Morphine paradoxically prolongs neuropathic pain in rats by amplifying spinal NLRP3 inflammasome activation

Pain after disease/damage of the nervous system is predominantly treated with opioids, but without exploration of the long-term consequences. We demonstrate that a short course of morphine after nerve injury doubles the duration of neuropathic pain. Using genetic and pharmacological interventions, and innovative Designer Receptor Exclusively Activated by Designer Drugs disruption of microglia reactivity, we demonstrate that opioid-prolonged neuropathic pain arises from spinal microglia and NOD-like receptor protein 3 inflammasome formation/activation. Inhibiting these processes permanently resets amplified pain to basal levels, an effect not previously reported. These data support the “two-hit hypothesis” of amplification of microglial activation—nerve injury being the first “hit,” morphine the second. The implications of such potent microglial “priming” has fundamental clinical implications for pain and may extend to many chronic neurological disorders

Cervical Epidural Electrical Stimulation Increases Respiratory Activity through Somatostatin-Expressing Neurons in the Dorsal Cervical Spinal Cord in Rats

We tested the hypothesis that dorsal cervical epidural electrical stimulation (CEES) increases respiratory activity in male and female anesthetized rats. Respiratory frequency and minute ventilation were significantly increased when CEES was applied dorsally to the C2-C6 region of the cervical spinal cord. By injecting pseudorabies virus into the diaphragm and using c-Fos activity to identify neurons activated during CEES, we found neurons in the dorsal horn of the cervical spinal cord in which c-Fos and pseudorabies were co-localized, and these neurons expressed somatostatin (SST). Using dual viral infection to express the inhibitory Designer Receptors Exclusively Activated by Designer Drugs (DREADD), hM4D(Gi), selectively in SST-positive cells, we inhibited SST-expressing neurons by administering Clozapine N-oxide (CNO). During CNO-mediated inhibition of SST-expressing cervical spinal neurons, the respiratory excitation elicited by CEES was diminished. Thus, dorsal cervical epidural stimulation activated SST-expressing neurons in the cervical spinal cord, likely interneurons, that communicated with the respiratory pattern generating network to effect changes in ventilation.SIGNIFICANCE STATEMENT A network of pontomedullary neurons within the brainstem generates respiratory behaviors that are susceptible to modulation by a variety of inputs; spinal sensory and motor circuits modulate and adapt this output to meet the demands placed on the respiratory system. We explored dorsal cervical epidural electrical stimulation (CEES) excitation of spinal circuits to increase ventilation in rats. We identified dorsal somatostatin (SST)-expressing neurons in the cervical spinal cord that were activated (c-Fos-positive) by CEES. CEES no longer stimulated ventilation during inhibition of SST-expressing spinal neuronal activity, thereby demonstrating that spinal SST neurons participate in the activation of respiratory circuits affected by CEES. This work establishes a mechanistic foundation to repurpose a clinically accessible neuromodulatory therapy to activate respiratory circuits and stimulate ventilation