0171 Parabrachial-CGRP neurons regulate awakenings to pain stimulus. (25th May 2022) - Lynch, Nicole
Abstract: Introduction: The external lateral part of the PB (PBel), receives spinal afferents carrying pain-signals, which are then relayed to other brainstem and forebrain arousal areas. Many PBel neurons that express calcitonin gene-related peptide [CGRP; PBelCGRP] also receive spinal afferents carrying pain signals. Selective activation of PBelCGRP neurons, in CGRP-CreER mice, cause short latency cortical arousal, while inhibition of these neurons prevents waking to hypercapnia. Also, blocking CGRP receptors has a potential to reverse opiate tolerance in rodents, however, mechanism of action or the circuitry remains unknown. Methods: To test if PBelCGRP is the critical relay node transmitting pain to induce cortical arousals, we conducted either genetic deletion or acute optogenetic inhibition of this node, in both inflammatory pain and opto-pain models. All mice were implanted for recording sleep.Inflammatory-pain model: CGRP-creER mice were bilaterally injected with AAV-Flex-DTA (n=10) to delete CGRP neurons in PBel or AAV-GFP (n=6; control). Both groups received a foot injection of either 5% formalin or saline (10 µl) and were recorded for the sleep.Opto-pain model: CGRP-ChR2 mice were injected in the PB with AAV-Flex-JAWS (n=5) and implanted with bilateral optical fibers directed to the PB, and miniature probes in the foot pad to facilitate stimulation of the nociceptors. We recorded sleep while stimulating nociceptors every 5 min with or without opto-inhibition of PBelCGRP. Results: In mice with intact CGRP neurons, foot stimulation with formalin produced 100% wakefulness for the first three hours, while with deletion of PBCGRP neurons, pain induced sleep loss could be attenuated in both males (by 83%) and females(by 60%). This recovery of NREM sleep significantly correlated with loss of PBelCGRP. In the opto-pain model, opto-stimulation of nociceptors at 10Hz for 5 sec woke up mice in 100% of the trials with a short latency of 4.4 ± 0.78 sec, while opto-inhibition of PBelCGRP that expressed JAWS, by red laser that preceded the nociceptive stimulus by 20s, prevented stimulus-induced awakenings. Conclusion: This suggests that PBelCGRP is the main relay node for awakening to pain. We are now investigating the terminal fields of the PBelCGRP neurons to assess which of them contribute to pain induced arousal. Support (If Any): NS112175