A new surgical approach to the study of vomeronasal system regeneration.
A new surgical approach to the study of vomeronasal system regeneration.
复制标题
研究犁鼻系统再生的新手术方法。
DOI：
10.1093/chemse/bjh148
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发表时间：
2005
期刊：
影响因子：
3.5
通讯作者：
Costanzo,RichardM
中科院分区：
文献类型：
--
作者：
Matsuoka,Masato;Norita,Masao;Costanzo,RichardM
犁鼻系统，包括犁鼻器、副嗅球和高级中枢，其功能是检测与社会和生殖行为相关的物质（Wysocki，1979；Halpern，1987）。犁鼻上皮的结构与嗅觉上皮相似，由受体神经元、支持细胞和前体细胞组成(Matsuoka et al., 2000)。犁鼻感觉细胞的轴突终止于副嗅球，而嗅觉感觉细胞的轴突直接投射到主嗅球（Matsuoka 等，1998；Rodriguez 等，1999）。在啮齿动物中，副嗅球位于主嗅球的后尾区。嗅觉和犁鼻系统为研究神经变性和再生提供了理想的模型，因为在成年动物的发育过程中和正常生理条件下，嗅觉和犁鼻神经元会发生持续的神经发生（Moulton，1974；Graziadei 和 Monti-Graziadei，1978）。然而，在受伤后观察到感觉细胞数量的急剧增加。先前的研究已经检查了不同损伤方法后两个系统的再生和恢复，包括去除嗅球和副嗅球、感觉上皮的化学损伤和神经感觉纤维的横断（Costanzo和Graziadei，1983；Samanen和Forbes，1984；Costanzo，2000；Matsuoka等，2002）。不同的手术方法有其优点和缺点。在筛板处切断嗅觉神经的方法也切断了犁鼻神经。使用目前的方法不可能在筛板水平选择性横断犁鼻神经。去除嗅球会导致犁鼻系统完全退化。然而，这种方法是不可逆的，因为没有留下目标组织供再生神经重新建立功能连接并实现完全恢复。为了研究犁鼻系统功能和恢复，需要一种方法来选择性地切断犁鼻神经，同时保持嗅觉系统完好无损。在这项研究中，我们试图建立一种仅选择性切断犁鼻神经的新方法。使用成年雄性小鼠，我们开发了一种新的手术方法，在副嗅球前方的一点切断犁鼻神经。麻醉后，嗅球上方的额骨被移除。这为嗅球和额叶皮层提供了良好的通道。然后，我们使用 vannis 微型解剖剪刀在嗅球和额叶皮层之间的边界处切割犁鼻神经。接下来，我们在术后 6、20、60 和 120 天检查了主嗅球和副嗅球内的犁鼻上皮、嗅上皮及其感觉神经末梢。为了研究犁鼻神经元的变性和再生，我们使用了一种称为嗅觉标记蛋白（OMP）的嗅觉细胞标记物。 OMP 是一种酸性蛋白，在嗅觉和犁鼻神经元（包括细胞体、轴突和神经末梢）中表达。尽管其功能尚不清楚，但 OMP 已被广泛用作嗅觉系统中成熟化学感应细胞的标记物（Margolis，1980）。
The vomeronasal system, including the vomeronasal organ, the accessory olfactory bulb and higher centers, functions to detect substances associated with social and reproductive behavior (Wysocki, 1979; Halpern, 1987). The vomeronasal epithelium has a structure similar to that of the olfactory epithelium, consisting of receptor neurons, supporting cells and precursor cells (Matsuoka et al., 2000). The axons of vomeronasal sensory cells terminate in the accessory olfactory bulb, while axons of olfactory sensory cells project directly to the main olfactory bulb (Matsuoka et al., 1998; Rodriguez et al., 1999). In rodents, the accessory olfactory bulb is located at the posterocaudal region of the main olfactory bulb. The olfactory and vomeronasal systems provide ideal models for the study of neural degeneration and regeneration, because a continual neurogenesis of olfactory and vomeronasal neurons occurs during development and under normal physiological conditions in adult animals (Moulton, 1974; Graziadei and Monti-Graziadei, 1978). However, more dramatic increases in the number of the sensory cells have been observed after injury. Previous studies have examined regeneration and recovery of both systems following different methods of injury, including removal of the olfactory and accessory olfactory bulbs, chemical lesion of the sensory epithelium and transection of nerve sensory fibers (Costanzo and Graziadei, 1983; Samanen and Forbes, 1984; Costanzo, 2000; Matsuoka et al., 2002). Different surgical methods have their advantages and disadvantages. Methods developed to cut the olfactory nerves at the cribriform plate also cut the vomeronasal nerves. Selective transection of the vomeronasal nerve at the level of the cribriform plate is not possible using current methods. Removal of the olfactory bulbs provides complete degeneration in the vomeronasal system. However, this method is not reversible, since there is no target tissue remaining for the regenerated nerves to reestablish functional connections and enable complete recovery. To study vomeronasal system function and recovery, a method is needed to selectively cut the vomeronasal nerves while leaving the olfactory system intact. In this study, we tried to establish a new method to selectively cut only the vomeronasal nerves.Using adult male mice, we developed a new surgical approach that cuts the vomeronasal nerves at a point just anterior to the accessory olfactory bulb. After anesthesia, the frontal bone over the olfactory bulb is removed. This provides good access to the olfactory bulbs and frontal cortex. We then cut the vomeronasal nerve at the borderline between the olfactory bulb and the frontal cortex using vannis micro dissecting scissors. Next we examined the vomeronasal epithelium, the olfactory epithelium and their sensory nerve terminals within the main and accessory olfactory bulb at 6, 20, 60 and 120 days after surgery. In order to study the degeneration and regeneration of vomeronasal neurons, we used an olfactory cell marker called the olfactory marker protein (OMP). OMP is an acidic protein which is expressed in both olfactory and vomeronasal neurons including cell bodies, axons and nerve terminals. Although its function is unknown, OMP has been used extensively as a marker for mature chemosensory cells in the olfactory system (Margolis, 1980).