| Summary: | 碩士 === 國立陽明大學 === 神經科學研究所 === 92 === To investigate the functions and expression regulation of LUZP in brain, we in collaboration with professor NCChang have established a Luzp-KO/lacZ-KI mutant mice model. In situ X-gal staining of the expressed β-galactosidase (encoded by lacZ) provides an alternative tracing method for the temporal and spatial expression patterns of LUZP. Spatial expression of LUZP was mapped to cerebral cortex, hippocampus, hypothalamus, mid-brain, brain stem and dorsal root ganglion (DRG) in the adult brain. Temporal course of expression reveals that LUZP is first detected in both CA1 and CA3 of hippocampus at P14. Differential regulation mechanism exists such that LUZP in CA1 is down-regulated as a function of time whereas that in CA3 persists. By P28, LUZP expression may be detected in all regions cited above and persists through adulthood.
As CA3 of hippocampus is implicated in spatial learning and memory and highly sensitive to stress; we have screened for phenotypic traits employing behavioral paradigms of spatial learning/memory and emotion memory out from fear or stress-anxiety. Despite the fact that homozygous Luzp-KO/lacZ-KI mice are embryonic lethal, the heterozygous mutant containing one allele of Luzp remains a valid model to unravel the physiological function of LUZP in adult CNS via the pharmacogenetic approach. When compared to their wild type littermates, Luzp-KO/lacZ-KI(+/-) mice exhibit (i) normal spatial learning capability and memory retention (water maze), (ii) normal level of anxiety-related behavior pattern (elevated plus maze), (iii) normal emotional memory retention for 24 hr, 8 days, 14days and 28 days after foot shock pairing(contextual fear conditioning). Luzp-KO/lacZ-KI(+/-) mice were administrated with different dosages of NMDA receptor antagonist(MK801) intraperitoneally 30 mins before training in contextual fear conditioning. Luzp-KO/lacZ-KI(+/-) mice displayed similar phenotype as wildtype littermates under different dosages of MK801 treatment. Whether differential responses between wild type and Luzp-KO (+/-) will result if the test animals were pretreated with drugs or subjected to stress paradigms should await further investigation.
To further delineate the role LUZP plays in CA3 of hippocampus during stress; serum corticosterone (CT) level of test mice was monitored by RIA either after natural stress paradigms; i. e., forced swimming and immobilization, or after adrenalectomy (ADX). Correlating stress receptor immunocytochemistry of glucocorticoid receptor(GR-ICC) and corticotrophin-releasing factor receptor I (CRF-RI-ICC) in hippocampal CA3 were examined. Qualitative data from ICC and in situ X-gal staining and quantitative morphometric data were presented that the expression of Luzp(lacZ) 、GR and CRF RI were not changed when compared to control at ADX-13d and ADX-20d. Whether down regulation of LUZP in response to ADX (low CT) and high CT level are mediated via the same mechanism should await further investigation.
We treated mice with the glutamate analog, kainic acid (KA) to elicits seizure and the results indicated the down regulation of LUZP and GR in the same time window observed in the CA3 region. To investigate the role of Luzp in epileptic seizures, we treated heterozygous Luzp-KO/lacZ-KI mice and wildtype littermates with either kainic acid or pentylenetetrazol(PTZ), a GABA receptor antagonist, intraperitoneally and scored the degrees of seizure. The heterozygous Luzp-KO/lacZ-KI mice show a higher degree of seizure and died more frequently, after extensive convulsive seizures, at 25 mg kg-1 of kainic acid than the control groups. According to the behavioral trait, c-Fos expression pattern representing neuronal activity was compared between Luzp(+/-) and wildtype littermates 1.5hr post-seizure. The c-fos expression in heterozygotes is more stronger in the CA1 region than in wildtype littermates with similar degrees of KA-induced seizure. These results suggest a possible link between the gene dosage of LUZP and the susceptibility of epilepsy. The enhanced c-Fos expression in Luzp(+/-) mice may provide a direction for the future studies to explore the molecular targets and cellular pathways which LUZP involved in the regulation of kainic acid-induced epilepsy.
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