Abstract
The slow cholinergic transmission in autonomic ganglia is known to be mediated by an inhibition of Kv7 channels via M1 muscarinic acetylcholine receptors. However, in the present experiments using primary cultures of rat superior cervical ganglion neurons, the extent of depolarisation caused by the M1 receptor agonist oxotremorine M did not correlate with the extent of Kv7 channel inhibition in the very same neuron. This observation triggered a search for additional mechanisms. As the activation of M1 receptors leads to a boost in protein kinase C (PKC) activity in sympathetic neurons, various PKC enzymes were inhibited by different means. Interference with classical PKC isoforms led to reductions in depolarisations and in noradrenaline release elicited by oxotremorine M, but left the Kv7 channel inhibition by the muscarinic agonist unchanged. M1 receptor-induced depolarisations were also altered when extra- or intracellular Cl− concentrations were changed, as were depolarising responses to γ-aminobutyric acid. Depolarisations and noradrenaline release triggered by oxotremorine M were reduced by the non-selective Cl− channel blockers 4-acetamido-4′-isothiocyanato-stilbene-2,2′-disulfonic acid and niflumic acid. Oxotremorine M induced slowly rising inward currents at negative membrane potentials that were blocked by inhibitors of Ca2+-activated Cl− and TMEM16A channels and attenuated by PKC inhibitors. These channel blockers also reduced oxotremorine M-evoked noradrenaline release. Together, these results reveal that slow cholinergic excitation of sympathetic neurons involves the activation of classical PKCs and of Ca2+-activated Cl− channels in addition to the well-known inhibition of Kv7 channels.
Highlights
Acetylcholine is the prime transmitter in the ganglia of the entire autonomic nervous system; it excites postganglionic neurons simultaneously via two different types of receptors: Pflugers Arch - Eur J Physiol (2014) 466:2289–2303 nicotinic and muscarinic acetylcholine receptors
The following results indicated that an inhibition of Kv7 channels contributed to this sequence of events: (1) retigabine, an activator of Kv7 channels, abolished noradrenaline release evoked by the mAChR agonist oxotremorine M, but not that triggered by electrical field stimulation [23]; (2) direct inhibition of Kv7 channels by Ba2+ and/or linopirdine elicited action potential- and Ca2+-dependent noradrenaline release from sympathetic neurons [7, 20]; (3) activation of B2 bradykinin receptors on sympathetic neurons caused an inhibition of Kv7 channels [18], on one hand, and led to noradrenaline release, on the other hand, again in an action potential- and Ca2+dependent manner [7]
The results demonstrate that the activation of M1 receptors can depolarise sympathetic neurons and induce noradrenaline release independently of Kv7 channels; the alternative signalling mechanisms include classical protein kinase C (PKC) enzymes and Ca2+-activated Cl− channels
Summary
Acetylcholine is the prime transmitter in the ganglia of the entire autonomic nervous system; it excites postganglionic neurons simultaneously via two different types of receptors: Pflugers Arch - Eur J Physiol (2014) 466:2289–2303 nicotinic (nAChRs) and muscarinic (mAChRs) acetylcholine receptors. Ganglionic transmission via these two receptors can occur independently of each other [8, 36]. Protein kinase C (PKC) can be activated via mAChRs [26], and PKC may contribute to the muscarinic inhibition of Kv7 channels [15], most probably by regulating the PIP2 sensitivity of the channel [24]. It is still unknown whether PKC might be involved in the excitation of postganglionic sympathetic neurons via M1 receptors and, if so, whether a PKC-dependent excitation of sympathetic neurons relies on an inhibition of Kv7 channels
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