Elucidating the Role of the vlPAG in Novel Pain-linked Escape Behavior Using C57 Sublines

PROJECT SUMMARY Investigating genetic differences that lead to significant variation in pain sensitivity and behavioral responsiveness have contributed to the thorough mapping of the peripheral pain pathway. In contrast, the pain pathway in the brain and the neural mechanisms underlying affective pain are relatively underexplored. By looking beyond the commonly used C57BL/6J (6J) mouse line, I have identified a C57 subline, C57BL/10SnJ (10SnJ), that shows a novel affective pain phenotype of increased pain-linked escape behavior in comparison to 6J mice. Preliminary work has found that 10SnJ mice show increased neural activity in the ventral lateral periaqueductal grey (vlPAG) and anterior cingulate cortex (ACC) during pain-linked escape behavior. A wealth of prior research has shown that the PAG is responsible for escape behavior initiation, and studies have identified a glutamatergic circuit from the ACC to vlPAG that modulates pain. My hypothesis is that 10SnJ mice have heightened ACC to vlPAG circuit activity that drives their massive pain-linked escape behavioral response. This proposal will utilize the 10SnJ outlier strain and existing genetic tools in 6J mice to elucidate the role of the vlPAG in pain-linked escape behavior. Studying this novel behavioral phenotype will increase our knowledge of the circuits and cell types that govern affective pain and heightened pain states. In Aim 1, I will elucidate the vlPAG cell type responsible for increased pain-linked escape behavior using first RNA in situ of known vlPAG specific cell types to identify a target. The sufficiency, necessity, and specificity of this cell type for pain-linked escape behavior will then be determined using excitatory and inhibitory Cre-dependent opsins and an associated 6J Cre line. Next in Aim 2 I will determine the contribution of the ACCGlu → vlPAG circuit to increased pain-linked escape behavior. I will first trace the circuit with anterograde and retrograde viral tracers in 10SnJ and 6J mice and compare the circuit activity during pain-linked escape behavior using cFos immunohistochemistry. A 6J VGlut Cre mouse line will then be used to determine the correlation of the ACCGlu → vlPAG circuit with pain-linked escape behavior using in vivo fiber photometry targeting the vlPAG measuring a Cre-dependent calcium indicator delivered via an anterograde tracer injected into the ACC. Finally I will determine the sufficiency and necessity of the ACCGlu → vlPAG circuit for pain-linked escape behavior by stimulating in the ACC either an excitatory or inhibitory cre dependent opsin delivered via a retrograde tracer injected into the vlPAG. The proposed study would identify a vlPAG cell type and circuit responsible for the novel behavioral type escape behavior in specifically painful situations. This proposal would add significantly to the field's understanding of how the vlPAG contributes to the affective aspect of pain, and prepare me for a career as an independent researcher.