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大鼠腓浅神经传入纤维在脊髓胶状质的定位投射——FRAP法*
http://www.100md.com 《解剖学报》 1998年第4期
     作者:丁继固 任立恩 陈小武 蔡丽霞 胡松林

    单位:咸宁医学院解剖学教研室,咸宁 437100

    关键词:腓浅神经;跨神经节溃变;胶状质;抗氟化物酸性磷酸酶法;大鼠

    解剖学报/980410 摘 要 为了解大鼠腓浅神经传入纤维在脊髓胶状质(SG)的投射定位,按照跨神经节溃变原理,用抗氟化物酸性磷酸酶(FRAP)法和显微测量进行定量研究。结果显示:大鼠腓浅神经向SG的纵向投射主要为L2~6,L2~6各节段SG水平切面“眉毛状反应带”弧形长均值分别为0.795、0.849、0.913、0.921、0.852mm;而腓浅神经向L2~6各节段SG水平切面投射均值分别为0.447~0.533、0.384~0.561、0.351~0.598、0.338~0.611、0.383~0.561mm的范围。结果表明,腓浅神经反应带占据SG中间区域。
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    有关周围神经传入纤维向SG投射的研究,国内外不少学者曾有报道,如Knyihar和Csillik[1]曾切断大鼠左侧坐骨神经,4d内导致L4~S1节段SG内FRAP反应完全消失(水平方向的定位未作描述)。邱树华[2]用血管钳挫伤左侧坐骨神经,结果于L3~S1节段左侧SG中间内侧2/3部位的FRAP活性消失。朱培纯[3]、胡松林[4]等均做过这方面的研究,但对其局部定位均未作测量,本研究试图在上述研究基础上对腓浅神经传入纤维在SG的定位进行显微测量研究,为针刺镇痛及内脏牵涉痛原理提供较为准确的形态学依据。

    材料和方法

    用健康Wistar大鼠20只,雌雄不拘,体重180~250g,水合氯醛腹腔注射麻醉(300mg/kg),然后在腓骨颈处切断腓浅神经(切断左侧5只,切断右侧5只,双侧切断10只)。非手术侧作为本实验的对照侧。依据跨神经节溃变时间[5],术后存活7~12d,活体断头迅速暴露腰骶部脊髓,按神经根确定各节段并做标记。取出L1~S1脊髓,放在-15℃冰台上数分钟,按标记再分段,将各段分别置于-15℃恒冷切片机上做20~25μm的连续横切,每个节段又按上、中、下3部分贴片,将切片迅速吹干,固定于甲醛钙液内10min,按FRAP法[6],以β-苷油磷酸为底物(pH 4.7),硝酸铅为捕获剂加入0.02mmol/L氟化钠,以抑制胶状质以外其他部位溶酶体内的酸性磷酸酶(AcP),在37℃恒温下孵育4~6h,经硫化铵处理,以显示胶状质内溶酶体外的FRAP。
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    经上述处理后,再将显色的切片置于(10×10)光镜下,用物镜测微尺(0.01mm)标定好目镜测微尺(每1刻度为0.0134mm),设胶状质的内侧端为“0”值,对各显色切片中双侧SG内FRAP活性的水平向“眉毛状反应带”长及其消失区进行观察测量,将所测值进行统计学处理,以确立腓浅神经传入纤维在SG的投射模式。

    结果

    非手术对照侧与手术侧L1下段、S1上段FRAP活性均呈黑色“眉毛状反应带”,未见消失区(图1);切断腓浅神经实验侧,L2~6节段SG中间带呈现宽窄不等的FRAP活性消失区(图2~6)。L2~6各节段SG“眉毛状反应带”长及消失区测值亦不同,反应带L2为0.795mm,消失区在0.447~0.533mm范围(但上部5例、中部9例、下部17例);L3为0.849mm,消失区在0.304~0.561mm范围;L4为0.913mm,消失区在0.351~0.598mm范围;L5为0.921mm,消失区在0.338~0.611mm范围;L6为0.852mm,消失区在0.383~0.561mm范围(但中部仅15例、下部9例),各节段上、中、下3部分测值见附表。
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    附表 大鼠腓浅神经切断侧脊髓胶状质FRAP活性消失区测微匀值(mm) N:20

