延髓腹外侧Btzinger复合体
呼吸时相转换效应的研究^*

李勤　吕明　宋刚　张衡　刘磊

　　摘要　实验在40只氨基甲酸乙酯麻醉的成年家兔上进行,观察电刺激Btzinger复合体(Bt.C)对呼吸节律的影响。结果表明: (1)吸气相早期短串电刺激Bt.C导致膈神经放电被短暂抑制,被抑制的程度与刺激强度呈正相关。当刺激落位在吸气相中、晚期时,可导致吸气提前终止(“吸气切断”),之后跟随一缩短的呼气相。导致“吸气切断”的阈强度与刺激落位呈负相关。(2)呼气相短串电刺激Bt.C,可诱导膈神经短暂放电,呼气相晚期诱导出现放电之后往往跟随一正常的吸气放电,结果导致该呼气相缩短。该效应亦具有刺激强度、刺激落位依赖性。结果提示: Bt.C参与吸气向呼气以及呼气向吸气的时相转换。
　　关键词: Btzinger复合体;呼气递增神经元;时相转换;电刺激
　　学科分类号: R332.3,Q473

STUDIES ON RESPIRATORY PHASE SWITCHING
EFFECTS OF Btzinger COMPLEX^*

LI　QIN,,SONG　GANG^***,ZHANG　HENG,LIU　LEI
Department of Physiology,Shandong Medical University,Jinan 250012
L　MING
Neurosurgery Division,Liaocheng People′s Hospital,Liaocheng 252000

　　ABSTRACT　The effects of electrical stimulation of Btzinger complex (Bt.C) on respiratory rhythm were investigated in 40 urethane anesthetized adult rabbits.The results were as follows.(1) A short train stimulation delivered in the early inspiratory phase produced a transient inhibition of phrenic discharge.The stimulus,when delivered in the mid or late inpiratory phase,could cause a premature termination of the inspiratory phase (“inspiratory off-switch”) and a switching to the expiratory phase,which was accompanied with the reduced duration of the consecutive expiratory phase.There was a negative linear correlation between the threshold intensity of inspiratory off-switching and delivery time of stimulation.(2) A short train stimulation delivery in the expiratory phase elicited a transient phrenic discharge.The discharge in the late expiratory phase was followed by a premature onset inspiration.This effect was also dependent on the strength and delivery time of the stimulus.The results suggest that the Bt.C is involved in the central control of respiratory phase-switching.
　　Key words: Btzinger complex;expiratory augmenting neurons;phase-switching;electric stimulus

　　Lipski 和Merril 1980年首次用电生理学方法研究证明位于延髓腹外侧、面神经后核腹内侧区域的一群神经元为呼气神经元^［1］,即Btzinger复合体 (Bt.C)。随后的许多神经解剖学及电生理学研究证实,Bt.C的呼气递增神经元(E-aug)的轴突与膈运动神经元及脑干呼吸神经元有着广泛的抑制性突触联系^［2～4］。新近的研究表明长串电刺激猫的Bt.C导致吸气抑制,吸气相短串刺激可诱发“吸气切断”^［5～7］,然而Bt.C在呼吸时相转换中的作用尚未见系统的研究报道。本研究运用膈神经放电周期触发短串电刺激方法,观察电刺激Bt.C对呼吸节律的影响。部分内容已以摘要形式发表^［8］。
　　实验选用成年家兔40只,体重2～2.5 kg,雌雄不拘。动物制备、Bt.C定位及膈神经放电记录方法与前文吕明等^［9］报道的方法相同。需指出的是,本实验所用的刺激电极为电解研磨、绝缘的钨丝微电极（尖端直径1～3 μm,电阻5～10 MΩ）。电刺激由电刺激器（Nihon Kohden）经隔离器（Nihon Kohden）提供。短串电刺激参数: 强度10～100 μA,波宽0.3 ms,频率200 Hz,25～125 ms。短串电刺激的起始落位由动物自身膈神经放电控制,实验中刺激落位时间分别用吸气相刺激延迟系数（IDC）和呼气相刺激延迟系数(EDC)表示,刺激延迟系数=从吸（呼）气开始到刺激开始之间的时程/对照吸（呼）气时程。
　　实验结束后,在刺激部位通以阳极直流电（30 μA,15 s）制造损伤点标记。过量麻醉处死动物,取脑并置于10%甲醛溶液中固定,2周后做100 μm组织切片,观察刺激位点。统计学处理用实验前后配对t检验,以±s表示,P<0.05即认为有统计学意义,两变量间依存相关关系用直线回归与相关分析。

　　1.　吸气相短串刺激
　　吸气相早期（IDC≤0.4）短串电刺激Bt.C可导致膈神经放电短暂抑制(图1A: arrow 1B）。在一定范围内,抑制持续时间与刺激强度呈正相关（r=0.8773,P<0.001,n=99）。抑制潜伏期（从刺激开始至放电完全终至）为(4.7±0.3) ms。在吸气相中,晚期给予短串电刺激可导致吸气完全中断,即“吸气切断”。“吸气切断”之后往往跟随一缩短的呼气相(图1A: arrow 2C）。导致“吸气切断”的阈强度与刺激落位呈负相关（r=－0.8806,P<0.001,n=53）（图1D）,即阈强度随刺激落位的延后而下降。在吸气中期给予强度为50 μA的短串电刺激并导致“吸气切断”时,与前对照相比,随后的呼气时程缩短了(23.8±1.2)%(P<0.01,n=18)。

