脑桥呼吸调整中枢向延髓Botzinger复合体轴突投射的研究

实验在３３只成年猫上进行。Ｂｏｔｚｉｎｇｅｒ复合体内微量注入麦角辣根过氧化酶（ＷＧＡ－ＨＲＰ,３０－６０ｎｌ,５％,９例）后,在脑桥呼吸调整中枢结合臂外侧核及Ｋｏｌｌｉｋｅｒ－Ｆｕｓｅ核观察到大量ＨＲＰ标记神经元。在２０例动物中,检测了９１个在呼吸调整中枢记录到的呼吸神经元对电刺激Ｂｏｔｚｉｎｇｅｒ复合体的反应。其中１３个神经元（吸气性１１个,跨时相２个）可被逆行兴奋。实验结果表明,发自脑桥呼吸调整中枢神经元的轴突可投射到Ｂｏｔｚｉｎｇｅｒ复合体,这一投射通路可能与呼吸调节有关。     

用微电泳方法给予吗啡对家兔脑桥结合臂旁内侧核区呼吸性单位活动的影响

在家兔延髓曾观察到,孤束区和疑核区的呼吸性单位对微电泳给予吗啡的敏感性并不相同,本实验进一步观察微电泳给予吗啡对脑桥呼吸调整中枢(结合臂旁内侧核区)呼吸性单位放电的影响。在67个呼吸性单位中,微电泳给予吗啡引起兴奋的有40个,阻遏的4个,其余23个活动无明显改变。在18个吗啡引起兴奋的呼吸性单位中,有12个单位的吗啡效应可被纳洛酮对抗。6个微电泳给予吗啡引起兴奋的呼吸性单位,在静脉给予吗啡时也出现兴奋效应;这种兴奋效应能被微电泳或静脉给予纳洛酮所对抗。本工作说明,脑桥呼吸调整中枢的呼吸性单位对吗啡很敏感,而且主要起兴奋反应。     

KF核及Btzinger复合体内GABA能神经元向膈神经核的投射

实验在 6只成年猫上进行。将WGA HRP微量注入C5膈神经核内 ,通过逆行追踪及GABA免疫组织化学FITC荧光双重标记方法 ,研究了脑干内GABA能神经元向膈神经核的投射。结果在脑桥KF核和面神经后核周围区 (即B¨otzinger复合体 )观察到GABA HRP双标神经元。另外 ,在中缝大核、旁巨细胞外侧核及前庭神经核也观察到双标神经元。本实验结果表明 :发自上述脑干神经核团 ,特别是KF核及B¨otzinger复合体的GABA能神经元的轴突可投射到膈神经核     

家兔脑桥臂旁内侧核区微量注射吗啡对中缝大核区单位放电的影响

实验在62只家兔上进行。结果观察到,中缝大核(NRM)区562个单位中,有118个单位的自发放电频率低,放电比较规则,动作电位时程长,易被微电泳5-羟色胺所阻遏,称为A 组单位。其余444个单位的自发放电频率高,动作电位时程短,称为B 组单位。大多数 B组单位对微电泳5-羟色胺不起反应。脑桥臂旁内侧核(NPBM)区微量注射吗啡(200μg/2μl)或静脉注射吗啡(3mg/kg)后,20个A 组单位中有19个发生兴奋效应,而49个B 组单位中仅有29个发生兴奋效应,而且A组单位发生兴奋的程度也比B组单位的高。这些结果提示,NRM区的A 组单位可能是5-羟色胺神经元,吗啡对这些神经元有相对选择性的兴奋作用。 在另外11只家兔上,应用辣根过氧化物酶(HRP)逆行追踪技术观察到,NPBM 区与NRM 区有纤维联系。 本实验结果提示,静脉注射吗啡所致的呼吸抑制,可能与吗啡作用于 NPBM,通过纤维联系,引起NRM 5-羟色胺神经元兴奋有关。     

KF核及B(o)tzinger复合体内GABA能神经元向膈神经核的投射

实验在６只成年猫上进行。将ＷＧＡ－ＨＲＰ微量注入Ｃ５膈神经核内,通过逆行追踪及ＧＡＢＡ免疫组织化学ＦＩＴＣ荧光双重标记方法,研究了脑干内ＧＡＢＡ能神经元向膈神经核的投射。结果在脑桥ＫＦ核和面神经后核周围区（即Ｂｏｔｚｉｎｇｅｒ复合体）观察到ＧＡＢＡ－ＨＲＰ双标神经元。另外,在中缝大核、旁巨细胞外侧核及前庭神经核也观察到双标神经元。本实验结果表明：发自上述脑干神经核团,特别是ＫＦ核及Ｂｏｔｚｉｎｇｅ     

