The long term goals of this project are to identify the cellular and molecular mechanisms that lead to inflammation in the cerebral cortex following ischemia and to determine how this inflammatory response exacerbates neuronal damage. In peripheral tissues, the inflammatory response is marked by the cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNFalpha), and interleukin-6 (IL-6). Corticotropin-releasing hormone (CRH), a neuropeptide first identified as a hypothalamic releasing hormone that mediates the stress response, is also produced during inflammation and has been shown to induce IL-1 and IL-6. We and others have demonstrated an important potential link between CRH and ischemic damage in that infusion of a CRH antagonist produces significant attenuation of neural damage induced by cerebral ischemia. Furthermore, there is recent evidence that expression levels of CRH are rapidly upregulated in the cortex following ischemia. We have observed that in addition to neurons, a number of cells in the CNS contain functional CRH receptors including astrocytes, endothelial cells from brain microvasculature, and microglia. Importantly, CRH immunoreactivity is observed in nerve terminals throughout cerebral cortex and over varicosities located on cerebral microvessels. Our observations suggest that release of CRH induced by ischemia participates in the inflammatory reaction. We hypothesize that CRH modulates ischemia-induced inflammation via actions on non-neural cells in the CNS, leading th increased activation of resident inflammatory cells and increased infiltration by peripheral leukocytes. We propose to test these possible mechanisms in vitro and in an animal model of stroke using CRH receptor gene knockout mice to examine the contribution of CRH to the inflammatory response following MCAO.
Specific Aim 1 : To characterize the inflammatory response following ischemia in vivo using flow cytometry to quantify microglial activation and leukocyte infiltration in the brain and to correlate these changes with CRH and CRH receptor expression, cytokine expression, and induction of endothelial cell adhesion molecules.
Specific Aim 2. To characterize the effect of CRH on purified non-neuronal cells from the CNS in culture.
Specific Aim 3. To test the role of CRH in the inflammatory response following ischemia in vivo.
