5-HT2A受体在尼可刹米增强新生大鼠延髓脑片呼吸放电中的作用

本文旨在探讨中枢呼吸兴奋剂尼可刹米对新生大鼠基本节律性呼吸的产生和调节的影响及5-HT2A受体在其中的作用.制作新生大鼠离体延髓脑片标本,含面神经后核内侧区(the medial region of the nucleus retrofacialis,mNRF)并保留舌下神经根,灌流改良Kreb'S液(modified Kreb'S solution,MKS),记录舌下神经根呼吸相关节律性放电活动(respiratory-re-lated rhythmic discharge activity,RRDA),观察不同浓度尼可刹米、5-HT2A受体特异激动剂2,5-二甲氧基-4-碘苯基丙烷[1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane,DOI]、5-HT2A受体特异拮抗剂酮舍林(ketanserine)以及联合使用尼可刹米和酮舍林对舌下神经根RRDA的影响.结果显示,尼可刹米在0.5～7μg/mL时对延髓脑片RRDA有兴奋作用,在5 μg/mL时对吸气时程(inspiratory time,TI)、放电积分幅度(integral amplitude,IA)、呼吸周期(respiratory cycle,Re)等呼吸指标综合效果最显著.DOI明显延长TI、增强IA、缩短RC,对RRDA有兴奋作用.酮舍林明显缩短TI、减弱IA、延长RC,对RRDA有抑制作用.联合使用DOI和酮舍林对RRDA无明显作用.酮舍林可完全阻断尼可刹米对RC的作用,部分阻断尼可刹米对IA的作用,对尼可刹米引起的TI变化无明显影响.结果提示,尼可刹米增强新生大鼠离体延髓脑片舌下神经根RRDA,5-HT2A受体可能足尼可刹米作用途径之一.     

NMDA与非NMDA受体激动剂对新生大鼠延髓脑片呼吸节律性放电的作用

目的：探讨N－甲基－D－天门冬氨酸（NMDA）和非NMDA类受体在基本呼吸节律发生和调节中的可能作用。方法：以改良的Kerb's液灌流新生SD大鼠离体延髓脑片,记录片与之相连的舌下神经的呼吸节律性放电活动（respiratory rhythmical discharge activity,RRDA),在灌流中给予兴奋性氨式酸类递质及相应的拮抗剂,观察其对RRDA的影响。结果：使用非NMDA受体激动剂海人酸（KA）后,可见呼吸周期及呼吸时间有所延长,NMDA受体激动剂NMDA对呼吸活动则没有明显影响;相应的拮抗剂6－氰基－7－硝基喹恶啉土卫四（DNAX）和2－氨基酸戊酸（AP5）均可使放电频率和积分幅值明显降低,吸气时间显著缩短,但DNQX同时可致呼吸周期和呼气时间明显缩短。结论：在哺乳动物基本呼吸节律的产生和调节中,NMDA类受体主要对呼吸活动的强度产生调节作用;而非NMDA类受体不仅可以影响呼吸的强度,同时对呼吸的频率也发挥调节作用。     

cAMP-PKA通路参与Ⅱ组代谢性谷氨酸受体对新生鼠离体延髓脑片呼吸节律性放电的调节

本研究旨在探讨cAMP-PKA通路在Ⅱ组代谢性谷氨酸受体对离体延髓脑片呼吸节律性放电的影响中的作用。制作新生大鼠离体延髓脑片标本,主要包含延髓面神经后核内侧区(medial region of the nucleus retrofacialis,mNRF),并完整保留舌下神经根,以改良Kreb’s液(modified Kreb’s solution,MKS)恒温灌流脑片,用吸附电极记录舌下神经根呼吸节律性放电活动(respiratory rhythmical discharge activity,RRDA)。待放电活动稳定后,第1组灌流Ⅱ组代谢性谷氨酸受体特异性拮抗剂(2S)-α-ethylglutamic acid(EGLU)10min,第2组先给予cAMP-PKA通路激动剂Forskolin灌流10min,而后MKS洗脱至正常,灌流cAMP-PKA通路抑制剂Rp-cyclic3’,5’-hydrogen phosphorothioate adenosine triethylammonium salt(Rp-cAMPS)10min,第3组首先给予Rp-cAMPS10min,洗脱后联合Rp-cAMPS+EGL...     

