Cerebral vasospasm (CV) is a significant cause of mortality (11,000 per year in the US) and morbidity in patients surviving a subarachnoid hemorrhage (SAH). Onset of CV occurs 3-10 days after the SAH, and this affords us a valuable therapeutic window. There are currently no effective therapies for this debilitating pathology. Our long term goal is to determine the mechanism of smooth muscle contraction during CV after SAH. We believe this knowledge will enable clinicians to use the therapeutic window to prophylactically treat all SAH patients and thus prevent onset of CV. Our specific hypothesis is that signalling events in the smooth muscle comprise a protein kinase C-mediated contraction coupled with a rho-mediated inhibition of myosin light chain phosphatase (MLCP), leading to intractable vasoconstriction. We hypothesize that these events involve oxidation products of bilirubin (BOXes). This hypothesis is based on the observations that (i) CSFV (cerebrospinal fluid from patients that developed CV) but not CSFC (non-vasospastic SAH CSF), has been shown to inhibit MLCP in vitro, (ii) PKC and Rho activation have been implicated in the initiation and maintenance of CV in animal models, and (iii) BOXes are found in human (CSFV), but not in CSFN.
The specific aims are to: 1. Determine the involvement of PKC in our model. We will determine which isoforms of PKC are activated in response to CSFV in the smooth muscle. We will confirm these observations with isoform-specific PKC inhibitors. 2. Determine the involvement of phosphatase inhibition in our model. Activities of MLCP and upstream molecules in this cascade will be investigated to determine the level at which CSFV modulates phosphatase activity. 2. Determine the role of BOXes in the aetiology of CV. Are BOXes involved in the PKC activation, the MLCP inhibition, or both? Both pathways will be investigated and we will compare these findings with those seen in CSFV treated smooth muscle. The results obtained from completion of the aims stated above will be of clinical relevance, and application to the human pathology.
