Nonopioid actions of dynorphin and neuropathic pain

Abstract

The hypothesis of this dissertation is that peripheral nerve injury will elevate dynorphin in the spinal cord, which has a facilitating effect on the nociceptive pathway via a non-opioid mechanism. Two major questions are addressed in this dissertation: how dynorphin is up regulated, and how dynorphin produces a facilitating effect on the nociceptive pathway. The first question was approached by examining the activation of NF-κB, AP-1, CREB, and MAP kinase pathways following nerve injury. These experiments indicate that there was an increase in the activation of NF-κB, AP-1 and CREB. While it was not known whether the increase in AP-1 and NF-κB activity was occurring in dynorphin-expressing cells, phospho-CREB-ir did not colocalize with prodynorphin. Thus CREB may not play a direct role in dynorphin gene expression. Following nerve injury, there was an increase in phospho-ERK1/2, which was also seen in dynorphin-expressing cells. It is possible that the activation of ERK1/2 may contribute to the up-regulation of dynorphin in some dynorphin-expressing cells. There was also a dramatic increase in phosopho-p38-ir. However, it did not colocalize with prodynorphin. In summary, these experiments indicated that there are profound changes in the cellular signaling pathways following nerve injury, which may play an important role in initiating neuroplastic changes in the spinal cord, including the up-regulation of dynorphin. There is considerable evidence suggesting the involvement of NMDA receptors in the excitatory effect of dynorphin in vivo and possible interaction between dynorphin and NMDA receptors. The interaction between dynorphin and NMDA receptors was further investigated by radioligand binding assay using iodinated dynorphin A(2-17). These experiments suggest dynorphin binds to NMDA receptors with relatively high affinity. Dynorphin appears to interact preferentially with the closed state of NMDA receptors, and may be inhibitory on NMDA receptors. To look for mechanisms which may underlie the excitatory effect of dynorphin, I examined its effect on intracellular calcium level ([Ca²⁺]ᵢ). These experiments suggest that dynorphin A(2-17) elicited an increase in [Ca²⁺]ᵢ in the cortical neurons via a non-NMDA, non-opioid mechanism. This novel action of dynorphin could provide a potential mechanism for the excitatory actions of dynorphin in vivo.