Authors :
Presenting Author: Matthew Summerfield, BA – University of Iowa
Melanie Moquete, BA – University of Iowa
Katelyn Joyal, PhD – Tufts University School of Medicine
Gordon Buchanan, MD, PhD – University of Iowa
Rationale:
Epilepsy affects over 65 million people worldwide. Approximately one-third of this population will be refractory to anti-seizure drugs. These patients are at increased risk for sudden unexpected death in epilepsy (SUDEP). A potential mechanism that causes SUDEP is postictal dysfunction of serotonin (5-HT) neurons. Our lab has previously demonstrated that dorsal raphe nucleus (DRN) 5-HT neurons are critical for CO2 arousal, with increased arousal latency when these neurons are absent. Additionally, DRN neuronal activity is diminished following electrically-induced seizures in rats. Here, we specifically investigated the effects of electrically induced seizures on the 5-HT subpopulation of the DRN. We hypothesized that DRN 5-HT neuronal activity becomes refractory postictally and unresponsive to CO2.Methods:
Pet1::GCaMP mice, generated by crossing Pet1-Cre and Ai96(RCL-GCaMP6s) mice, allowing for the conditional expression of fluorescent Ca2+ indicator GCaMP6s under control of the Pet1 promoter. Mice were instrumented with EEG and EMG electrodes, a fiber photometry cannula placed targeting the DRN (AP: -4.60, ML: ±0.0, DV: -2.93), and a bipolar electrode placed unilaterally into the basolateral amygdala (AP: -1.34, ML: -2.80, DV: -4.70). Amygdala kindling was used to induce reliable electrically triggered tonic-clonic seizures. DRN 5-HT neuronal activity was measured via fiber photometry and the calcium-dependent fluorescence (ΔF/F) was recorded. Mice were placed in plethysmography chambers under room air (21% O2 / bal N2), and after baseline recording, seizures were induced. Following seizure termination, mice were exposed to either CO2 (7% CO2 / 21% O2 / bal N2) or to another room air tank. Histology was performed to confirm implant placement and expression. Results:
In Pet1::GCaMP mice, DRN 5-HT neuronal activity increased during the ictal period and diminished postictally. Seizure duration was positively correlated with the period of elevated fluorescence, which persisted throughout the seizure. Postictal exposure to CO2 did not significantly alter the duration of reduced fluorescent signal compared to room air, suggesting blunted DRN 5-HT responsiveness to CO2. There was no correlation between the duration of postictal generalized EEG suppression (PGES) and duration of reduced fluorescent signal. Furthermore, when exposed to CO2 postictally, averaged ΔF/F values were significantly lower than those when exposed to CO2 without seizure. The significant reduction in the averaged ΔF/F value during the postictal period in addition to CO2 exposure demonstrates how seizures impair the functionality of the chemosensitive DRN 5-HT neurons. Conclusions:
These findings demonstrate that DRN 5-HT neurons have increased activity during seizures but exhibit a refractory state during the postictal period. The lack of responsiveness to CO2 during this period suggests impaired chemosensory function, which may contribute to SUDEP vulnerability. These results underscore the importance of brainstem 5-HT neurons in recovery following seizures. Funding:
Funding: R01NS129722 (GFB), F31NS125955 (KGJ), T32NS007421 (MJS).