兔颈交感神经干动作电位的呼吸性节律

(一)兔颈交感神经干的动作电位和膈神经发放有明显的同步关系,是为交感发放的呼吸性节律。改变呼吸中枢和血管运动中枢的兴奋状态后,交感发放的呼吸性节律也随着发生相应的变化,但切断缓冲神经不影响交感的呼吸性节律发放。(二)必须保留脑桥和延髓完整,交感发放才能表现出呼吸性节律。在脑桥水平切断脑干而动物发生长吸式呼吸时,交感发放的呼吸性节律消失。(三)本文讨论了交感神经和膈神经同步化发放的机制,认为呼吸中枢的兴奋过程使血管运动中枢的活动被周期性地“强化”。于是交感发放出现呼吸性节律。呼吸中枢对血管运动中枢的“强化”的程度,乃取决于这两个中枢的兴奋性的高低,这一过程可能是在脑桥和延髓的网状结构中进行的。     

微电泳法给予吗啡对家兔延髓疑核区呼吸性单位放电的影响

用五管微电极在家兔疑核区记录单位活动并观察微电泳吗啡时对单位活动的影响。在54个呼吸性单位中,吗啡引起阻遏的有10个,兴奋的1个,其余43个活动无明显改变。在6个吗啡引起阻遏的单位中,有5个单位的吗啡效应可被纳洛酮对抗。在49个非呼吸性单位中,吗啡引起阻遏的有13个,兴奋的有10个,其余26个活动无明显改变。在6个吗啡引起阻遏的单位中,有5个单位的吗啡效应可被纳洛酮对抗;在7个吗啡引起兴奋的单位中,吗啡的兴奋效应均不能被纳洛酮对抗。本实验观察到疑核区的呼吸性单位有一部分能被吗啡阻遏,但呼吸性单位对谷氨酸和吗啡敏感的比例均明显地比非呼吸性单位的比例低。     

中缝大核在臂旁内侧核区注射吗啡所致呼吸抑制效应中的作用

实验在33只浅麻醉、肌肉麻痹、人工呼吸及切断双侧颈迷走神经的家兔上进行。观察中缝大核区电解损毁或微量注射利多卡因对呼吸活动及臂旁内侧核区微量注射吗啡所致呼吸抑制效应的影响。结果是:电解损毀中缝大核区,使呼吸频率增加,膈神经放电的幅度和频率均无明显变化,而臂旁内侧核区微量注射吗啡抑制呼吸的程度减轻;中缝大核区微量注射利多卡因,则部分消除臂旁内侧核区微量注射吗啡的呼吸抑制效应。中缝大核旁网状结构电解损毁或微量注射利多卡因,不影响吗啡的呼吸抑制效应。上述结果提示,中缝大核区可能在脑桥臂旁内侧核区微量注射吗啡抑制呼吸的机制中起一定作用。     

回苏灵兴奋家兔呼吸的机制分析

本实验在29只局部麻醉、肌肉麻痹的家兔,用电生理方法研究了呼吸复苏剂回苏灵(dime-fline,DIM)兴奋呼吸的机制。在20例完整动物,静脉注射 DIM 使膈神经放电增加,出现兴奋效应。有效剂量为0.05—0.3mg/kg。在上丘水平去大脑后,同样剂量的 DIM 也能出现类同效应。横断脑干使动物处于长吸或喘息状态下,静脉注射 DIM 都能使膈神经放电增加,并加快呼吸频率,调整呼吸型式接近于正常。在11例动物的延髓孤束核区,用微电泳给予 DIM、GABA 及 BIG(bicuculline),观察了60个单位的放电变化,看到 DIM 能使 GABA敏感单位的放电出现兴奋、阻遏及先兴奋后阻遏等不同效应。在36个 GABA 敏感单位(其中吸气相关单位27个,非呼吸相关单位9个)中,DIM 能对抗 GABA 效应的有32个(其中吸气相关单位24个,非呼吸相关单位8个),不能对抗的有4个(其中吸气相关单位1个,非呼吸相关单位3个)。在13个微电泳 DIM 能对抗 GABA 效应的单位中,静脉注射小剂量 DIM可使其出现兴奋效应,并可对抗 GABA 效应。以上结果提示,DIM 对膈神经放电的兴奋效应,可能是阻断了脑干呼吸中枢的内源性 GABA 对呼吸与非呼吸相关单位的抑制作用的结果。     

