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Department of Pharmacology, School of Pharmacy, University of London.

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.

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