用于嗅觉机理研究的MEMS微探针阵列

微探针芯片是基于微机电系统MEMS体加工技术的研究在体神经电生理或临床药理分析的新兴工具,主要应用动物脑区电生理的在体研究,可有助于在原位水平的网络系统中,了解神经元之间的信息耦合传导机制,探索神经元如何联系成整体来处理和储存信息等.首先分析细胞-微电极界面模型,然后介绍了原位式微探针的设计和工艺制作步骤,采用微探针对大鼠嗅球区附近和海马区的脑电位进行测试和分析,并通过对嗅粘膜施加气体刺激,观察嗅前核区电位响应变化,结果显示10-3M的甲基水杨酸盐对大鼠嗅上皮刺激时,会在嗅前核区产生响应.在此基础上,结合当前的研究工作,说明基于体加工MEMS技术的微探针阵列适合于在体神经元的多位点实时测量,对嗅觉传导机理的研究具有重要作用.     

基于微电极阵列的嗅球细胞网络传感器的研究

嗅球(OB)是嗅觉系统的第一中转站,在嗅觉信息的识别和处理中具有重要的作用.嗅球中具有多种类型的神经元,分别具有不同的生理特点和功能.本研究利用细胞培养技术,将嗅球神经元与微电极阵列(MEA)芯片耦合,构建一种细胞网络传感器,用于对多点的嗅球神经元电活动进行同步观察与分析.结果显示,MEA上培养的嗅球细胞生长良好,能够检测多个通道的嗅球神经元的自发电位以及谷氨酸作用下的诱发响应.研究表明,该嗅球细胞网络传感器能够实现信号的多通道同步检测及有效分辨神经元的自发信号和诱发响应,并且能够很好地捕捉不同通道神经元响应的特点.该研究对于进一步分析嗅觉信息在嗅球内的传导和编码具有重要的意义.     

仿生光学人工鼻及其呼吸气体检测的实验研究

针对呼吸气体诊断的需要，介绍了一种新型的具有仿生特点的光学人工鼻实验系统，用于探索通过呼吸气味诊断疾病的可行性。该系统由气敏光纤传感器阵列和信号处理系统组成，利用光纤传感器易与多种聚合物敏感材料相结合和其对气味的响应快、精度高以及易获得嗅觉动态信息的特点，通过计算化学方法，实现了多种聚合物敏感材料与荧光试剂的定量配制，研究了具有不同选择性的气味光纤传感器阵列。此外，将生物嗅觉机理模型用于人工鼻系统的设计过程，采用模拟生物嗅觉的感受器-嗅球-线性延迟神经网络算法，实现了从光纤传感器阵列的响应数据中同时分离出混合气味的成分和浓度信息。实现结果表明，该光学人工鼻传感器实验系统具有一定的仿生特性和应用前景。     

基于脊髓内功能电激励的大鼠后肢运动选择性控制

脊髓内功能电激励提供了一种恢复脊髓损伤患者运动功能的途径。本研究的目标是在大鼠脊髓上确定诱发后肢不同动作的区域,对支配相应动作的区域进行归一化描述,并验证脊髓内功能电激励能够用于后肢运动功能恢复。实验中利用了三维扫描式功能电激励技术对大鼠脊髓腰骶段进行刺激。结果表明,对腰骶段脊髓腹侧进行电刺激能够产生单关节或多关节的协调运动,合理选择刺激位点能够诱发后肢多种不同动作,且后肢运动向量图覆盖后肢运动的矢状平面的各个方向。本文绘制了大鼠脊髓运动功能图谱,对相关动作的控制区域进行归一化描述,为后续实验中电极植入位点的确定提供指导。     

电子鼻技术在医学中的应用

目的:探讨电子鼻技术方法及其在医学中的应用。方法:电子鼻是利用气体传感器阵列的响应图案来识别气味的电子系统,通常由气味取样器、气体传感器阵列和信号处理系统三部分组成。介绍了电子鼻的发展状况、论述了通用电子鼻的组成、原理及所用的传感器(金属氧化物传感器、导电型聚合物传感器、压电型传感器等)的特性及其医学检测中的应用。结果:电子鼻可用于检测肺部疾病(肺癌、肺结核)、糖尿病、尿毒症和细菌感染(厌氧菌、耳鼻喉感染细菌、不同类型的金黄葡萄球菌、伤口微生物感染)。结论:电子鼻作为一种无创的、快速诊断技术在疾病的早期诊断筛查及微生物感染的快速检测方面具有重要的临床应用价值。     

