| Osmoregulation is a fundamental cellular process that is most critical to brain homeostasis. A hyperosmotic stimulus within the brain triggers intracellular reactions that result in either neurodegeneration or neuroprotection in response to the cellular insult. The stress-responsive mitogen-activated protein kinase (MAPK), p38, is the central component of a signaling cascade that functions to regulate a diverse set of intracellular events when it is activated by injurious stimuli. In this study, the hyperosmotic stress-induced activation and function of p38 were characterized and the effects of hyperosmolarity on protein synthesis were determined. Hyperosmolarity, produced by the addition of sorbitol to rat brain slices incubated in vitro, produced rapid and prolonged p38 activation, most significantly within the hippocampus. Other MAPK family members, extracellular signal-regulated kinase (ERKI/2) and c-Jun NH2-terminal kinase (JNK) were transiently activated by hyperosmolarity or nonresponsive to it, respectively. Activation of the p38 pathway included phosphorylation of downstream substrates, mitogen-activated protein kinase-activated kinase-2 (MAPKAPK-2) and heat shock protein 27 (HSP27), a signaling pathway known to produce actin stability. Concurrently, hyperosmotic stress decreased total levels of protein synthesis. Nevertheless, the synthesis of a 30-kDa protein, identified as the anti-apoptotic protein, 14-3-3epsilon, was dose-dependently induced in response to increasingly severe hyperosmotic conditions; a finding independent of p38 pathway activation. Taken together, these results suggest the hippocampus responds to hyperosmotic challenge with an attempt at adaptation. However, stress-activated protein kinases are best characterized in association with apoptosis. Examination of apoptotic markers in hippocampal slices exposed to hyperosmotic conditions revealed the translocation of cytochrome c to the cytosolic fraction, a marker of declining mitochondrial function. To a greater extent than the apoptotic stimulus, staurosporine, hyperosmolarity activated caspase-3 resulting in the cleavage of poly(ADP-ribose) polymerase (PARP). Inhibition of p38 attenuated cytochrome c release and dose-dependently reduced caspase-3 activation. These results reveal the apoptotic pathway to be a second downstream correlate of p38 signaling. Taken as a whole, p38 mediates parallel events, simultaneously producing both apoptosis and cytoprotective mechanisms within the hippocampus. Therefore, p38 is uniquely poised to function as a pleiotropic mediator of the hippocampal response to hyperosmolarity. |
