A GABAergic inhibitory microcircuit controlling cholinergic outflow to the airways

Case Western Reserve University, Cleveland, Ohio, United States
Journal of Applied Physiology (Impact Factor: 3.06). 02/2004; 96(1):260-70. DOI: 10.1152/japplphysiol.00523.2003
Source: PubMed


GABA is the main inhibitory neurotransmitter that participates in the regulation of cholinergic outflow to the airways. We have tested the hypothesis that a monosynaptic GABAergic circuit modulates the output of airway-related vagal preganglionic neurons (AVPNs) in the rostral nucleus ambiguus by using a dual-labeling electron microscopic method combining immunocytochemistry for glutamic acid decarboxylase (GAD) with retrograde tracing from the trachea. We also determined the effects of blockade of GABAA receptors on airway smooth muscle tone. The results showed that retrogradely labeled AVPNs received a significant GAD-immunoreactive (GAD-IR) terminal input. Out of a pooled total of 3,161 synaptic contacts with retrogradely labeled somatic and dendritic profiles, 20.2% were GAD-IR. GAD-IR terminals formed significantly more axosomatic synapses than axodendritic synapses (P < 0.02). A dense population of GABAergic synaptic contacts on AVPNs provides a morphological basis for potent physiological effects of GABA on the excitability of AVPNs. GAD-IR terminals formed exclusively symmetric synaptic specializations. GAD-IR terminals were significantly larger (P < 0.05) in both length and width than unlabeled terminals synapsing on AVPNs. Therefore, the structural characteristics of certain nerve terminals may be closely correlated with their function. Pharmacological blockade of GABAA receptors within the rostral nucleus ambiguus increased activity of putative AVPNs and airway smooth muscle tone. We conclude that a tonically active monosynaptic GABAergic circuit utilizing symmetric synapses regulates the discharge of AVPNs.

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    • "We used triple immunohistochemical methods to label neural tracers with DAB (above) and parvalbumin (PV) or calbindin (CB) neurons with either silver-enhanced gold-conjugated or tetramethylbenzidine (TMB) labeled secondary antibodies. These methods show distinct labeling at the EM: DAB appears as a dark uniform precipitate, silver-enhanced gold particles as circular clumps of variable size, and TMB as rod-shaped crystals (e.g., Pinto et al., 2003; Gonchar and Burkhalter, 2003; Moore et al., 2004; Zikopoulos and Barbas, 2007). "
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    • "In the quantitative analysis of neurons which project to the antrum we found that of all the synapses observed with these neurons, 23.0 +/− 3.6% contained GAD67-IR. This percentage is very similar to data of others (Moore et al., 2004) who reported findings on GAD-IR terminals synapsing with airway-related vagal preganglionic neurons (AVPNs) in the rostral nucleus ambiguous. From all of the synaptic contacts with retrogradely labeled AVPNs, 20.2% were GAD-IR. "
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    • "The three methods show distinct labeling at the EM: DAB appears as a dark uniform precipitate, silver-enhanced gold particles as circular clumps of variable size, and TMB as rod-shaped crystals [e.g., (Pinto et al., 2003; Gonchar and Burkhalter, 2003; Moore et al., 2004; Medalla et al., 2007; Zikopoulos and Barbas, 2007); and personal observations]. "
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