聚酰亚胺薄膜电极与大鼠视神经胶质细胞的生物相容性研究

为了研究视神经胶质细胞与聚酰亚胺薄膜电极的黏附性及生物相容性、最佳体外电脉冲刺激参数,为人工视觉假体材料的植入提供实验依据,本实验从大鼠神经胶质细胞的体外培养入手,采用细胞毒性试验、黏附试验等多种生物学方法对材料的生物相容性进行系统的评价,并建立电刺激大鼠神经胶质细胞体外模型,观察脉冲电流对神经胶质细胞生物学特性的影响。结果显示:聚酰亚胺薄膜材料的细胞毒性为0～1级,无明显细胞毒性。聚酰亚胺薄膜电极表面的细胞黏附生长良好,该材料对大鼠体外培养的细胞形态无损害,对细胞的生长和增殖均无明显的抑制作用;扫描电镜下观察到神经胶质细胞紧密贴附在材料表面,铺展良好并连接成片。在固定刺激脉冲的宽度和幅度以及刺激时间的情况下,频率越小,对细胞影响越小。以上研究结果提示聚酰亚胺薄膜电极与大鼠视神经胶质细胞有较好的黏附性和生物相容性,并能形成有效的电脉冲刺激,是一种优良的视神经植入材料。     

嗅觉模型的理论研究

嗅觉反应的主要机理包括以下五个连续的主要过程 :1 周围环境中的气味分子通过鼻腔中对流与扩散的双重作用 ,传输到嗅区 ;2 气味分子在嗅区的上方侧向传输到达嗅粘膜表面 ;3 气味分子被嗅粘膜表面的粘液所吸附 ;4 气味分子在嗅粘膜层中扩散 ,到达嗅细胞 ;5 气味分子的浓度在嗅细胞处达到浓度阈值 ,刺激嗅细胞放电 ,产生嗅觉。我们建立了揭示这五个主要过程的嗅觉模型 ,并通过理论分析得到模型的精确解。精确解定量揭示了嗅觉生理参数之间的内在关系 ,为进一步研究嗅觉机理提供了理论依据     

稳态视觉刺激频率与诱发响应相关性的fMRI研究

稳态视觉诱发电位(SSVEP)及脑电逆问题(inverse EEG)的研究显示了大脑皮层对于不同频率的稳态视觉刺激的响应具有幅度及空间位置上的差异.本文利用功能性磁共振成像(fMRI)研究稳态视觉刺激频率与诱发响应之间的相关性.根据SSVEP的实验结果进行真fMRI实验设计,对5名被试者给予不同频率的稳态视觉刺激,同时进行了脑部的fMRI扫描,采用SPM软件进行数据处理和分析.结果显示,不同频率刺激下的响应区域均集中于初级视皮层(V1),并且均具有单侧优势效应,即右侧半球的响应强于左侧半球.该结果显示了初级视皮层单侧优势对于稳态视觉诱发响应的表现形式.本文讨论了单侧优势与刺激性质之间存在相关性的可能,以及对此做进一步研究和应用的前景.     

糖尿病神经病理性疼痛大鼠脑异常活动的功能磁共振研究

目的　构建糖尿病神经病理性疼痛大鼠模型,利用功能磁共振成像技术,观察脑功能活动变化情况,探索异常功能活动的脑区。 方法　雄性SD大鼠48只,随机分为糖尿病组（n=36）和对照组（n=12）。糖尿病组通过腹腔注射60 mg/kg链脲佐菌素构建糖尿病模型,同时测定50％机械刺激缩足反射阈值的变化情况,根据触觉诱发性疼痛标准,糖尿病组大鼠被分成疼痛组及无痛组。利用锰离子增强功能磁共振成像技术进行大鼠脑功能成像,图像后处理分析糖尿病神经病理性疼痛大鼠脑内异常功能活动区域。 结果　成功构建糖尿病神经病理性疼痛大鼠模型,发现脑内存在显著性功能活动增强的区域,包括感觉皮层、腹内侧前额叶皮层、前扣带皮层、下橄榄核、梨状皮层、杏仁核及岛叶部分皮层。 结论　糖尿病神经病理性疼痛大鼠脑内部分脑区具有异常功能活动的表现,这些脑区很有可能参与该疼痛调控的中枢机制。     