    Table The micromeasurement's mean value of the FRAP activity disappearea area in SG of the

    spinal cord on the rat's superficial peroneal nerve cut side

    L2

    ΣX

    L3

    ΣX

    L4
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    ΣX

    上

    upper

    中

    medial

    下

    lower

    上

    upper

    中

    medial

    下

    lower
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    上

    upper

    中

    medial

    下

    lower

    眉样反应带长

    the length of the eye-

    brow-like reactive zone

    N

    20

    20
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    20

    20

    20

    20

    20

    20

    20

    VMax

    0.871

    0.871

    0.871

    0.938

    0.938
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    0.938

    1.005

    1.005

    1.005

    VMin

    0.670

    0.670

    0.670

    0.670

    0.737

    0.737

    0.804

    0.804
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    0.804

    X

    0.791

    0.794

    0.801

    0.795

    0.834

    0.851

    0.864

    0.849

    0.901

    0.911

    0.925
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    0.913

    S

    0.060

    0.054

    0.051

    0.055

    0.063

    0.054

    0.048

    0.055

    0.051

    0.051

    0.051
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    0.051

    消失区起点值

    the original value

    of the extinct area

    N

    5

    9

    17

    20

    20

    20

    20

    20
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    20

    VMax

    0.469

    0.469

    0.469

    0.469

    0.469

    0.469

    0.402

    0.402

    0.402

    VMin

    0.402
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    0.402

    0.402

    0.335

    0.335

    0.335

    0.268

    0.268

    0.268

    X

    0.456

    0.461

    0.427

    0.447
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    0.397

    0.383

    0.373

    0.384

    0.369

    0.345

    0.338

    0.351

    S

    0.029

    0.022

    0.033

    0.028
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    0.041

    0.043

    0.039

    0.041

    0.046

    0.050

    0.051

    0.049

    消失区止点值

    the terminal value

    of the extinct area

    N
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    5

    9

    17

    20

    20

    20

    20

    20

    20

    VMax

    0.536

    0.603

    0.603

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    0.603

    0.603

    0.670

    0.670

    0.670

    VMin

    0.469

    0.469

    0.469

    0.469

    0.469

    0.536
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    0.536

    0.536

    0.536

    X

    0.523

    0.536

    0.540

    0.533

    0.547

    0.566

    0.569

    0.561

    0.586
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    0.596

    0.613

    0.598

    S

    0.030

    0.033

    0.037

    0.034

    0.041

    0.040

    0.034

    0.039

    0.029
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    0.037

    0.039

    0.035

    L5

    ΣX

    L6

    ΣX

    上

    upper

    中

    medial

    下

    lower
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    上

    upper

    中

    medial

    下

    lower

    眉样反应带长

    the length of the eye-

    brow-like reactive zone

    N

    20

    20
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    20

    20

    20

    20

    VMax

    1.005

    1.005

    1.005

    0.938

    0.871

    0.871

    VMin

    0.804
