Characterization of brain-derived neurotrophic factor (BDNF) and its receptor, tropomyosin receptor kinase B (TrkB), expression and function in the spinal nociceptive circuitry in a model of chronic neuropathic pain

University of Minnesota Ph.D. dissertation. 2021. Major: Neuroscience. Advisor: Lucy Vulchanova. 1 computer file (PDF); vi, 116 pages.Brain derived neurotrophic factor (BDNF) is widely studied for its role inplasticity that underlies learning and memory, and the maladaptive plasticity that gives rise to chronic pain. BDNF acts through its receptor, tropomyosin kinase B (TrkB), to initiate signaling cascades that result in time-, sex-, region-specific changes in nociceptive circuitry due to nerve injury. Despite great interest in the role of BDNF in neuroplasticity responsible for producing chronic pain states, including neuropathic pain, very few studies include more than one timepoint, region, sex, and methods to evaluate the presence and activity of BDNF. These gaps in the literature are especially problematic when considering reported sex-dependent or contradictory effects of BDNF on pain behaviors. Understanding the scope and exact mechanism of BDNF-TrkB signaling in neuropathic pain is crucial for the development of novel pain therapies that are safe, effective, and unbiased. My thesis work focuses on providing a comprehensive assessment of BDNF and TrkB expression and function which includes both sexes, multiple regions of the nociceptive pathway, cell types, and timepoints after nerve injury. Using a highly sensitive in situ hybridization method, I show that changes in BDNF and TrkB mRNA in DRG and spinal cord are time- and sex-dependent. Although males had higher amounts of TrkB protein in dorsal horn (DH) of the spinal cord, the degree of TrkB activation due to injury was comparable between the sexes. A novel TrkB specific inhibitor, ANA-12, modestly but significantly reduced hypersensitivity in males in a time-dependent manner but not in females. Further investigation into this revealed that ANA-12, instead of inhibiting TrkB activity, increased DH TrkB phosphorylation in both sexes. Finally, the role of microglia in BDNF-TrkB signaling was investigated and although BDNF mRNA was not detected in any microglial profiles in DH, TrkB mRNA was reliably identified in DH microglia after nerve injury. The microglial TrkB phosphorylation was elevated in DH of injured animals pointing to a functional role. To test that hypothesis, I generated a tamoxifen inducible microglia specific TrkB knockout line and conducted behavioral experiments to evaluate the effect of microglial TrkB on pain behaviors. No behavioral differences were detected at any timepoint in either sex. Surprisingly, tissue analysis revealed an elevation in BDNF and TrkB along with the surface area of microglia in DH indicating increased microglial activation in response to microglial TrkB deletion. These data collectively suggest that BDNF-TrkB activity is region, sex, and time-dependent and highlights the importance of validating not just the tools used, but also the effect of the manipulations on the underlying process that is being studied