G‐proteins and G‐protein subunits mediating cholinergic inhibition of N‐type calcium currents in sympathetic neurons

One postsynaptic action of the transmitter acetylcholine in sympathetic ganglia is to inhibit somatic N-type Ca²⁺ currents: this reduces Ca²⁺-activated K⁺ currents and facilitates high-frequency spiking. Previous experiments on rat superior cervical ganglion neurons have revealed two distinct pathways for this inhibitory action: a rapid, voltage-dependent inhibition through activation of M₄ muscarinic acetylcholine receptors (mAChRs), and a slower, voltage-independent inhibition via M₁ mAChRs ( 1 –536]. We have analysed the mechanistic basis for this divergence at the level of the individual G-proteins and their α and βγ subunits, using a combination of site-directed antibody injection, plasmid-driven antisense RNA expression, over-expression of selected constitutively active subunits, and antagonism of endogenously liberated βγ subunits by over-expression of βγ-binding β-adrenergic receptor kinase 1 (βARK1) peptide. The results indicate that: (i) M₄ mAChR-induced inhibition is mediated by G_oA; (ii) α and βγ subunits released from the activated G_oA heterotrimer produce separate voltage-insensitive and voltage-sensitive components of inhibition, respectively; and (iii) voltage-insensitive M₁ mAChR-induced inhibition is likely to be mediated by the α subunit of G_q. Hence, Ca²⁺ current inhibition results from the concerted, but independent actions of three different G-protein subunits.