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    0.804

    0.804

    0.737

    0.737

    0.737

    X

    0.928

    0.918

    0.918

    0.921

    0.874

    0.844

    0.838
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    0.852

    S

    0.045

    0.044

    0.044

    0.044

    0.046

    0.046

    0.046

    0.046

    消失区起点值

    the original value

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    N

    20

    20

    20

    20

    15

    9

    VMax

    0.402

    0.402

    0.402

    0.469
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    0.469

    0.469

    VMin

    0.268

    0.268

    0.268

    0.335

    0.335

    0.335

    X

    0.332

    0.335

    0.348
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    0.338

    0.378

    0.384

    0.387

    0.383

    S

    0.046

    0.038

    0.041

    0.042

    0.039

    0.039

    0.045
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    0.041

    消失区止点值

    the terminal value

    of the extinct area

    N

    20

    20

    20

    20

    15

    9

    VMax

    0.670
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    0.670

    0.670

    0.670

    0.670

    0.603

    VMin

    0.536

    0.536

    0.536

    0.469

    0.469

    0.469

    X
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    0.623

    0.609

    0.599

    0.611

    0.576

    0.563

    0.543

    0.561

    S

    0.044

    0.043

    0.034

    0.040
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    0.051

    0.042

    0.040

    0.044

    讨论

    1. 大鼠腓浅神经向SG纵向投影分析

    我们此前的研究[7]发现,腓总神经传入纤维主要投射至L2~6,部分实验动物其纤维投向S1的上、中部(上部7例、中部7例)。本次实验切断腓浅神经,发现其纵向投射也主要在L2~6,而S1无1例发现消失区,并且L2上部仅5例、中部9例、下部17例;L6中部15例、下部9例出现消失区。由此看来,腓浅神经向SG纵向投射首尾向,尤其是尾侧比腓总神经投射约少半个节段,这可能是腓总神经其他分支的投射区域。
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    2. 大鼠腓浅神经向SG水平投射分析

    从统计表各测值分析,腓浅神经主要投向SG的中间带,从上至下逐渐增宽,至L6才稍变窄。如果把腓总神经投射区值[8]与腓浅神经投射区值比较,发现这两条神经从L2~6的起点值之差分别是:0.024(0.423与0.447之差(mm)、0.011(0.395与0.384之差)mm、0.001(0.352与0.351之差)mm、0.019(0.319与0.338之差)mm、0.039(0.344与0.383之差)mm;其止点值之差分别为:0.004(0.537与0.533之差)mm、0.054(0.615与0.561之差)mm、0.089(0.687与0.598之差)mm、0.088(0.699与0.611之差)mm、0.108(0.669与0.561之差)mm。从上述数据看,两神经起点值之差值除L2、L6稍大外,其他各节段之差值不大;而止点值的差值除L2相差不大外,其余各节段差值从上至下逐渐增大,由此看来腓浅神经投射在腓总神经投射区域的内侧(约占2/3),至于腓总神经比腓浅神经多额的投射区域(尤其是外侧),可能是腓总神经其他分支(如腓肠外侧皮神经和腓深神经等)的投射区域所致,而其他分支投射有待今后进一步研究。
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    图7 腓浅神经投射区模式图

    Fig.7 Projection area mould figures of the superficial peroneal nerve

    上述研究结果较为精确地确立了大鼠腓浅神经在SG的定位,为针刺治疗、针麻原理及内脏牵涉痛原理提供了较为准确的形态学依据。

    本研究根据杨安峰[8]引用Waibl报道的脊髓各节段高度,其中L2~6分别为3.9、3.7、3.3、2.9、2.5(mm)。“眉毛状反应带”长按1∶5,腓浅神经投射至各相应节段的主要测量均值按1∶40绘制成腓浅神经投射区模式图(附图)。

    图1 大鼠脊髓L1下部.切断右侧腓浅神经9d后,切断神经侧和非切断侧SG呈现FRAP活性 ×15
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    图2 大鼠脊髓L2中部.切断两侧腓浅神经10d后,两侧SG中间带1/3范围呈现FRAP活性消失 ×15

    图3 大鼠脊髓L3中部.切断两侧腓浅神经10d后,两侧SG中间带1/2范围呈现FRAP活性消失 ×15

    图4 大鼠脊髓L4中部.切断左侧腓浅神经8d后,左侧SG中间带2/3范围呈现FRAP活性消失 ×15

    图5 大鼠脊髓L5中部.切断左侧腓浅神经8d后,左侧SG中间带呈现FRAP活性 ×15

    图6 大鼠脊髓L6上部.切断右侧腓浅神经11d后,右侧SG中间带2/3范围呈现FRAP活性消失 ×15

    Fig.1 The low part L1 spinal segment of rat.On the 9th day after superficial peroneal nerve of right side were cut,the activity of FRAP in all SG was found on the cut nerve side and uncut side.×15
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    Fig.2 The middle part of L2 spinal segment of rat.On the 10th day after superficial peroneal nerve of both side were cut,the activity of FRAP in 1/3 of the intermediate zone of both side SG disappeared on the cut both side nerve.×15

