Unraveling respiratory rhythmic integration from rhythm generation to motor outpu

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DESCRIPTION (provided by applicant): Mammalian breathing is composed of three phases: inspiration (I), post-inspiration (E1), and active expiration (E2). It is well known that the transverse slice preparation isolating the preB¿tzinger complex (preB¿tC) spontaneously generates inspiratory activity. The absence of expiratory population activity is consistent with th theory that a larger network is required to generate the three different phases. The preB¿tC is only one of several interacting networks that exist bilaterally and extend rostrocaudally in the ventrolateral medulla of the brainstem and is collectively termed the ventral respiratory column (VRC). The functional interactions between VRC networks, and more specifically how expiratory rhythms emerge from the respiratory network are largely unknown Based on these observations, we hypothesize that the longer burst durations in the VRC slice are due to the partial separation of inspiratory (I) and post-inspiratory (E1) phases, and the infrequent rostral burst activity reflects active expiration (E2). We further hypothesize that the different phases ar shaped by synaptic inhibition and synaptic interactions dissociate when exposed to episodic hypoxia, a common occurrence in multiple pathologies. These interactions have so far only been studied from in vivo and in situ preparations. Unfortunately neither in vivo nor in situ preparations are amenable to the same degree of cellular rigor as a slice preparation. This becomes a major disadvantage when trying to understand the cellular mechanisms underlying the generation of respiratory phases. We have developed a novel horizontal slice (VRC slice, mice, p5-p7; 700-1100¿m) that isolates the entire VRC from the rostral edge of the facial nucleus to C3 in the spinal cord and that retains bilateral connectivity. Initial observations from population recordings in the VRC slice reveal: broader burst durations compared to the transverse slice, a second rostral phase, synchronized preB¿tC and rostral phases in the presence of gabazine, and frequency irregularities after exposure to episodic hypoxia, similar to results seen in vivo. Thus, our overarching goal is to functionally characterize network-to-network interactions that underlie the generation of the three phases of respiration. We combine electrophysiological, optogenetic, and pharmacological techniques in three approaches toward this goal: (1) mapping population rhythms across the slice and correlating pre-motor activity with motor output, (2) investigating the role of synaptic interactions in establishing different respiratory phases, (3) examining the interactions between the preB¿tC and rostral rhythms following exposure to acute intermittent hypoxia. These experiments will further our understanding into the interacting networks responsible for generating the phases of breathing rhythms and provide insight into the destabilizing network effects of episodic hypoxia throughout the medullary VRC.