Use of Integrated Optical Clearing and 2-Photon Imaging to Investigate Sex Differences in Neuroimmune Interactions After Peripheral Nerve Injury

Abstract

Peripheral nerve injury induces a myriad of immune-derived symptoms that negatively impacts pain, depression, and overall quality of life. Neuroimmune differences underlie sexual dimorphisms in various pain states. The innate immune system is a source of these sex differences, which promotes inflammation and pro-nociception through bidirectional signaling with the nervous system. Spatiotemporal interactions between leukocytes and sensory neurons could hold the key to explain ascribed differences between sexes. To date, studies have found it difficult to display these interactions. We are poised to answer important questions regarding the recruitment of peripheral leukocytes to key tissues of the pain system, the dorsal root ganglia (DRG) and sciatic nerve after nerve injury. We optically clear whole DRGs and sciatic nerves and concomitantly use multi-photon microscopy and transgenic reporter lines, to visualize leukocyte dynamics involved in neuropathic pain development following nerve injury. We observed robust sexual dimorphisms in leukocyte recruitment to the lumbar DRGs after nerve injury. We also assessed immune cell size and morphology to understand activation states in the context of nervous tissue inflammation. The altered mechanisms by which the male and female immune systems respond to nerve injury are still topics of further research, however; the continued use of next-generation imaging with advanced whole tissue image analysis remains an important tool in understanding the reciprocal interactions between neuronal and non-neuronal cells.

Mediators of Neuropathic Pain; Focus on Spinal Microglia, CSF-1, BDNF, CCL21, TNF-α, Wnt Ligands, and Interleukin 1β

“…Importantly, the findings illustrate how sexual convergence onto shared cellular and behavioral endpoints, such as allodynia, pain sensitivity or KCC2 downregulation, may mask sex differences in underlying molecular and cellular mechanisms ( 28 ). Other recent work has shown that macrophage invasion of DRG is predominant in males and not in females although both show similar amounts of allodynia following peripheral nerve injury ( 454 ).…”

Peripheral Voltage-Gated Cation Channels in Neuropathic Pain and Their Potential as Therapeutic Targets

“…Among the cellular mechanisms so far identified, it has been reported that spinal microglia activation is required for injury-induced hypersensitivity in males whereas activation and invasion of adaptive immune cells such as T-lymphocytes is required in females ( 451 , 452 ). Macrophage invasion of DRG is predominant in males and not in females ( 453 ) and nociception is regulated by spinal serotonin and noradrenaline in male but not in female mice ( 454 ). It has also recently been shown that ex vivo treatment of live human organ donor spinal cord tissue with BDNF downregulates markers of inhibition and upregulates markers of facilitated excitation in dorsal horn neurons from males but not females ( 455 ).…”