On the transduction mechanism for muscarine-induced inhibition of M-current in cultured rat sympathetic neurones.
1. Dissociated adult or fetal rat superior cervical ganglion cells were voltage-clamped through a single patch pipette. The voltage-dependent K+ current, IM (M-current), was maintained by including MgATP in the pipette solution and by buffering the solution pH to 6.7. 2. Bath-applied muscarine (0.4 microM) produced a reversible inhibition of IM. 3. Addition of Gpp(NH)p (200 microM) or GTP-gamma-S (500 microM) to the pipette solution induced a slowly developing inhibition of IM and prevented recovery from subsequent muscarine-induced inhibition. 4. Addition of GDP-beta-S (500 microM) to the pipette solution reduced the amount of IM inhibition produced by 0.4 microM-muscarine by 42% and reduced the associated inward shift of the holding current by 56%. 5. Cells responded normally to muscarine after pre-treatment for 4-27 h with 500 ng ml-1 pertussis toxin (PTx). 6. IM was not diminished by extracellular addition of 1 mM-dibutyryl cyclic AMP, 8-bromo-cyclic AMP or dibutyryl cyclic GMP, or of 10 microM-forskolin. 7. IM was not reduced by inclusion of Li+ (2 mM) or inositol 1,4,5-trisphosphate (IP3, 100 microM) in the patch pipette, nor by ionophoretic injection of IP3 from an inserted micropipette. 8. Addition of 4-beta-phorbol 12,13-dibutyrate (PDBu, 0.5-2 microM) to the extracellular medium partly inhibited IM and reduced an additional component of resting membrane current. This effect was not replicated by 4-alpha-phorbol 12,13-didecanoate. 9. It is concluded that the inhibition of IM by muscarine is mediated through activation of a PTx-insensitive GTP-binding protein. The effect of muscarine appears not to be mediated by cyclic nucleotides or IP3 but may possibly involve the generation of diacylglycerols and activation of protein kinase C.
Brown
D A
DA
Department of Pharmacology, School of Pharmacy, University of London.
Marrion
N V
NV
Smart
T G
TG
D000818:Animals; D002478:Cells, Cultured; D005333:Fetus; D005728:Ganglia, Sympathetic / Q000166:cytology* / Q000196:embryology; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology; D009116:Muscarine / Q000494:pharmacology*; D009433:Neural Inhibition / Q000187:drug effects*; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects*; D051381:Rats; D011919:Rats, Inbred Strains; D013873:Thionucleotides / Q000494:pharmacology
D015221:Potassium Channels; D013873:Thionucleotides; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D015240:Phorbol 12,13-Dibutyrate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C023427:guanosine 5'-O-(2-thiodiphosphate); D009116:Muscarine; D015544:Inositol 1,4,5-Trisphosphate; D006160:Guanosine Triphosphate