HUMAN-BASED EX VIVO ASSAYS FOR PAIN CIRCUIT RESEARCH: A TRANSLATIONAL ALTERNATIVE TO ANIMAL MODELS

Chronic pain affects more than one-third of the global population, reduces quality of life, and imposes substantial socioeconomic costs. Although preclinical studies have clarified many mechanisms underlying persistent pain, translation to patients remains limited, underscoring the need for more predictive assays and improved recognition of interspecies differences. Here, we developed a surgical approach to rapidly recover intact human spinal cord from recently deceased organ donors and evaluated the preservation of neuronal and glial populations by immunolabeling tissue from 16 donors (11 males and 5 females). In parallel, we established organotypic cultures of lumbar spinal cord slices that remained viable for several weeks and were compatible with functional assays. Using adeno-associated viral vectors, we drove somatic GFP expression in dorsal horn cells, enabling targeted whole-cell recordings from visually identified neurons (n=44; 3 males, 4 females). Neuronal capacitance, input resistance, rheobase, spike threshold, and resting membrane potential did not differ significantly by sex; however, input resistance and rheobase varied with donor medical history. To assess network dynamics, we performed calcium imaging to quantify coordinated dorsal horn activity evoked by electrical and chemical stimulation. This human ex vivo platform provides a translationally relevant approach to interrogate spinal pain circuitry and to evaluate candidate analgesics.