兔延髓腹侧面化学敏感区对通气的作用

本工作探查了28只家兔延髓腹侧面化学敏感区的定位以及该区在机体对高 CO_2通气反应中的作用。用 pH 值为7.0、7.34和8.0的人工脑脊液滤纸敷贴在延髓腹侧表面观察通气变化。在锥体外方、第 Ⅶ一Ⅺ 对脑神经根部内侧和舌下神经根内侧两个区域用 pH7.0的滤纸敷贴时可使通气增加。反之,贴以浸有 pH 8.0的滤纸时可使通气减小。因此分别将此两区称为头端(R)和尾端(C)化学敏感区。在舌下神经根部上方,在 R 区和 C 区之间敷贴不同 pH 的滤纸则无通气变化。此区称为中间区(Ⅰ区)。PH7.34的滤纸敷贴以上三区均无通气变化。完整动物用 2%普鲁卡因麻醉 R 区和 C 区后,高 CO_2仍能引起通气增加,麻醉Ⅰ区后,高 CO_2引起通气减少。切断双侧颈动脉窦神经的动物,用2%普鲁卡因麻醉 R 区和 C 区,高 CO_2亦导致通气减少;麻醉Ⅰ区,则呼吸停止。上述结果表明,家兔延髓腹侧面存在着对称的化学敏感区。在机体对高 CO_2的通气反应中,Ⅰ区和 R、C 区都起着一定的作用。     

延髓腹外侧区微量注射L－NNA和SNP对大鼠血压、心率和肾交感神经活动的影响

在麻醉大鼠观察了向延髓腹外侧区微量注射ＮＯ合成酶抑制剂Ｎ－硝基左旋精氨酸（ＬＮＮＡ）和硝普钢（ＳＮＰ）对血压、心率和肾交感神经活动的影响,旨在探讨中枢左旋精氨酸－ＮＯ通路在动脉血压调节中的作用及其机制。实验结果如下：（１）向延髓腹外侧头端区（ＲＶＬＭ）注射Ｌ－ＮＮＡ后,平均动脉压（ＭＡＰ）升高,肾交感神经活动（ＲＳＮＡ）增强;心率（ＨＲ）减慢,但无统计学意义。ＭＡＰ和ＲＳＮＡ的变化持续３０ｍｉｎ以上;此效应可被预先静注左旋精氨酸所逆转。（２）向ＲＶＬＭ微量注射ＳＮＰ,ＭＡＰ降低,ＲＳＮＡ减弱;但ＨＲ的变化无统计学意义。（３）向延髓腹外侧尾端区（ＣＶＬＭ）注射Ｌ－ＮＮＡ,ＭＡＰ降低,ＨＲ减慢,ＲＳＮＡ减弱。（４）向ＣＶＬＭ微量注射ＳＮＰ,ＭＡＰ升高,ＲＳＮＡ增强,而心率无明显变化。以上结果表明,中枢左旋精氨酸－ＮＯ通路对延髓腹外侧部的神经元活动有调变作用。     

家兔延髓区域阻滞对呼吸的影响

实验在麻醉、切断迷走神经的家兔上进行。记录双侧膈神经放电和气管内压作为呼吸的指标。用尖端直径为70—100μm的玻璃微管接微量注射器,脑内注入1％普鲁卡因,双侧对称地阻滞闩前2.0—3.0mm,中线旁开2.0—2.5mm,背侧表面下3.5—4.5mm的区域后,38只兔都出现了可逆性呼吸停止。双侧性电损毁此区,则可造成不可逆性呼吸停止。以同法注入同量生理盐水,则对呼吸无影响。经组织学检查,此区范围相当于Meessen图谱的面后核内侧区——包括面后核(Nr.Ⅶ)内侧部、网状小细胞核(R·pc)腹侧部、网状巨细胞核(R·gc)背外侧部和外侧网状核(Rl)的内侧部分。用同样方法阻滞延髓头端区、孤束核区及疑核尾端区、对呼吸节律均无明显影响。用1％普鲁卡因局部麻醉延髓腹侧面的中枢化学感受器I(s)区,呼吸也无明显变化。结果提示,面后核内侧区的一些结构可能在形成呼吸节律中起重要作用。     

