Mapping inhibitory and excitatory projections to the preBӧtzinger complex

Abstract

Breathing relies on rhythmic activity generated by the preBӧtzinger complex (preBӧtC), a small brainstem region. Because breathing is influenced by many different physiological, behavioral, and emotional contexts, the preBӧtC is not only highly connected with nearby respiratory related regions in the hindbrain, but also has widespread connections with other mid-and forebrain regions not generally implicated in the regulation of breathing. The specific functions for many of these preBӧtC inputs remain unclear. Although afferent pathways indiscriminately target excitatory and inhibitory neurons in the preBӧtC, we hypothesized that inputs to the preBӧtC would map to distinct brain regions based on the inhibitory or excitatory phenotype of the projecting neurons. To test this, mice that express tdTomato only when both Cre and FlpO recombinases are present (Ai65) were bred with mice that express Cre in inhibitory neurons ( VgatCre). Adult offspring ( VgatCre; Ai65) received a unilateral preBӧtC injection of a retrograde AAV that expresses FlpO recombinase (AAVrg-Flpo), thereby specifically activating tdTomato expression in any inhibitory neurons that project directly to the preBӧtC. Whole-brain imaging revealed many labeled neurons along the ipsilateral ventral respiratory column (VRC), far rostral and caudal of the injection site, extending along the intermediate reticular zone (IRt) from the VII nucleus through the caudal end of the ventral respiratory group (VRG). Interestingly, this well-defined and continuous column of labeled neurons was confined to the ventral IRt at its caudal end but moved dorsally at its rostral end. Only a small number of neurons were labeled along the contralateral VRC. Sparse labeling was present in the medial parabrachial, Kolliker-Fuse, and reticular regions of the pons. Many projecting neurons were observed across subregions of the amygdala, which were almost exclusively ipsilateral. The hypothalamus contained diffuse projecting neurons along its rostral-caudal extent that were also mostly ipsilateral. However, denser labeling was present in a distinctly curved cluster of the lateral hypothalamus, which was observed on both the ipsilateral and contralateral side. Consistent with the preBӧtC having distinct inhibitory and excitatory input pathways, we did not observe significant labelling in other regions with known projections to the preBotC identified previously using non-cell-type specific tracing strategies. These included cortex, lateral parabrachial, NTS, and RVM. This will be tested further by replicating this experiment with targeting of glutamatergic ( Vglut2) neurons. Defining these cell-type-specific inputs to the preBӧtC will provide an essential foundation for future functional studies to determine how these circuits may regulate breathing. R01 HL1660317 (Baertsch) R00 HL145004 (Baertsch). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.