神經調節素-1的基因突變對成體嗅球神經細胞新生的影響

Abstract

成鼠腦室下區（subventricular zone）中的神經幹細胞藉由新生神經（neurogenesis）的過程，將神經成體細胞分化為神經母細胞，並沿著吻測遷移流（ rostral migratory stream） 到達嗅球（main olfactory bulb）補充因計畫性細胞死亡減少的中間神經元；之前的研究報告顯示，嗅球內中間神經元的改變會好發生在精神疾病的患者身上，尤其精神分裂症（schizophrenia），已經在許多研究指出有較高產生嗅覺功能喪失的可能性，但僅有少數的研究提出是和新生神經的過程有關 性。神經調節素-1（NRG1）是許多造成精神分裂症致病基因的其中一個，利用神經調節素-1突變的老鼠探討是否影響成鼠新生神經的過程，以瞭解神經調節素-1在新生神經上扮演的功能，造成而精神分裂患者嗅覺異常的原因。實驗過程中利用免疫螢光技術觀察腦室下區的背側吻側、吻側遷移流及嗅球中嗅小球旁細胞（periglomerular cell）三個區域，可以發現神經調節素-1突變後腦室下區的背側吻側產生較多個神經成體細胞及神經母細胞，給予bromodeoxyuridine（BrdU）後也相同在腦室下區的背側吻側觀察到被標定BrdU的神經成體細胞及神經母細胞個數增加，且吻側遷移流的頭側部的神經母細胞也比正常老鼠多，另外，嗅小球旁細胞中酪胺酸羥化酶（ tyrosine hydroxylase ） 增加， 但鈣離子結合蛋白（calbindin）減少，並同時在嗅球的西方墨點法中得到相同的結果，所以我們推測神經調節素-1基因的突變會不僅影響神經成體細胞在腦室下區的增生、也改變了在吻側遷移流移行及嗅球中間神經元的分化能力，並藉由型態學的研究結果可有助於釐清新生神經過程及精神分裂症之間的關係。

A region of the brain where adult neurogenesis takes place is in the subventricular zone (SVZ). Various types of interneurons in the olfactory bulbs are periodically replenished throughout adulthood to sustain neuronal homeostasis. The main supply of new interneurons arises from the neural stem cells in the SVZ. These neural progenitor cells (NPC) differentiate into neuroblasts (NB) and migrate in a chain-like formation through the rostral migratory stream (RMS) to the main olfactory bulbs (MOB), more specifically the granule cell layer (GCL) and the glomerular cell layer (GL). In previous studies, olfactory deficits, such as in odor perception impairment, may result from changes in the population of interneurons within the MOB. These olfactory deficits are some phenotypic indicators of psychological diseases, for example, schizophrenia. According to some studies, most schizophrenics have higher probability of acquiring olfactory deficits compared to non-psychotics. But, only a few studies propose correlations between adult neurogenesis and schizophrenia. Since neuregulin-1 (NRG1) is one of many schizophrenia susceptible genes, our study used transgenic mice with mutation in nrg1 gene to investigate adult neurogenesis. Immunofluorescence analysis of brain sections prepared from 7 to 9 weeks old wildtype (nrg1+/+) and mutant (nrg1+/-) NRG1 mice revealed that mutants tend to have 63% more in number of NPC (nestin) and 12% increase in migrating NB (doublecortin, DCX) in the dorsal-rostral part of SVZ. Consistent with the observations above, in the mutant NRG1 mice treated with 3 bromodeoxyuridine (BrdU), the number of newly generated NPC and NB in SVZ seemed to have increased 65% and 22% respectively. Interestingly, unlike the above, mutant NRG1 mice treated with BrdU appeared to exhibit 21% more newly produced NB in the rostral region (near the MOB) of the RMS. The western blot analysis results showed differences between wildtype and mutant NRG1 mice’s MOB in protein levels of tyrosine hydroxylase (TH), calretinin (CR), or calbindin (CB), but not GAD67, where expression level of TH and CR increased by 51% and 15.9% respectively, while CB expression level decreased by 53.2% after depletion of the ligand, NRG1. Immunofluorescence staining of different interneuron subtypes in the GL for TH and CB periglomerular (PG) cells are consistent with the results from western blot analysis with 33% increase for TH and 27% decrease for CB in cell numbers. We are suggesting the role of NRG1 is involved in proliferation of adult neural progenitor cells and distribution of migrating neuroblasts in the SVZ, RMS and MOB, as these are mostly precursors of MOB’s interneurons. Also, NRG1 may influence the differentiation of interneurons in the GL layer of MOB. This animal model for schizophrenia may contribute to the field in understanding the relationship between adult neurogenesis and pathogenesis of psychological diseases from cellular morphological standpoint.