Cholinergic collaterals arising from noradrenergic sympathetic neurons in mice.

The sympathetic nervous system vitally regulates autonomic functions, including cardiac activity. Postganglionic neurons of the sympathetic chain ganglia relay signals from the central nervous system to autonomic peripheral targets. Disrupting this flow of information often dysregulates organ function and leads to poor health outcomes. Despite the importance of these sympathetic neurons, fundamental aspects of the neurocircuitry within peripheral ganglia remain poorly understood. Conventionally, simple monosynaptic cholinergic pathways from preganglionic neurons are thought to activate postganglionic sympathetic neurons. However, early studies suggested more complex neurocircuits may be present within sympathetic ganglia. The present study recorded synaptic responses in sympathetic stellate ganglia neurons following electrical activation of the pre- and postganglionic nerve trunks and used genetic strategies to assess the presence of collateral projections between postganglionic neurons of the stellate ganglia. Orthograde activation of the preganglionic nerve trunk, T-2, uncovered high jitter synaptic latencies consistent with polysynaptic connections. Pharmacological inhibition of nicotinic acetylcholine receptors with hexamethonium blocked all synaptic events. In order to prove that high jitter, polysynaptic events were due to the presence of cholinergic collaterals from postganglionic neurons within the stellate ganglion, we knocked out choline acetyltransferase in adult noradrenergic neurons. This genetic knockout eliminated orthograde high jitter synaptic events and EPSCs evoked by retrograde activation. These findings suggest that cholinergic collateral projections arise from noradrenergic neurons within sympathetic ganglia. Identifying the contributions of collateral excitation to normal physiology and pathophysiology is an important area of future study and may offer novel therapeutic targets for the treatment of autonomic imbalance. KEY POINTS: Electrical stimulation of a preganglionic nerve trunk evoked fast synaptic transmission in stellate ganglion neurons with low and high jitter latencies. Retrograde stimulation of a postganglionic nerve trunk evoked direct, all-or-none action currents and delayed nicotinic EPSCs indistinguishable from orthogradely-evoked EPSCs in stellate neurons. Nicotinic acetylcholine receptor blockade prevented all spontaneous and evoked synaptic activity. Knockout of acetylcholine production in noradrenergic neurons eliminated all retrogradely-evoked EPSCs but did not change retrograde action currents, indicating noradrenergic neurons have cholinergic collaterals connecting neurons within the stellate ganglion. Abstract figure legend Sympathetic ganglia such as the stellate ganglion relay sympathetic signals from the central nervous system to autonomic targets, including the heart. Conventionally, simple monosynaptic cholinergic pathways from preganglionic neurons via T-2 activate postganglionic sympathetic neurons. However, early studies suggested that collateral projections may be present between postganglionic neurons of the stellate ganglion. To test this hypothesis, we developed a new mouse model that conditionally and selectively deletes choline acetyltransferase (ChAT), the synthetic enzyme for ACh, from noradrenergic neurons (TH ChAT KO ). Our experimental approach centered on retrogradely activating these postganglionic neurons using the inferior cardiac nerve (iCN). In Control (TH WT ) SG neurons, single shocks to the iCN evoked large inward somatic currents consistent with retrogradely-conducted action potentials invading the recorded SG neuron cell body (i.e. retrograde action currents in voltage-clamp; rACs; upper panels). These rACs were often followed by excitatory synaptic currents (eEPSCs) with unique stimulus thresholds consistent with activation of axons from other postganglionic neurons. Genetic knockout of ACh from noradrenergic sympathetic neurons (lower panels) abolished retrogradely-activated eEPSCs, confirming the presence of cholinergic collateral transmission between postganglionic neurons in the SG. This article is protected by copyright. All rights reserved.