Molecular mechanisms in purine-mediated protection

Stroke caused by cerebral ischemia is the third leading cause of death and the leading cause of neurological disability. During hypoxia, ATP production is diminished and the energy status of neurons is compromised, ionic gradients dissipate, and cell membranes depolarize. Concomitant with energy depletion is the formation of the purine nucleoside adenosine, which acts as a powerful endogenous neuroprotectant during ischemia-induced energy failure, i.e. by decreasing neuronal metabolism and increasing cerebral blood flow. Purine nucleosides thereby are inducing several distinct intercellular signaling pathways. This is particularly the case in the brain, which expresses high concentrations of adenosine receptors. These effects are mediated through interaction of adenosine with specific receptors, and stimulation and subsequent signaling functions of the adenosine receptors was hypothesized to result in an effective treatment of stroke. Similar effects were observed with the related purines inosine and guanosine, which positively regulate neuronal development and/or support neurite outgrowth. Many aspects of the mechanisms involved in purine-mediated protection remain unclear. Our data on the activation and potential causal roles of p42/p44 MAPK and, very recently, p120/140 PRK1/2 in these pathways, to our opinion, now strongly support a detailed investigation of 1.) the critical roles of serinehreonine protein kinases p42/p44 MAPK (together with the established effectors ELK-1 and HIF-1a) as well as PRK1/2, in purine nucleoside-mediated signaling leading to regeneration and/or survival of neurons, and finally 2.), the concept of potential use of these signaling mechanisms for future ischemia/reperfusion drug therapies.

To reduce apoptosis in the brain is central to functional recovery after stroke. In response to hypoxia/ischemia purine nucleosides are formed and act as powerful endogenous neuroprotectants. Specific aims of the project P19578-B05 were studies of: 1.) The impact of purine nucleoside-mediated signaling on regeneration and survival of neurons, 2.) The potentially critical roles of serinehreonine protein kinases p42/44 MAPK and PRK1/2 in purine-mediated neuroprotection and 3.) The evaluation of the concept of the potential use of these signaling functions for future ischemia/reperfusion drug therapies. In our work, using two neuronal cell models (A2a receptor positive PC12 cells and A1 receptor positive primary cerebellar granule neurons) and a low oxygen incubator, we have identified the importance of two key signaling modules. We found that, viability and neurite formation critically depend on functional activity of the mitogen- activated protein kinase (MAPK); (Tomaselli, 2008) and the stabilization of the Hypoxia- inducible factor-1 alpha (HIF-1alpha); (Zur Nedden, 2008). Furthermore, employing siRNA knockdown-techniques, pioneer studies of our group showed the involvement of the Proteinkinase-C-related kinase (PRK1) in purine-mediated neuroprotection (Thauerer, 2010).