Anatomical and functional mapping of vagal nociceptive sensory nerve subsets innervating the mouse lower airways by intersectional genetics

Most vagal sensory afferents innervating the lower airways are activated by noxious stimuli including irritants (e.g. TRPV1 agonist capsaicin) and inflammatory mediators, causing nociceptive cardiorespiratory reflexes (e.g. cough, bronchospasm, changes in respiratory drive and heart rate). Vagal ganglia comprise embryologically distinct nodose and jugular neurons, but little is known of their specific contribution to nociceptive reflexes. Using a novel TRPV1^Flp mouse in combination with P2X2^Cre, Tac1^Cre, intersectional reporter mice, and adeno-associated virus we mapped and modulated distinct nociceptive afferents. TRPV1⁺P2X2⁺ neurons were found exclusively in the nodose ganglion and were activated by αβmATP and capsaicin, but rarely expressed Tac1. TRPV1⁺P2X2⁺ fibres innervated the lungs (many projected into the alveoli) but not the trachea. Centrally they innervated the nucleus tractus solitarius (nTS). More than 90% of TRPV1⁺Tac1⁺ neurons were found in the jugular ganglion and were activated by capsaicin but not αβmATP. TRPV1⁺Tac1⁺ fibres innervated the lungs (although none projected into the alveoli) and the trachea submucosa. They terminated solely in the paratrigeminal complex (Pa5). Many TRPV1^-Tac1⁺ neurons were found in both nodose and jugular ganglia that innervated the trachea and large pulmonary airways. These projected to both nTS and Pa5. Using intersectional chemogenetics, we selectively stimulated lower airway afferent subsets using intravenous injections of clozapine-N-oxide. Activation of TRPV1⁺, TRPV1⁺P2X2⁺ or TRPV1⁺Tac1⁺ fibres evoked bradycardia and bradypnoea. Activation of Tac1⁺ fibres evoked tachycardia and tachypnoea. Activation of vagal TRPV1^-Tac1⁺ neurons only evoked tachycardia. These data show the distinct innervation patterns and reflex function of multiple nociceptive vagal afferent subsets. KEY POINTS: Most sensory innervation of the lower airways is provided by vagal afferents, which are divided into distinct nodose and jugular subsets. Our intersectional approach showed that co-expression of P2X2 and TRPV1 labelled nodose nociceptors, that co-expression of Tac1 and TRPV1 labelled jugular nociceptors, and that Tac1 expression with TRPV1 labelled a mixed population of nodose and jugular neurons. Nodose nociceptors, jugular nociceptors and vagal TRPV1-negative Tac1-positive (TRPV1^-Tac1⁺) afferents project to distinct medulla subnuclei, and innervate different regions and tissues within the lower airways. Chemogenetic activation (via intravenous clozapine-N-oxide) of nodose and jugular nociceptors evoked bradypnoea and bradycardia, whereas activation of TRPV1-negative Tac1-positive (TRPV1^-Tac1⁺) afferents evoked tachycardia. These data show that genetically distinct vagal afferent subsets have distinct innervation patterns and reflex function.