大鼠下丘脑-缰核系统对中缝大核痛反应神经元放电的影响

在大鼠尾部给以伤害性刺激后,外侧缰核和中缝大核的单位按其反应型式可分为四种类型,即痛兴奋单位、广动力型单位、痛抑制单位和无反应单位。电刺激下丘脑外侧区对外侧缰核中各种单位的自发放电主要产生抑制作用,对其中痛兴奋单位和痛抑制单位的自发放电尤为明显。刺激下丘脑外侧区对中缝大核中痛兴奋单位的自发放电有明显兴奋作用,刺激外侧缰核则有抑制作用,损毁外侧缰核后,下丘脑外侧区的兴奋作用消失。分别刺激下丘脑外侧区和外侧缰核对中缝大核中痛抑制单位的自发放电都有明显的抑制作用;损毁外侧缰核后下丘脑外侧区的抑制作用仍存在。以上结果提示,下丘脑外侧区影响中缝大核活动的途径有二。其一可能是通过去除外侧缰核对中缝大核中痛兴奋单位的紧张性抑制作用;另外还可能通过外侧缰核以外的途径抑制中缝大核中痛抑制单位的活动。     

G protein-coupled receptor kinase 2 mediates mu-opioid receptor desensitization in GABAergic neurons of the nucleus raphe magnus

Nucleus raphe magnus (NRM) sends the projection to spinal dorsal horn and inhibits nociceptive transmission. Analgesic effect produced by mu-opioid receptor agonists including morphine partially results from activating the NRM-spinal cord pathway. It is generally believed that mu-opioid receptor agonists disinhibit spinally projecting neurons of the NRM and produce analgesia by hyperpolarizing GABAergic interneurons. In the present study, whole-cell patch-clamp recordings combined with single-cell RT-PCR analysis were used to test the hypothesis that DAMGO ([D-Ala(2),N-methyl-Phe(4),Gly-ol(5)]enkephalin), a specific mu-opioid receptor agonist, selectively hyperpolarizes NRM neurons expressing mRNA of glutamate decarboxylase (GAD(67)). Homologous desensitization of mu-opioid receptors in NRM neurons could result in the development of morphine-induced tolerance. G protein-coupled receptor kinase (GRK) is believed to mediate mu-opioid receptor desensitization in vivo. Therefore, we also investigated the involvement of GRK in mediating homologous desensitization of DAMAMGO-induced electrophysiological effects on NRM neurons by using two experimental strategies. First, single-cell RT-PCR assay was used to study the expression of GRK2 and GRK3 mRNAs in individual DAMGO-responsive NRM neurons. Whole-cell recording was also performed with an internal solution containing the synthetic peptide, which corresponds to G(betagamma)-binding domain of GRK and inhibits G(betagamma) activation of GRK. Our results suggest that DAMGO selectively hyperpolarizes NRM GABAergic neurons by opening inwardly rectifying K(+) channels and that GRK2 mediates short-term homologous desensitization of mu-opioid receptors in NRM GABAergic neurons.     

Changes in the background activity of neurons of the central gray substance when serotonin is applied to it or its synthesis is blocked

The changes in the background activity (BA) of neurons of the central gray substance upon the iontophoretic application of serotonin (ST) to them in a concentration of 1·10^–4 mole/liter, and following the blockage of its synthesis by pchlorophenylalanine (PCPA), were investigated in hexenal-anesthetized rats. The application of ST led to changes in the BA only in group III neurons, in which the BA was distinguished by continuous action potential generation. The application of ST elicited an increase in the frequency of the BA in 21.2% of such neurons, suppression in 45.5%, and the BA remained unchanged in 33.3% of the units. The identified changes in the BA in these neurons coincided with those which arose as the result of stimulation of the monoaminergic structures. When the application of ST was combined with stimulation of the locus coeruleus (LC), the substantia nigra (SN), and the nucleus raphe magnus (NRM), the suppression of the BA was intensified; however, when the LS and the SN were stimulated, the suppression was more pronounced than with stimulation of the NRM. The intensity of the suppressant effect of the NRM decreased after preliminary administration of PCPA, but it did not disappear completely, and the influences elicited by the LS and the SN intensified. The mechanisms of the serotoninergic control of the activity of the neurons of the central gray substance are discussed.A. A. Bogomolets Institute of Physiology, Ukrainian Academy of Sciences, Kiev. Translated from Neirofiziologiya, Vo. 24, No. 2, pp. 169–177, March–April, 1992.     

Relationship of glutamate and aspartate to the periaqueductal gray-raphe magnus projection: analysis using immunocytochemistry and microdialysis.