电刺激兔延髓腹侧化学敏感区和压力敏感区对呼吸、血压,的影响及其中枢递质机制

电刺激麻醉兔延髓腹侧化学敏感区头端区引起潮气量(V_T)增加,呼吸频率(f)增快;电刺激压力敏感区(中间区)则使V_T减小,f亦增快。弱刺激时,两者均产生降压反应;刺激增强可诱发双相或升压反应。在出现周期性呼吸时,电刺激化学敏感区可使呼吸节律正常化、V_T增大,而电刺激压力敏感区则导致呼吸暂停。电刺激压力敏感区时,吸气时间(TI)和呼气时间(T_E)均缩短,以T_E变化更明显;由于V_T减小和T_I缩短,V_T/T_I保持相对不变,提示吸气终止的中枢阈值降低。在准备刺激的相应局部预先应用阿托品,可使电刺激化学敏感区产生的通气增强效应翻转,而对电刺激压力敏感区引起的通气抑制无明显影响;用印防己毒素则可选择性消除电刺激压力敏感区的通气抑制和降压效应。本工作表明延髓腹侧存在两个不同的中枢机制,其中化学敏感区产生的通气增强与胆碱能系统有关;压力敏感区产生的通气减弱效应与GABA系统有关。     

心室内和心外膜应用腺苷对延髓PGL神经元电活动的影响

在35只切断两侧缓冲神经和迷走神经的麻醉大鼠,观察心室内注射腺苷和心外膜涂布腺苷对延髓腹外侧头端区PGL神经元自发电活动的影响。结果如下：（1）35只大鼠共记录到121个自发放电单位,平均放电频率为22．5±1．9spikes／s。（2）心室内冲击注射腺苷（0．5μmol/kg,0．1ml）时,BP先升（△1．7±0．2kPa,P＜0.001）后降（△4.6土0.5kPa,P＜0．001）,HR减慢（△126．5±12.3bpm,P＜0.001）;35个PGL神经元自发放电单位中,30个单位的放电频率由21．9士2．6增至29．2土3．4spikes／s（P＜0．001）,3个单位不变,2个单位减少。（3）心外膜涂布腺苷（20mmol/L）,动脉血压和心率的变化不明显,22个PGL神经元自发放电频率由18．8土1．9增至26．9土2．8spikes／s（P＜0．001）,3个单位的放电频率无变化。（4）静脉注射选择性腺苷A1受体拮抗8－cyclopentyl-1,3－dipropylxanthine（DPCPX,500μg／kg）可完全阻断腺苷对PGL神经元的兴奋效应。（5）在左右房室沟涂布85％酚或切除双侧星状神经节后,腺苷激活PGL神经元的效应即行消失。结果提示,腺苷可通过人受体激活心交感神经传入纤维,进而兴奋PGL神经元。     

Location of central respiratory chemoreceptors in the developing tadpole

The location of central respiratory chemoreceptors in amphibian larvae may change as the central chemoreceptive function shifts from driving gill to driving lung ventilation during metamorphosis. We examined this possibility in the in vitro brain stem of the pre- and postmetamorphic Rana catesbeiana tadpole by microinjecting hypercapnic artificial cerebrospinal fluid (aCSF) while recording fictive lung ventilation. The rostral and caudal brain stem were separately explored systematically using injections of 11 nl of aCSF equilibrated with 100% CO2 that transiently acidified a 500-microm region, producing a maximum reduction in pH of 0.23 +/- 0.06 at the site of injection. In postmetamorphic tadpoles, chemoreceptive sites were concentrated in the rostral compared with the caudal brain stem. No such segregation was observed in the premetamorphic tadpole. We conclude that, as in lung rhythmogenic function, respiratory chemosensitivity emerges rostrally in the amphibian brain stem during development.     

cAMP-PKA通路参与Ⅱ组代谢性谷氨酸受体对新生鼠离体延髓脑片呼吸节律性放电的调节

本研究旨在探讨cAMP-PKA通路在Ⅱ组代谢性谷氨酸受体对离体延髓脑片呼吸节律性放电的影响中的作用.制作新生大鼠离体延髓脑片标本,主要包含延髓面神经后核内侧区(medial region of the nucleus retrofacialis,mNRF),并完整保留舌下神经根,以改良Kreb's液(modified ...     