自适应干扰对消的大鼠视觉诱发电位提取

研究大鼠的视觉诱发电位对认知人类视觉机理和疾病发生的位置具有重要意义.为获得大鼠视觉刺激诱发电位的特征信息,本研究设计了一种新的用于自适应干扰对消的大鼠视觉诱发电位信号采集方案,在大鼠初级视皮层不同位置和深度分别植入检测电极和干扰参考电极,有效提取了同源干扰信号.并基于该方案进行了大鼠视觉诱发电位的快速提取研究.仿真实验和实际应用结果表明,应用该方案实现自适应干扰对消,信噪比提高约6 dB,能够有效提取出大鼠单次闪光刺激诱发电位波形,并可从单次提取的结果中获得准确的潜伏期信息.该方案为实时获取大鼠视觉诱发电位特征信息提供了有效手段.     

Activity-induced manganese-dependent functional MRI of the rat visual cortex following intranasal manganese chloride administration

Manganese-enhanced MRI (MEMRI) of the brain requires delivery of manganese into the target brain regions. It was previously shown that, following intranasal application, ongoing olfactory stimulation facilitates manganese transport along the olfactory nerve into the olfactory bulb, so bypassing the blood–brain barrier (BBB). We report on experiments to evaluate whether visual stimulation can permit manganese transport onwards from the olfactory bulb to the visual cortex. Rats in intact olfactory bulb group were reserved intact olfactory bulb, while those in olfactory bulbectomy group received bilateral bulbectomy. After intranasal MnCl₂ administration, olfactory and visual stimulations were performed on all the animals for a consecutive 20 h. The visual cortex was then examined using MEMRI. Enhanced imaging on T1WI was noted in the visual cortex of the intact olfactory bulb group. Image subtraction revealed that the signal intensity in the visual cortex of the intact olfactory bulb group was significantly higher than that of olfactory bulbectomy group. Volume of interest (VOI) analysis also showed that normalized intensities in the visual cortex of the intact olfactory bulb group were significantly higher as compared with those of the olfactory bulbectomy group. Inductively coupled plasma mass spectrometry (ICP-MS) confirmed that the manganese content in the visual cortex of the intact olfactory bulb group was increased in comparison with that of the olfactory bulbectomy group. These findings indicate that activity-induced manganese-dependent functional MRI (AIM fMRI) of the rat visual cortex can be performed following intranasal administration of manganese and demonstrate that manganese can migrate from the olfactory bulb to the visual cortex.     

AMPA autoreceptors drive correlated spiking in olfactory bulb glomeruli

Information processing in the brain may rely on temporal correlations in spike activity between neurons. Within the olfactory bulb, correlated spiking in output mitral cells could affect the odor code by either binding or amplifying signals from individual odorant receptors. We examined the timing of spike trains in mitral cells of rat olfactory bulb slices. Depolarization of mitral cell pairs elicited spikes that were correlated on a rapid timescale (< or =10 ms) for cells whose primary dendrites projected to the same glomerulus. Correlated spiking was driven by a novel mechanism that depended on electrical coupling at mitral cell primary dendrites; the specific synchronizing signal was a coupled depolarization ( approximately 20 ms) that was mediated by dendritic AMPA autoreceptors. We suggest that glomerulus-specific correlated spiking in mitral cells helps to preserve the fidelity of odor signals that are delivered to the olfactory cortex.     

