Heme oxygenase inhibitors reduce formalin-induced Fos expression in mouse spinal cord tissue

Recent work from our laboratory and others supports a role for heme oxygenase in nociception and pain of several etiologies including inflammatory, incisional and neuropathic. Since it has been observed that heme oxygenase inhibitors reduce formalin-induced pain behaviors in mice and rats, we attempted to determine if this analgesic effect was reflected in a reduction in formalin-induced spinal cord Fos expression, an index of neuronal activation. To perform these studies, it was necessary to first examine the cytoarchitecture of the mouse lumbar spinal cord so that histological sections from known segmental levels could be chosen, and Fos-positive nuclei could be assigned to established dorsal horn laminae. After documenting the segmental and laminar distribution of Fos-positive nuclei following a 5% formalin injection, we went on to determine that the heme oxygenase inhibitor tin-protoporphyrin or morphine reduced this Fos expression as analyzed using confocal fluorescence microscopy. It was also observed that mice lacking expression of heme oxygenase type 2, an isozyme of heme oxygenase found in high abundance in the spinal cord, had lowered Fos expression after the formalin injection. Additional confocal microscopy studies demonstrated widespread expression of heme oxygenase type 2 in spinal cord neuron cell bodies. Double-labeling experiments showed that a high percentage of Fos-positive nuclei identified after administration of formalin were located within heme oxygenase type 2-positive cell profiles. Our studies support the hypothesis that heme oxygenase type 2 plays a role in formalin-induced nociception. Furthermore, from these results we suggest that the heme oxygenase type 2 located in spinal cord dorsal horn neurons participates in this nociceptive pathway.