BDNF in Phrenic Motor Neurons: Effects of Cervical Spinal Injury and Intermittent Hypoxia
Authors: Oberto Jose Rafael , Ciesla Marissa C. , Seven Yasin B. , Kelly Mia N. , Allen Latoya L. , Smith Kristin , Asa Zachary , Simon Alec , Holland Ashley , Santiago Juliet , Stefan Kelsey , Ross Ashley , Gonzalez-Rothi Elisa J. , Mitchell Gordon S. 
Published: April 2020

Cervical spinal cord injury (cSCI) compromises breathing. Since respiratory impairment increases mortality, there is a critical need for new treatments to enhance breathing function after cSCI. One emerging strategy to enhance breathing function after cSCI is to elicit spinal respiratory plasticity via exposure to acute intermittent hypoxia (AIH). Brain derived neurotrophic factor (BDNF) is both necessary and sufficient for AIH‐induced phrenic motor plasticity. Repetitive AIH exposure enhances BDNF expression within phrenic motor neurons in spinally intact rats, but its effects after cSCI are not well known. In contrast to the beneficial effects of AIH, chronic intermittent hypoxia (CIH), similar to that experienced during sleep apnea elicits neuropathology (including inflammation) that may undermine plasticity. The balance between the therapeutic vs pathogenic effects of intermittent hypoxia is a matter of “dose.” Since the impact of various intermittent hypoxia protocols on phrenic motor neuron BDNF expression in phrenic motor neurons has not been investigated, we tested the hypothesis that BDNF expression in phrenic motor neurons is enhanced by repetitive AIH, but not CIH, in spinally intact and injured rats. BDNF expression was assessed in the phrenic motor neurons of male rats with and without C2 hemisection (C2Hx; 12 wks post‐injury) exposed to 28 days of: 1) normoxia; 2) daily AIH (10, 5‐min 10.5% O2 episodes per day; 5‐min normoxic intervals); 3) mild CIH (5‐min 10.5% O2 episodes, 5‐min intervals; 8 hrs/day); and 4) moderate CIH (2‐min 10.5% O2 episodes, 2‐min intervals; 8 hrs/day). Rats received intrapleural Cholera toxin B (CtB) injections 14 days before C2Hx to label phrenic motor neurons. After the study, spinal cords were harvested and BDNF expression assessed via immunofluorescence using a custom MATLAB algorithm to determine BDNF optical density within identified phrenic motor neurons and in a region of interest (50 mm radius) around the center of gravity of CtB labeled neurons. Spinally intact and injured rats have differential responses to acute and chronic intermittent hypoxia. Whereas spinally intact rats exposed to daily AIH increase BDNF expression within and around phrenic motor neurons; there were no effects of mild or moderate CIH. In contrast, BDNF within and around phrenic motor neurons in spinal cord injured rats was unaffected by daily AIH, and was highest after moderate CIH. Thus, whereas only daily AIH elicits neurochemical plasticity in intact rats, similar effects were observed only with moderate CIH after C2Hx. These data support the idea that the “injured spinal cord is a new spinal cord,” and highlight the potential of intermittent hypoxia preconditioning to enhance the capacity for BDNF‐dependent phrenic motor plasticity and (possibly) functional recovery after cSCI.Support or Funding InformationAPS HEARST (JRO), SPARC (OT2OD023854) NIH T32 HD043730 (LLA) and HL134621 (MNK), NIH K12 HD055929 (EGR) and the UF McKnight Brain Institute