Low Dose Ampakine Stimulates Diaphragm Activity and Increases Tidal Volume following Cervical Spinal Cord Injury in Non‐Anesthetized Freely Behaving Rats

Abstract

Sabhya Rana1, 2, 3, Michael D. Sunshine1, 2, 3, David D. Fuller1, 2, 3 1Department of Physical Therapy, 2Center for Respiratory Research and Rehabilitation, 3McKnight Brain Institute, University of Florida, Gainesville, FL Ampakines are allosteric modulators of AMPA receptor channel kinetics, and can stimulate breathing during hypoventilation such as following opioid overdose or in neuromuscular disorders. Our laboratory has shown that acute treatment with an ampakine, CX717 can increase inspiratory phrenic motor output in rat models of incomplete cervical spinal cord injury (SCI), when studied under anesthesia. The next step in the translational pathway to is to determine if the drugs are safe and effective in the awake animal. The goal of the current work was to evaluate the effect of two low impact ampakines (CX717 and CX1739) on diaphragm muscle activation and breathing in freely moving non-anesthetized rats following SCI. Ventilation and metabolic rate (VCO2) were measured using whole body plethysmography and diaphragm EMG was assessed using in-dwelling muscle electrodes. At 4 and 14 days following C2 spinal hemisection injury (C2Hx), rats were given a single intravenous bolus of CX717 (5 mg/kg, n=8), CX1739 (5 mg/kg, n=8) or vehicle (2-hydroxypropyl-beta-cyclodextrin; HPCD; n=8). At the 4-day time point, infusion of CX717 or CX1739 increased peak diaphragm EMG (recorded ipsilateral to C2Hx) by 40-60% compared to vehicle. However, only the CX1739 group showed an accompanying increase in tidal volume (20%). During an acute respiratory challenge with hypoxia-hypercapnia (10% O2, 7% CO2), both the CX717 and CX1739 groups showed increased diaphragm EMG compared to vehicle (65-75%); the CX1739 group showed a larger increase in tidal volume. At 14 days post injury, CX717 infusion increased the peak inspiratory ipsilateral diaphragm EMG activity (~30%) and tidal volume (~20%) during baseline breathing. In contrast, CX1739 was less effective at 14 days and produced an increase in ipsilateral diaphragm EMG activity of ~10% with no change in tidal volume. During the hypoxia-hypercapnia challenge, both the CX1739 and CX717 groups showed increased ipsi-lesional diaphragm EMG (25-35%) but no increase in tidal volume as compared to vehicle group. Neither ampakine had an impact on respiratory rate at either time point, suggesting a spinal (vs. medullary) mechanism of action. Additionally, no adverse off-target effects (e.g., changes in heart rate or hypoactive state) were apparent. Improved respiratory muscle activation is often a desirable therapeutic outcome after incomplete cervical SCI. Our data indicate that low dose, low impact ampakine treatment can increase diaphragm muscle output after cervical SCI, and therefore may provide a pharmacologic strategy that could be useful in the context of respiratory rehabilitation. The divergent response to CX717 vs. CX1739 at 4 vs. 14 days post-injury merits further study; one possibility is that ongoing neuroplastic changes in AMPA receptor expression after SCI may differentially influence the efficacy of different ampakines.