Effects of Bilobalide on Hypoxia/Hypoglycemia-Stimulated Glutamate Efflux from Rat Cortical Brain Slices

The aim of this study was to investigate the effects of bilobalide, the postulated active constituent of Ginkgo biloba, on the release of glutamate elicited by hypoxia/hypoglycemia. Cortical slices were prepared from rat brain and perfused with normal artificial cerebrospinal fluid (aCSF) or aCSF made hypoxic by gassing with nitrogen, and hypoglycemic by removal of glucose. The perfusate was assayed for glutamate by HPLC. After 30 minutes, perfusion with hypoxic/hypoglycemic aCSF glutamate levels in the perfusate were increased approximately 5-fold. Bilobalide at 1, 10, and 100 M, when perfused together with hypoxic/hypoglycemic aCSF, significantly reduced the release of glutamate. This study suggests that the reported neuroprotective properties of bilobalide may, in part, be mediated through its ability to reduce glutamate efflux, thus leading to a decrease in the excitotoxic effects of this neurotransmitter.     

一氧化氮对呼吸节律性放电的调节作用

实验旨在探讨一氧化氮(nitric oxide,NO)在基本呼吸节律产生和调节中可能的作用。制作新生大鼠离体延髓脑片标本,主要包含面神经后核内侧区,前包钦格复合体、腹侧呼吸组以及背侧呼吸组的一部分。同时保留舌下神经根,用改良Kreb′s液灌流脑片并记录与之相连的舌下神经根呼吸节律性放电(respiratory rhythmical discharge activity,RRDA),在灌流液中分别给予不同浓度的NO供体硝普钠(sodium nitroprusside,SNP),NO合成前体L—精氨酸(L—Arginine,L-Arg)以及神经元型一氧化氮合酶(neuronal nitric oxide synthase,nNOS)特异性抑制剂7-nitro indazole (7-NI),观察其对RRDA的影响。结果显示,nNOS的特异性抑制剂7-NI对吸气时程和放电强度有明显抑制,而NO合成前体L—Arg,以及NO供体SNP对呼吸放电活动没有明显的影响。这提示,在哺乳动物基本呼吸节律的产生和调节中,NO可能对吸气中止和呼吸幅度具有调节作用。     

Glial Cells are Involved in the Exciting Effects of Doxapram on Brainstem Slices In Vitro

This study tested whether the glial cells are involved in the exciting effects of doxapram on brainstem slice in vitro. Experiments were performed in brainstem slice preparations from neonatal rats. The medial area of nucleus retrofacialis (mNRF) and the hypoglossal nerve (XII nerve) were contained in the preparations. The slices were perfused with modified Kreb’s solution (MKS), and the rhythmical respiratory discharge activity (RRDA) was simultaneously recorded from the XII nerve by using suction electrodes, including the discharge time course of inspiratory (Ti), expiratory (Te), respiratory cycle (RC), and integrity amplitude of inspiratory discharge (IA). After applying of doxapram (5 μM) to the MKS for 10 min, Ti and IA increased significantly (85.0 ± 25.0%, 13.2 ± 2.5%, respectively, P < 0.05), the Te and the RC decreased significantly (19.0 ± 1.4%, 12.8 ± 1.4%, respectively, P < 0.05) when compared with control group. When the methionine sulfoximine (MS, 10 μM), a blockage of glutamine synthetase, was applied, all the exciting effects of doxapram on RRDA were reversed. After the glutamine (20 μM) was applied to the MKS for 10 min, the exciting effects were revealed again. Our results suggest that the normal metabolism of glial cells takes a key role in the modification of the RRDA in the slices. In conclusion, glial cells are involved in the exciting effects of doxapram on brainstem slice in vitro.     