    Fig.3 The middle part of L3 spinal segment of rat.On the 10th day after superficial peroneal nerve of both side were cut,the activity of FRAP in the 1/2 of the intermediate zone of both side SG disapeared on the cut both side nerve.×15
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    Fig.4 The middle part of L4 spinal segment of rat.On the 8th day after superficial peroneal nerve of left side were cut,the activity of FRAP in the 2/3 of the intermediate zone of left side SG disappeared on the cut nerval side.×15

    Fig.5 The middle part of L5 spinal segment of rat.On the 8th day after superficial peroneal nerve of left side were cut,the activity of FRAP in the intermediate zone of left side SG disappeared on the cut nerval side.×15
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    Fig.6 The upper part of L6 spinal segment of rat.On the 11th day after superficial peroneal nerve of right side were cut,the activity of FRAP in the 2/3 the intermediate zone of right side SG disappeared on the cut nerval side.×15

    收稿1997-04 修回1998-04

    参考文献

    [1]Knyihar E.Csillik B.Effect of peripheral anatomy on the fine structure and histochemistry of the Rolando substance:degenerative atrophy of central processes of pseudounipolar cells.Exp Brain Res,1976,26(6):73
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    [2]邱树华,戴世吉,吴海霞.大鼠坐骨神经对脊髓胶状质投射的节段定位——酸性磷酸酶(ACP)法研究.解剖学通报,1982,5(增刊2,上):41

    [3]朱培纯,吴海霞,许 红.大鼠面部各区皮肤对胶状质的躯体定位投射——酸性磷酸酶法研究(Ⅰ).解剖学报,1981,12(2):155

    [4]胡松林,胡圣望,蔡丽霞.大鼠正中神经感觉在脊髓胶状质定位投射——酸性磷酸酶(ACP)法研究.解剖学报,1988,19(2):159

    [5]Grant G,Ygge J,Molander C.Peripheral sensory nerve projection studied by transganglionic degeneration and transganglionic transport of horseradish peroidase.In:Brown AG,Rethety M(eds).Spinal cord sensation.Edinburgh:Scottish Academic Press,1981:33-36
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    [6]皮尔斯(马仲魁等译).组织化学.北京:人民卫生出版社,1965:657~568

    [7]丁继固、胡松林、蔡丽霞.大鼠坐骨神经、胫神经、腓总神经在脊髓胶状质定位投射的定量分析——FRAP法.中国组织化学与细胞化学杂志:1993;2(1):75

    [8]杨安峰,王 平.大鼠的解剖和组织.北京:科学出版社,1985:160~161

    THE ORIENTATIONAL PROJECTIONS OF THE AFFERENT

    FIBERS OF THE SUPERFICIAL PERONEAL NERVE TO

    THE SUBSTANTIA GELATINOSA OF THE SPINAL
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    CORD IN RATS——FRAP METHOD

    Ding JiguΔ, Ren lien,Chen Xiaowu, Cai Lixia, Hu Songlin

    (Department of Anatomy,Xianning Medical College,Xianning)

    To research the orientational projections of the afferent fibers of the rat's superficial peroneal nerve to the substantia gelatinosa(SG) of spinal cord.Methods: In accordance with the principle of transganglion degeneration, a quantitative research was made,adopting fluoride-resistant acid phosphatas (FRAP) and micromeasurement technique were used.Result: The longitudinal projection of the superficial peroneal nerve to SG was in L2-6.The mean value of the arched length of horizontal eyebrow like reactive zone of the SG in L2-6 was measured as 0.795、0.849、0.913、0.921 and 0.852mm respectively.And the mean value measured in the horizontal projection of the superficial peroneal nerve to L2-6 was in 0.447-0.533、0.384-0.561、0.351-0.598、0.338-0.611 and 0.383-0.561mm area respectively. Conclusion: The results show that the reactive zone of the superficial peroneal nerve is occurred intermediate area of SG.

    KEY WORDS Superficial peroneal nerve; Transganglion degeneration; Substantia gelationosa;Fluorid-resistant acid phosphatase; Rat

    △Department of Anatomy,Xianning Medical College,Xianning 437100,China

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