Neural correlates of conditioned odor avoidance in infant rats

Newborn rat pups can learn to either approach or avoid odor cues through associative conditioning. The present results demonstrate that preference conditioning and avoidance conditioning both modify olfactory bulb responses (focal 2-deoxyglucose uptake and mitral-tufted cell single unit responses) to the conditioned odor. Despite opposing behavioral responses to the conditioned odor, however, olfactory bulb neural responses did not detectably differ between learned odor cues signaling approach and those signaling avoidance. Control pups exhibited neither the behavioral nor neural changes. Furthermore, both the behavioral and neural changes to these odor cues could be extinguished. These results suggest that the olfactory bulb in neonates may code learned odor importance, but specific information attached to that importance may require processing in other brain regions.     

Dissociation of behavioral and neural correlates of early associative learning

Wistar rat pups were trained in an olfactory associative conditioning task on postnatal Day 6, 12, or 20. The training consisted of 20 pairings of a novel odor (peppermint) with footshock (1.5 mA, 1 s) with an intertrial interval of 3 min. Additional pups were trained in either unpaired or naive control conditions. On the day following training, pups were either tested for their behavioral response to the conditioned odor in a two-odor choice test, or injected with ¹⁴C-2-deoxyglucose and exposed to the odor for examination of olfactory bulb neural responses to the odor. The results demonstrate that, although pups at all ages learned to avoid the odor, only pups trained during the first postnatal week had a modified olfactory-bulb glomerular-layer response to the odor. These results suggest that although olfactory memory is correlated with modification of olfactory bulb glomerular layer function in newborns, these changes are not required for normal memory in older pups. © 1995 John Wiley & Sons, Inc.     

The olfactory conditioning in the early postnatal period stimulated neural stem/progenitor cells in the subventricular zone and increased neurogenesis in the olfactory bulb of rats

The olfactory memory acquired during the early postnatal period is known to be maintained for a long period, however, its neural mechanism remains to be clarified. In the present study, we examined the effect of olfactory conditioning during the early postnatal period on neurogenesis in the olfactory bulb of rats. Using the bromodeoxyuridine-pulse chase method, we found that the olfactory conditioning, which was a paired presentation of citral odor (conditioned stimulus) and foot shock (unconditioned stimulus) in rat pups on postnatal day 11, stimulated the proliferation of neural stem/progenitor cells in the anterior subventricular zone (aSVZ), but not in the olfactory bulb, at 24 h after the conditioning. However, the number of newborn cells in the olfactory bulb was increased at 2 weeks, but not 8 weeks, after such conditioning. Neither the exposure of a citral odor alone nor foot shock alone affected the proliferation of neural stem/progenitor cells in the aSVZ at 24 h after and the number of newborn cells in the olfactory bulb at 2 weeks after. The majority of newborn cells in the olfactory bulb of either the conditioned rats or the unconditioned rats expressed the neural marker NeuN, thus indicating that the olfactory conditioning stimulated neurogenesis in the olfactory bulb. These results suggest that olfactory conditioning during the early postnatal period temporally stimulates neurogenesis in the olfactory bulb of rats.     

Synchronized oscillatory discharges of mitral/tufted cells with different molecular receptive ranges in the rabbit olfactory bulb.

Individual glomeruli in the mammalian olfactory bulb represent a single or a few type(s) of odorant receptors. Signals from different types of receptors are thus sorted out into different glomeruli. How does the neuronal circuit in the olfactory bulb contribute to the combination and integration of signals received by different glomeruli? Here we examined electrophysiologically whether there were functional interactions between mitral/tufted cells associated with different glomeruli in the rabbit olfactory bulb. First, we made simultaneous recordings of extracellular single-unit spike responses of mitral/tufted cells and oscillatory local field potentials in the dorsomedial fatty acid-responsive region of the olfactory bulb in urethan-anesthetized rabbits. Using periodic artificial inhalation, the olfactory epithelium was stimulated with a homologous series of n-fatty acids or n-aliphatic aldehydes. The odor-evoked spike discharges of mitral/tufted cells tended to phase-lock to the oscillatory local field potential, suggesting that spike discharges of many cells occur synchronously during odor stimulation. We then made simultaneous recordings of spike discharges from pairs of mitral/tufted cells located 300-500 microm apart and performed a cross-correlation analysis of their spike responses to odor stimulation. In approximately 27% of cell pairs examined, two cells with distinct molecular receptive ranges showed synchronized oscillatory discharges when olfactory epithelium was stimulated with one or a mixture of odorant(s) effective in activating both. The results suggest that the neuronal circuit in the olfactory bulb causes synchronized spike discharges of specific pairs of mitral/tufted cells associated with different glomeruli and the synchronization of odor-evoked spike discharges may contribute to the temporal binding of signals derived from different types of odorant receptor.     

