Possible involvement of induction of brain-derived neurotrophic factor in the neuroprotective effect of a 5-phenylpyrimidine derivative.

When primary cortical neurons prepared from the brains of rat embryos (E18) were cultured in the absence of serum, most of the neurons died after 3 days in vitro. We used this model to discover compounds which support neuronal survival, and found that a new 5-phenylpyrimidine derivative named FU248 (2-amino-5-(2,4-dichlorophenyl) pyrimidine) inhibited the neuronal cell death in a dose-dependent manner up to 1 microg/mL. Semiquantitative RT-PCR analysis revealed that an exposure of the primary cortical neurons to 1 microg/mL of FU248 transiently and significantly enhanced the expression of genes including brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3). The enhancement of the gene expression was maximal 6 hr after the addition of FU248, and the expression returned to the basal level after 24 hr. Expression of neurotrophin-4 was not detectable throughout the experimental period. The amount of the transcript for BDNF was approximately nine times and sixteen times more abundant than those for NT-3 and NGF, respectively (t=6 hr). Moreover, an anti-BDNF antibody suppressed the effect of FU248, whereas the control antibody did not show any effects on the neuronal survival. These findings strongly suggest that FU248 exerts its neuroprotective effect, at least in part, through induction of BDNF.

Effect of TTC-909 on cerebral infarction following permanent occlusion of the middle cerebral artery in stroke prone spontaneously hypertensive rats.

We investigated the effect of TTC-909, a drug preparation of the stable prostaglandin I(2) analogue clinprost (isocarbacyclin methylester; methyl 5-[(1S,5S,6R,7R)-7-hydroxy-6-[(E)-(S)-3-hydroxy-1-octenyl] bicyclo[3.3.0]oct-2-en-3-yl] pentanoate) incorporated into lipid microspheres, on cerebral infarction 7 days after permanent occlusion of the middle cerebral artery (MCA) in stroke prone spontaneously hypertensive rats (SHRSP). Under the anesthesia, the MCA was permanently occluded above the rhinal fissure. In schedule 1, vehicle or TTC-909 was injected i.v. once daily over 7 days starting immediately after MCA occlusion. In schedule 2, vehicle or TTC-909 was infused for 3 h starting immediately after MCA occlusion. In schedule 3, vehicle or TTC-909 was infused for 3 h starting immediately after MCA occlusion followed by bolus injection once daily over 6 days. Seven days later, the infarct volume was estimated following hematoxylin and eosin staining. Cerebral infarction produced by permanent occlusion of MCA was limited to the cerebral cortex. While this volume was reduced significantly in case of schedule 3, the infarct volume was not reduced significantly in schedules 1 and 2. Ozagrel, a thromboxane A(2) synthetase inhibitor, had no effect on the infarct volume in schedule 3. These results suggest that cerebral infarction can be developed progressively not only during the first few hours but also after a permanent occlusion of MCA in SHRSP. TTC-909 inhibited cerebral infarction, maybe by improving cerebral blood flow and by protecting against neuronal damage.

Effect of TTC-909 in a middle cerebral artery thrombosis model in stroke-prone spontaneously hypertensive rats.

We investigated the effect of TTC-909, a preparation of the stable prostaglandin I(2) analogue clinprost (isocarbacyclin methylester; methyl 5-[(1S,5S,6R,7R)-7-hydroxy-6-[(E)-(S)-3-hydroxy-1-octenyl] bicyclo[3.3.0]oct-2-en-3-yl] pentanoate) incorporated into lipid microspheres, on infarct volume 24 h after photochemically induced thrombotic occlusion of the middle cerebral artery in stroke-prone spontaneously hypertensive rats (SHR). Under anesthesia, the photosensitizing dye rose bengal (20 mg/kg) was administered intravenously and photoirradiation with green light (wavelength 540 nm) on the middle cerebral artery above the rhinal fissure was achieved using a xenon lamp for 10 min. Infarct volume 24 h after the photochemically induced thrombotic occlusion of the middle cerebral artery was significantly larger in stroke-prone SHR than in Wistar rats. When TTC-909 in doses of 100, 300 and 900 ng/kg/h was intravenously infused for 3 h, starting immediately after the end of the 10-min photoirradiation, the infarct volume was dose-dependently reduced and was statistically significant at a dose of 900 ng/kg/h (p < 0.05). Ozagrel, a thromboxane A(2) synthetase inhibitor, significantly reduced the infarct volume. The model of photochemically induced thrombotic occlusion of the middle cerebral artery in stroke-prone SHR is very useful, because the cerebral infarction is large enough and reproducible. TTC-909 may be effective for the treatment of acute ischemic stroke.