Headaches of intracranial origin have been suggested to result from the activation of primary afferent nociceptive neurons innervating intracranial blood vessels and the meninges. Endogenous factors underlying this neuronal activation remain poorly understood. Mast cells were implicated in numerous chronic painful conditions, including intracranial headaches. Given the ability of mast cells to release an abundance of algesic mediators and their proximity to nociceptors, a functional relationship is suggested. However, a direct link between mast cell activation, release of mediators and enhanced nociceptor excitability leading to activation of nociceptive pathways remained unexplored. Based on our preliminary data we hypothesized that dural mast cell degranulation, release of mediators and subsequent leukocyte recruitment promote activation of intracranial dural nociceptors, enhance their mechanosensitivity, and induce activation of dorsal horn neurons that process intracranial nociceptive input. We propose to use a multidisciplinary approach including electrophysiological, pharmacological, histological, and immunocytochemical methods to test our hypothesis. Three specific aims are proposed. In specific aim 1, single-unit recording of dural nociceptors from the trigeminal ganglion will be used to determine whether mast cell degranulation promotes activation and mechanical sensitization of dural nociceptors. In specific Aim 2, using both electrophysiological single unit recording, and c-fos expression as a marker of nociceptive neuronal activation, we will determine whether dural mast cell degranulation is sufficient to promote activation/sensitization of trigeminovascular dorsal horn neurons. In specific aim 3, single-unit recording combined with a pharmacological approach will be used to examine the relative contribution of the mast cell mediators; nitric oxide, TNF-alpha and tryptase and the role of leukocyte recruitment to dural mast cell-induced nociceptor activation and mechanical sensitization. Data obtained in these studies could provide a better understanding on the link between mast cells and deep tissue nociceptors, especially those innervating the intracranial meninges. Results may help identify pharmacological targets that could lead to the development of novel analgesic drugs for the treatment of headaches and possibly other intractable pain syndromes. [unreadable] [unreadable] [unreadable]