图1　吸气相短串电刺激Bt.C导致的膈神经放电短暂抑制及“吸气切断”效应
Fig.1　Transient inhibition of phrenic discharge and　“inspiratory off- switch” by short train stimulation to Bt.C in inspiratory phase
A.Short train stimulation (30 μA,5 pulses) to Bt.C at 0.2 s（IDC=0.33 arrow 1）and 0.4 s（IDC=0.66 arrow 2）after the onset of inspiration.Upper traces: integrated phrenic discharge;lower traces: original phrenic discharge.B.A higher sweep speed showing the transient inhibition of A (arrow 1).Dots showing 5 pulses.C.A higher sweep speed showing “inspiratory off-switch” of A (arrow 2).D.Relation between the delivery time of stimulus and threshold for “inspiratory off-switch”.

　　2.　呼气相短串刺激

　　呼气相短串电刺激Bt.C可诱导膈神经短暂放电。产生此效应的阈强度为(35±5) μA,该效应亦具有刺激强度和刺激落位依赖性,如图2所示,膈神经短暂放电积分幅度与刺激落位两者呈直线正相关,（r=0.9459,P<0.001,n=59）,即落位于呼气相晚期的刺激较落位于呼气相早期的刺激易于诱发膈神经短暂放电,并且当刺激落位于呼气相晚期时诱导出现的短暂放电之后往往跟随一正常的吸气放电,结果导致该呼气相的缩短。在呼气相晚期给予强度50 μA短串刺激,与前对照相比呼气时程缩短了(14.8±6.8)%(P<0.01,n=17)。从刺激结束至膈神经短暂放电开始出现的时间（潜伏期）为(36±3.7) ms。这一时间不随短串刺激的时程及强度的变化而变化。

图2　呼气相短串电刺激Bt.C诱导的膈神经短暂放电效应
Fig.2　Transient phrenic discharge in the expiratory phase by short train stimulation to Bt.C
A.Short train stimulation (50 μA,5 pulses) to Bt.C in early,mid and late expiratory phase elicited transient phrenic discharge (arrows).The discharge of late expiratory phase was followed by a premature onset inspiration.Upper traces: integrated phrenic discharge;lower traces: original phrenic discharge.B.A higher sweep speed showing the transient discharge of mid expiratory phase.C.A higher sweep speed showing the transient discharge of late expiratory phase.D.Relation between delivery time of stimulation and integrated transient phrenic discharge.

　　3.　对照实验

　　区域对照实验（n＝46）。在Bt.C的嘴端,内外侧各旁开1～1.5 mm三个区域,分别在吸气相或呼气相给予与效应区相同的短串电刺激,膈神经放电和呼吸时程均无明显变化。在Bt.C的尾端旁开1～1.5 mm处给予短串电刺激,可引起膈神经放电短暂抑制,这是由于Bt.C的投射纤维受到刺激所致^［3］。
　　本研究结果表明,吸气早期短串电刺激Bt.C可导致膈神经放电短暂抑制,吸气中、晚期短串电刺激Bt.C诱发“吸气切断”。呼吸神经元网络学说^［10］认为,E-aug的兴奋可进一步抑制在吸气相已被抑制的吸气递增神经元（I-aug）,彻底切断吸气形成呼气相。Bt.C恰好是E-aug集中存在的部位。细胞外记录及脉冲触发叠加（STA）等研究已表明Bt.C的E-aug可强烈抑制I-aug^［11］,显然我们的结果是符合上述假说的,与E-aug可强烈抑制I-aug的结果一致。实验中还发现,“吸气切断”之后紧跟一缩短的呼气相,这可能是由于“吸气切断”后E-aug和I-aug的交互抑制时程缩短,因此呼气相缩短。进一步分析表明,诱导“吸气切断”的阈强度在吸气相中逐渐降低,此结果提示,越接近吸气末期,Bt.C对外来刺激引起兴奋的阈值就越低,从而容易兴奋引起“吸气切断”,可见Bt.C控制由吸气向呼气的时相转换效应在吸气相中逐渐增强。研究还观察到,呼气相短串电刺激Bt.C可诱导出短暂的膈神经放电。我们认为这是因吸气神经元脱抑制后短暂兴奋所致。当刺激落位于呼气相晚期时,诱导出现的放电之后跟随一正常的吸气放电,结果导致该呼气相的缩短,由此可见Bt.C对吸气活动的抑制作用不仅促进吸气向呼气的时相转换,而且与吸气的启动,即呼气向吸气的时相转换有关。

*山东省自然科学基金（Q95C0824）和卫生部人才基金资助项目
^*Supported by Natural Science Foundation of Shandong Province (Q95C0824) and Talent Foundation of the Ministry of Public Health

^***Correspondence author

作者简介：宋刚：通讯作者
Correspondence author

作者单位：李勤　张衡　刘磊：山东医科大学基础医学院生理教研室,济南 250012
　　　　　吕明：聊城市人民医院神经外科,山东省聊城市　252000

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1997-12-15收稿　　1998-03-09修回