This study tested the hypothesis that the excitatory amino acid transmitters glutamate and/or aspartate are associated with the periaqueductal gray (PAG)-raphe magnus (NRM) projection. Retrograde neuroanatomical tracing procedures utilizing the tracers WGA-HRP or D-[3H]-aspartate were combined with immunocytochemical localization of glutamate or aspartate to determine if glutamate and/or aspartate immunostained neurons projected to the NRM. Both glutamate- and aspartate-immunoreactive cells in the PAG were found to project to the NRM. Double labeling immunocytochemichemical procedures indicated that glutamate and aspartate are co-localized in many PAG neurons, suggesting the following possibilities: (a) one of these two amino acids may serve as a precursor to the other; (b) both amino acids may be co-released from the same PAG neuron; or (c) both amino acids are present in high levels in the perikarya for metabolic purposes. At the EM level, both glutamate- and aspartate-immunoreactive terminals were identified in the NRM, strengthening the concept that both amino acids participate in synaptic transmission in this medullary nucleus. To determine if glutamate and aspartate are in fact released from PAG-NRM axons, the PAG was stimulated chemically with homocysteic acid (HCA) and amino acids were collected from the NRM using a microdialysis probe. Microinjection of HCA, but not vehicle, into the PAG resulted in the release of both glutamate and aspartate in the nucleus raphe magnus. These data suggest that both glutamate and aspartate are released from PAG fibers terminating in the NRM and provide strong support for the hypothesis that excitatory amino acids play a neurotransmitter role in the PAG-NRM pathway.     

Modulation of neuron activity of the midbrain periaqueductal gray matter influenced by monoaminergic brainstem structures

A study was done on base activity and changes in base activity (BA) of neurons in periaqueductal gray matter (PAG) during stimulation of monoaminergic structures of the brainstem: the nucleus raphe magnus (NRM), the locus coeruleus (LC), and the substantia nigra (SN), in rats anesthetized with hexenal (200 mg/kg). Three types of PAG neurons that differed in BA structure were identified. NRM, LC and SN stimulation changed BA only in type III neurons. Stimulation of these structures evoked an increase in BA frequency in 11.0–14.5%, and inhibition in 31.0–47.5% of type III neurons. Simultaneous stimulation of two structures led to a marked drop in intensity of effects. A depressing effect on BA was always detected if stimulation of one of the structures suppressed BA. Stimulation of two structures, with one being the NRM, was most effective. The role of PAG in the organization of the brain-stem component of the antinociceptive mechanism is discussed.A. A. Bogomolets Institute of Physiology, Ukrainian Academy of Sciences, Kiev. Translated from Neirofiziologiya, Vol. 24, No. 1, pp. 52–60, January–February, 1992.     

Opiocortin and catecholamine projections to raphe nuclei

Afferent projections to the nucleus raphe magnus (NRM) and dorsal raphe nucleus (DRN) were identified using retrograde transport of horseradish peroxidase conjugated wheat germ agglutinin (HRP-WGA). Neurons were labeled in important nociceptive regions including periaqueductal gray (PAG), arcuate nucleus, lateral hypothalamus and medial thalamic nuclei following both injections. We have immunocytochemically identified opiocortin/WGA neurons in the arcuate nucleus following NRM and DRN injections. Dual stained catecholamine/WGA perikarya were found in zona incerta, locus coeruleus, substantia nigra, nucleus tractus solitarius and adjacent A2, C2 and C3, lateral paragigantocellular reticular nucleus/C1 and lateral reticular nucleus/A1 following DRN injections and in zona incerta, substantia nigra, nucleus tractus solitarius/A2 and lateral reticular nucleus/A1 after NRM injections. These results provide further evidence for opiocortin and catecholamine modulation of analgesia.     

兔边缘系统背侧海马呼吸相关神经元

本工作对35只家兔在背侧海马区用玻璃微电极记录了103个神经元单位放电,分类如下:56个神经元属非呼吸节律性自发的间歇性放电,可分为两种类型:一为长时振荡型放电;另一为短时振荡型放电。47个神经元出现呼吸节律同步性自发放电,可分为三种类型:吸气型15个单位;呼气型9个单位;跨时相型23个单位。上述结果提示:边缘系统海马的呼吸信息在情绪活动的协调方面可能是一个有趣的问题。     

Effect of upper airway negative pressure and lung inflation on laryngeal motor unit activity in rabbit.

Distortion of the upper airway by negative transmural pressure (UANP) causes reflex vagal bradycardia. This requires activation of cardiac vagal preganglionic neurons, which exhibit postinspiratory (PI) discharge. We hypothesized that UANP would also stimulate cranial respiratory motoneurons with PI activity. We recorded 32 respiratory modulated motor units from the recurrent laryngeal nerve of seven decerebrate paralyzed rabbits and recorded their responses to UANP and to withholding lung inflation using a phrenic-triggered ventilator. The phasic inspiratory (n = 17) and PI (n = 5) neurons detected were stimulated by -10 cmH(2)O UANP and by withdrawal of lung inflation (P < 0.05, Friedman's ANOVA). Expiratory-inspiratory units (n = 10) were tonically active but transiently inhibited in postinspiration; this inhibition was more pronounced and prolonged during UANP stimuli and during no-inflation tests (P < 0.05). We conclude that, in addition to increasing inspiratory activity in the recurrent laryngeal nerve, UANP also stimulates units with PI activity.