Cryopreservation of rat hippocampal slices by vitrification

Although much interest has attended the cryopreservation of immature neurons for subsequent therapeutic intracerebral transplantation, there are no reports on the cryopreservation of organized adult cerebral tissue slices of potential interest for pharmaceutical drug development. We report here the first experiments on cryopreservation of mature rat transverse hippocampal slices. Freezing at 1.2 degrees C/min to -20 degrees C or below using 10 or 30% v/v glycerol or 20% v/v dimethyl sulfoxide yielded extremely poor results. Hippocampal slices were also rapidly inactivated by simple exposure to a temperature of 0 degree C in artificial cerebrospinal fluid (aCSF). This effect was mitigated somewhat by 0.8 mM vitamin C, the use of a more "intracellular" version of aCSF having reduced sodium and calcium levels and higher potassium levels, and the presence of a 25% w/v mixture of dimethyl sulfoxide, formamide, and ethylene glycol ("V(EG) solutes"; Cryobiology 48, pp. 22-35, 2004). It was not mitigated by glycerol, aspirin, indomethacin, or mannitol addition to aCSF. When RPS-2 (Cryobiology 21, pp. 260-273, 1984) was used as a carrier solution for up to 50% w/v V(EG) solutes, 0 degree C was more protective than 10 degrees C. Raising V(EG) concentration to 53% w/v allowed slice vitrification without injury from vitrification and rewarming per se, but was much more damaging than exposure to 50% w/v V(EG). This problem was overcome by using the analogous 61% w/v VM3 vitrification solution (Cryobiology 48, pp. 157-178, 2004) containing polyvinylpyrrolidone and two extracellular "ice blockers." With VM3, it was possible to attain a tissue K(+)/Na(+) ratio after vitrification ranging from 91 to 108% of that obtained with untreated control slices. Microscopic examination showed severe damage in frozen-thawed slices, but generally good to excellent ultrastructural and histological preservation after vitrification. Our results provide the first demonstration that both the viability and the structure of mature organized, complex neural networks can be well preserved by vitrification. These results may assist neuropsychiatric drug evaluation and development and the transplantation of integrated brain regions to correct brain disease or injury.     

家兔面神经后核内侧区在呼吸节律起源中的作用

从腹侧面暴露家兔延髓,脑内微量注射1%普鲁卡因阻滞面神经后核内侧区(mNRF),全部动物(n=20)一次注射(0.3—1.0μl)后即能可逆地消除呼吸节律。区域对照显示此区非常局限,范围约1.0×1.0×1.0mm。组织学检查表明为面神经后核内侧区。本文分析了 mNRF的呼吸相关神经元(RRNs)的放电形式。在 mNRF 有较多的呼气(E)神经元和呼气-吸气跨时相(E-IPS)神经元。在阻滞 mNRF 引起呼吸停止期间,观察到低位延髓背侧呼吸群(DRG)和腹侧呼吸群(VRG)尾端区 RRNs 放电的节律性消失,表现连续放电或停止放电。电刺激DRG,VRG 尾端区,只能诱发短串的膈神经放电,而不能产生节律性发放。说明这些区域的RRNs 无自动节律性活动的能力。结果表明,面神经后核内侧区与呼吸节律发生有关,它可能是呼吸节律发生器的一个重要的所在部位。     

组胺H1和H2受体在新生大鼠离体延髓脑片呼吸节律性放电调节中的作用

本研究探讨组胺H1和H2受体在新生大鼠基本节律性呼吸的发生和调节中的作用.以改良的Kreb's液恒温灌流新生Sprague-Dawley大鼠离体延髓脑片标本,稳定记录与之相连舌下神经根的呼吸节律性放电活动(respiratory-related rhythmical discharge activity, RRDA).实验分为5组:第1、2、3组分别单独给予组胺(histamine, HA)、 H1受体特异阻断剂pyrilamine和H2受体特异阻断剂cimetidine;第4组分别先后给予HA和HA pyrilamine;第5组分别先后给予HA和HA cimetidine,观察舌下神经根RRDA的变化.结果显示,单独给予HA后呼吸周期(respiratory cycle, RC)及呼气时程(expiratory time, TE)明显缩短,而吸气时程(inspiratory time, TI)及放电积分幅度(integral amplitude, IA)无明显变化;给予pyrilamine后RC、 TE明显延长,TI、 IA也无明显变化,且HA的作用可以被pyrilamine逆转;给予cimetidine后RC、 TE、 TI、 IA均无明显变化,且HA的作用不能被cimetidine逆转.结果提示,H1受体参与哺乳动物基本呼吸节律的产生和调节,H2受体对哺乳动物基本节律性呼吸的调控无明显影响.     

Hypoxia/hypoglycemia-induced amino acid release is decreased in vitro by preconditioning

The aim of this study was to investigate the effects of preconditioning on amino acid neurotransmitter release, induced by hypoxia/hypoglycaemia, from rat brain cortical slices. Tissue, perfused with artificial cerebrospinal fluid (aCSF) at 37 degrees C with zero glucose and gassed with 95% nitrogen and 5% carbon dioxide, showed a fivefold increase in glutamate release with little effect on gamma-aminobutyric acid (GABA) release. Preconditioning, with three 5-min periods of hypoxia/hypoglycaemia preceding continuous hypoxia/hypoglycaemia, significantly decreased glutamate release whilst significantly elevating GABA release. These results suggest that GABA may reduce the release of glutamate and consequently decrease the neurotoxic effects of glutamate.