Scholar
  33. 33.
    Hershey AD, Krause JE. Molecular characterization of a functional cDNA encoding the rat substance P receptor. Science, 1990,247(4945):958–962CrossRefPubMedGoogle Scholar
  34. 34.
    Huang RQ, Dillon GH. Maintenance of recombinant type A gamma-aminobutyric acid receptor function: role of protein tyrosine phosphorylation and calcineurin. J Pharmacol Exp Ther, 1998,286(1):243–255PubMedGoogle Scholar
  35. 35.
    Masu Y, Nakayama K, Tamaki H, et al. cDNA cloning of bovine substance-K receptor through oocyte expression system. Nature, 1987,329(6142): 836–838CrossRefPubMedGoogle Scholar
  36. 36.
    Nicoll RA, Schenker C, Leeman SE. Substance P as a transmitter candidate. Annu Rev Neurosci, 1980,3:227–268CrossRefPubMedGoogle Scholar
  37. 37.
    Lecci A, Maggi CA. Peripheral tachykinin receptors as potential therapeutic targets in visceral diseases. Expert Opin Ther Targets, 2003,7(3):343–362CrossRefPubMedGoogle Scholar
  38. 38.
    Maggi CA. Tachykinins as peripheral modulators of primary afferent nerves and visceral sensitivity. Pharmacol Res, 1997,36(2):153–169CrossRefPubMedGoogle Scholar
  39. 39.
    Quartara L, Altamura M, Evangelista S, et al. Tachykinin receptor antagonists in clinical trials. Expert Opin Investig Drugs, 2009,18(12):1843–1864CrossRefPubMedGoogle Scholar

Copyright information

© Huazhong University of Science and Technology and Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Li Li (李 丽)
    • 1
    • 2
  • Lei Zhao (赵 磊)
    • 1
  • Yang Wang (王 洋)
    • 1
    • 2
    • 3
  • Ke-tao Ma (马克涛)
    • 1
    • 2
  • Wen-yan Shi (石文艳)
    • 1
    • 2
  • Ying-zi Wang (王英姿)
    • 2
  • Jun-qiang Si (司军强)
    • 1
    • 2
    • 3
    • 4
  1. 1.Department of PhysiologyMedical College of Shihezi UniversityShiheziChina
  2. 2.Key Laboratory of Xinjiang Endemic and Ethnic DiseaseShihezi University School of MedicineShiheziChina
  3. 3.Basic Medical School of Wuhan UniversityWuhanChina
  4. 4.Basic Medical School, Tongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina

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