IN-SITU HYBRIDIZATION OF MESSENGER-RNA EXPRESSION FOR IP3 RECEPTOR AND IP3-3-KINASE IN RAT-BRAIN AFTER TRANSIENT FOCAL CEREBRAL-ISCHEMIA

Loss of intracellular calcium homeostasis has been regarded an importa nt factor underlying neuron cell death after cerebral ischemic insult. In the brain, a major mechanism for regulation of intracellular calci um is through the signal transduction pathway involving hydrolysis of poly-phosphoinositides and release of the second messenger, inositol 1 ,4,5-trisphosphate (IP3). IP3 mobilizes calcium by interacting with an intracellular receptor. Upon its release after agonist stimulation, t his second messenger is catabolized by a 3-kinase and a 5-phosphatase. In this study, in situ hybridization was carried out to examine the m RNA expression of IP3 receptor (IP(3)R) and IP3 3-kinase (IP3K) in rat brain cortex after transient focal cerebral ischemia induced by tempo rary occlusion of the middle cerebral artery (MCA) and the common caro tid arteries (CCAs). Results indicate a large decrease (52%) in IP(3)R mRNA levels in the ischemic cortex as compared to that in the contral ateral side at 4 h after a 45 min ischemic insult. By 16 h, practicall y no IP(3)R mRNA could be detected in the ischemic cortex. On the othe r hand, IP3K mRNA levels remained unaltered until 16 h after reperfusi on, during which time, expression in the infarct core decreased but th at surrounding the core area increased instead. Hybridization of adjac ent brain sections with probes for neuron specific enolase (NSE) and b eta-actin indicated also a time-dependent decrease in mRNA levels afte r ischemia, but these changes were less dramatic as compared to IP(3)R . At 16 and 24 h after reperfusion, there was an increase in beta-acti n mRNA in cortical areas outside the MCA cortex, suggesting of reactiv e gliosis. The exquisite sensitivity of IP(3)R gene expression to foca l cerebral ischemic insult and its perturbation during the early reper fusion period suggests an important role of the IP(3)R in cerebral isc hemic injury.