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Reduced output of the phrenic neuromuscular system and the resultant respiratory insuffciency occurs in many neuromuscular disorders. We hypothesized that expressing an excitatory DREADD (designer receptors exclusively activated by designer drugs) in phrenic motoneurons would enable DREADD-ligand induced increases in inspiratory tidal volume in awake, freely behaving rats. ChAT-Cre rats (n= 9; female n= 3) underwent bilateral injections of a Cre-dependent viral construct, AAV9-hSyn-DIO-hM3D(Gq)-mCherry (titer: 2.07x10 12 vg/mL) into the mid-cervical (C4) ventral horn (1 μl injected per side). The combination of focal AAV injection and Cre-dependent expression was expected to drive hM3D(Gq) expression primarily in phrenic motoneurons, which are ChAT-positive and located in the C3-C5 ventral horn. Whole-body plethysmography was used to measure breathing frequency, tidal volume, and minute ventilation before and after intravenous delivery of a DREADD ligand, JHU37160 (J60), or saline (sham). Delivery of the ligand produced an increase in inspiratory tidal volume compared to saline infusion (two-way RM ANOVA, p= 0.037). Respiratory rate was similar between the two conditions (p= 0.582). Two weeks after plethysmography recordings, rats were urethane anesthetized and phrenic nerve electrical activity was directly recorded using suction electrodes. This procedure was done to directly assess the impact of intravenous delivery of the DREADD ligand on phrenic motoneuron output. The J60 ligand caused a rapid, sustained, and bilateral increase in phrenic nerve efferent burst amplitude (one-way RM ANOVA, p< 0.001), whereas saline injection had no impact. The increase in phrenic burst amplitude lasted up to 100 minutes, at which point the experiment was stopped. We conclude that expression of an excitatory DREADD in phrenic motoneurons enables DREADD ligand induced increases in tidal volume in the awake, unrestrained rat. The phrenic nerve recordings verify that this response very likely reflects activation of phrenic motoneurons. Funding: R01HD052682-14 (DDF), R01HL153140-04 (DDF), University of Florida Graduate Fellowship (ESB). 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.
Benevides et al. (Wed,) studied this question.