Anatomical Basis of Interactions Between Locomotor and Respiratory Systems

Abstract

Functional interaction between locomotor and respiratory networks has been shown in multiple previous studies. Here, we demonstrate the anatomical basis of these interconnections using the retrograde transsynaptic tracer, pseudorabies virus (PRV), in adult Sprague Dawley rats. In particular, PRV‐152 with a green fluorophore (GFP) was injected into the left biceps brachii muscle and PRV‐614 with a red fluorophore (RFP) was applied onto the left hemidiaphragm to map biceps and diaphragm motor circuits. Cervical spinal cord and brainstem tissue were sectioned and immunostained to enhance RFP and GFP signals and stain acetylcholinergic neurons using choline acetyltransferase (ChAT) antibodies. Double‐labeled (GFP+RFP) and triple‐labeled (GFP+RFP+ChAT) neurons were identified within the spinal cord and brainstem. In the spinal cord, double and triple‐labeled interneurons were seen from C2 cervical to T1‐T2 thoracic levels ipsilateral and contralateral to PRV labeling. Interestingly, most of these neurons were found in the Rexed lamina 7 and 10. The presence of ChAT positive shared interneurons within these laminae allows us to suggest that they could be Voc interneurons, but more studies are required to support this hypothesis. In the brainstem, the double‐labeled neurons were seen in the following structures: 1) midbrain – in all parts of Periaqueductal Gray substance (PAG), Dorsal Raphe (DR) and Deep Mesncephalic (DMe) nuclei; 2) pons – in Kölliker‐Fuse and Parabrachial Nuclei (PB/KF), in areas of noradrenergic groups of cells (Locus Coeruleus and A5) and sub‐Coeruleus areas, in oral and caudal Pontine nuclei (PnO and PnC); 3) medulla – in all parts of Raphe (Obscurus, Magnus and Pallidus), Gigantocellular Reticular (Gi), Internal Reticular (IRt), and lateral Paragigantocellular (LPGi) nuclei, in para‐Fascial/Retrotrapezoid area (pF/RTN), and in Bötzinger and pre‐Bötzinger Complexes (BC and pBC). Triple‐labeled cells were found in KF, Raphe and LPGi structures. Our work confirmed the previous electrophysiological studies revealing shared interneurons in the spinal cord and pontomedullary respiratory compartments (PB/KF, pF/RTN) and uncovered the new brainstem structures that also could play a significant role in functional interactions between locomotor and respiratory systems.