Odor specificity of habituation in the rat anterior piriform cortex

Exposure to odorants results in a rapid (<10 s) reduction in odor-evoked activity in the rat piriform cortex despite relatively maintained afferent input from olfactory bulb mitral cells. To further understand this form of cortical plasticity, a detailed analysis of its odor specificity was performed. Habituation of odor responses in anterior piriform cortex single units was examined in anesthetized, freely breathing rats. The magnitude of single-unit responses of layer II/III neurons to 2-s odor pulses were examined before and after a 50-s habituating stimulus of either the same or different odor. The results demonstrated that odor habituation was odor specific, with no significant cross-habituation between either markedly different single odors or between odors within a series of straight chain alkanes. Furthermore, habituation to binary 1:1 mixtures produced minimal cross-habituation to the components of that mixture. These latter results may suggest synthetic odor processing in the olfactory system, with novel odor mixtures processed as unique stimuli. Potential mechanisms of odor habituation in the piriform cortex must be able to account for the high degree of specificity of this effect.     

Optical imaging of odor preference memory in the rat olfactory bulb

Early olfactory preference learning in rat pups occurs when novel odors are paired with reinforcing tactile stimulation that activate the noradrenergic locus coeruleus. Pairing of odor and a noradrenergic agonist in the olfactory bulb is both necessary and sufficient for odor preference learning. This suggests the memory change occurs in the olfactory bulb. Previous electrophysiological experiments demonstrated that odor preference training induces an increase in the field excitatory postsynaptic potential to olfactory nerve input and an alteration, after training, in glomerular [14C]2- deoxyglucose uptake and in single-unit responses of principal cells. We investigate here whether, 24 h after olfactory preference training, there is an alteration in intrinsic optical signals at the glomerular level. Six-day-old rat pups were trained, as previously, for a peppermint odor preference. Trained pups and control littermates were subjected to imaging of odor-induced intrinsic optical signals 1 day after the training session. Trained pups exhibited significantly larger responses to the peppermint compared with untrained littermates previously exposed to the same odor. The response of trained pups to a control odor (amyl acetate) was, however, not significantly different from that of untrained littermates. These observations demonstrate that odor preference memory can be read-out by optical imaging techniques.     

Both electrical and chemical synapses mediate fast network oscillations in the olfactory bulb

Odor perception depends on a constellation of molecular, cellular, and network interactions in olfactory brain areas. Recently, there has been better understanding of the cellular and molecular mechanisms underlying the odor responses of neurons in the olfactory epithelium, the first-order olfactory area. In higher order sensory areas, synchronized activity in networks of neurons is known to be a prominent feature of odor processing. The perception and discrimination of odorants is associated with fast (20-70 Hz) electroencephalographic oscillations. The cellular mechanisms underlying these fast network oscillations have not been defined. In this study, we show that synchronous fast oscillations can be evoked by brief electrical stimulation in the rat olfactory bulb in vitro, partially mimicking the natural response of this brain region to sensory input. Stimulation induces periodic inhibitory synaptic potentials in mitral cells and prolonged spiking in GABAergic granule cells. Repeated stimulation leads to the persistent enhancement in both granule cell activity and mitral cell inhibition. Prominent oscillations in field recordings indicate that stimulation induces high-frequency activity throughout networks of olfactory bulb neurons. Network synchronization results from chemical and electrical synaptic interactions since both glutamate-receptor antagonists and gap junction inhibitors block oscillatory intracellular and field responses. Our results demonstrate that the olfactory bulb can generate fast oscillations autonomously through the persistent activation of networks of inhibitory interneurons. These local circuit interactions may be critically involved in odor processing in vivo.