Optogenetic stimulation of respiratory related neurons modulates swallow and swallow‐breathing coordination

Abstract

Swallow and breathing are tightly coordinated behaviors in which disruption of this coordination leads to aspiration pneumonia, the leading cause of death in many adult and pediatric diseases. Little is known about the neural coordination of these two behaviors. Previous work in the respiratory column has shown activity in swallow‐related and breathing‐related neurons, as well as neurons active during both swallow and breathing, suggesting coordination at a central network level. Evidence in the intermediate reticular nucleus (postinspiratory Complex, PiCo) suggest involvement in swallow behavior but not swallow‐breathing coordination. We hypothesize that swallow predominately occurs during the expiratory phase of breathing due to inspiratory related neurons suppressing swallow activity. Thus, inhibition of excitatory neurons within the Pre‐Bötzinger complex will increase swallow excitability, while exciting glutamatergic neurons within this network will have the opposite effect. To evaluate swallow circuitry and its coordination with respiratory related networks we use optogenetic approaches to manipulate the activity of inhibitory (Vgat‐Cre) and excitatory (Dbx1‐ERT2Cre, Vglut2‐Cre) neuron populations within the ventral respiratory centers as well as modulation of PiCo (ChAT‐Cre) in the presence of light stimulation. Optogenetic activation (blue laser, 447nm) of Dbx1 cells was performed in adult Dbx1‐ERT2Cre/Ai32(ChR2) mice after exposing the ventral surface of the brainstem in vivo, and placing bilateral glass fiber optics over the ventral surface in the region of the Pre‐Bötzinger complex. Swallow activity was measured by monopolar suction electrodes of the hypoglossal (XII) and vagus (X) nerves as well as cervical sympathetic nerve (cSNA). Phases of breathing was measured via monopolar electromyogram (EMG) of the diaphragm and swallow was stimulated orally by injection of 0.1cc of 2.5% citric acid water. Our preliminary data suggest that stimulating Dbx1 cells decreased the frequency and duration of swallows. Interestingly, we observed in the presence of light stimulation multiple swallow attempts (oral phase without the pharyngeal phase) resulting in hyoid elevation but failure to induce pharyngeal phase. The average swallow in the absence of light stimulation had a duration of 1025ms whereas swallow during light stimulation was 809ms. We believe that activation of Dbx1 neurons in the Pre‐Bötzinger complex suppresses swallow activity and decouples oral and pharyngeal phases of swallow.Support or Funding InformationNIH: P01 HL090554, R01 HL144801 and R01 HL126523