Effects of blockade of voltage-sensitive Ca2+/Na+ channels by a novel phenylpyrimidine derivative, NS-7, on CREB phosphorylation in focal cerebral ischemia in the rat

NS-7 is a novel blocker of voltage-sensitive Ca2+ and Na+ channels, and it significantly reduces infarct size after occlusion of the middle cerebral a rtery. Persistent activation of cyclic AMP response element binding protein (CREB), which can be induced by increase in intracellular Ca2+ concentrati ons or other second messengers, has recently been found to be closely assoc iated with neuronal survival in cerebral ischemia. The present study was th erefore undertaken to evaluate the neuroprotective effects of NS-7 by analy zing changes in CREB phosphorylation in a focal cerebral ischemia model. CR EB phosphorylation in the brain of rats was investigated immunohistochemica lly at 3.5-48-h recirculation after 1.5-h occlusion of the middle cerebral artery. NS-7 (1 mg/kg; NS-7 group) or saline (saline group) was intravenous ly injected 5 min after the start of recirculation. The NS-7 group showed s ignificantly milder activation of CREB phosphorylation in various cortical regions after 3.5 h of recirculation than the saline group. The inner borde r zone of ischemia in the NS-7 group subsequently exhibited a moderate, but persistent, increase in number of phosphorylated CREB-positive neurons wit h no apparent histological damage. By contrast, the saline group displayed a marked, but only transient, increase in number of immunopositive neurons in this border zone after 3.5 h of recirculation, and this was followed by clear suppression of CREB phosphorylation and subsequent loss of normal neu rons. These findings suggest that: (1) the marked enhancement of CREB phosp horylation in the acute post-ischemic phase may be triggered largely by an influx of calcium ions as a result of activation of the voltage-sensitive C a2+ and Na+ channels; and that (2) the neuroprotective effects of NS-7 may be accompanied by persistent activation of CREB phosphorylation in the inne r border zone of ischemia. (C) 2000 Elsevier Science B.V. All rights reserv ed.