Investigating Mechanisms in Nociceptors Driving Ongoing Activity and Ongoing Pain

Ongoing (apparently spontaneous) pain at rest is a major complaint of patients suffering from many forms of acute and chronic pain, including acute and persistent postsurgical pain. Accumulating evidence suggests ongoing activity in nociceptors is a major driver of ongoing pain. Ongoing activity can be generated in sensory neurons in the absence of sensory generator potentials if one or more of three neurophysiological alterations occur – prolonged depolarization of resting membrane potential (RMP), hyperpolarization of action potential (AP) threshold, and/or increased amplitude of depolarizing spontaneous fluctuations of membrane potential (DSFs) to bridge the gap between RMP and AP threshold. Cellular signaling pathways that increase DSF amplitude and promote ongoing activity acutely in nociceptors were unknown for any neuromodulator, as were the neurophysiological alterations that underly ongoing activity in nociceptors after deep tissue incision. The work presented in this dissertation sought to identify a cellular signaling pathway and injury- and inflammation-related neuromodulator that induces alterations that drive hyperexcitability, such as enhanced DSFs, and thereby potentiates ongoing activity in nociceptors, as well as to determine the alterations that contribute to nociceptor hyperactivity associated with postsurgical pain. A combination of whole-cell patch clamp electrophysiology, pharmacology, and high content microscopy was used to define the effects of low concentrations of the injury-related proinflammatory mediator serotonin and the major pathway by which it exerts its effects. This study shows that serotonin enhances DSFs, hyperpolarizes AP threshold, and thereby potentiates OA in isolated rat DRG neurons at a concentration of 100 nM, and serotonin exerts these effects on nonpeptidergic nociceptors via the 5-HT4 receptor and downstream cAMP signaling via PKA and EPAC. Furthermore, whole-cell patch clamp recordings of primary DRG neurons, analysis of spontaneous pain behavior, and RNA sequencing were used to reveal an unexpected persistence of hyperexcitability in isolated DRG neurons and potentially associated differential gene expression, both of which were found after the behavioral expression of postsurgical pain had resolved. In addition to elucidating cell signaling mechanisms that can contribute to acute ongoing activity in nociceptors, these studies set the stage for future investigations addressing important questions about nociceptor contributions to pain