投射到兔颈上神经节的脊髓交感节前神经元——HRP法研究

本研究用成年家兔12只、猴1只。将20μl％HRP注射于兔的颈上神经节内,在猴则用15％的HRP。动物存活3～6天。切片按Mesulam氏联苯胺蓝色法作成色反应,中性红复染。脊髓内HRP标记细胞仅见于注射例,12只兔共有标记细胞7908个。但每例的标记细胞数目不等,最多1690个,最少71个。标记细胞的纵长分布为12个脊髓节(C_6～T_9),但主要集中在T_1～T_4(占86.18％),特别是T_2和T_3(占56.22％)。高峰则位于T_2(29.10％)。标记的节前神经元细胞柱的首尾长度在各动物也不一样,最长的11节,占C_7～T_9;最短的5节,占C_3～T_4。标记细胞的高峰在各例动物中分别位于T_1～T_3,但主要集中在T_2和T_3。在脊髓的横切面上,标记细胞集中于4个细胞群,即:中间外侧核本部(ILp)、中间外侧核侧索部(ILf)、中介核(IC)和中介核旁室管部(ICpe)。后者又分为背侧部(ICped)和腹侧部(ICpev)。HRP标记细胞主要位于ILp,12只兔有92.99％位于此。位于ILf的标记细胞较少,占6.25％。位于IC和ICpe者则更少,分别占0.68％和0.08％。此外,在前角背侧部可偶见标记细胞。本文虽仅用1只猴,但标记细胞的分布模式与兔者基本相同。     

氟伐他汀对心肌缺血兔颈上神经节神经元延迟整流钾通道特性的影响

目的 探讨氟伐他汀对心肌缺血情况下支配心脏的颈上交感神经节(SCG)神经元电生理特性的影响.方法 利用异丙肾上腺素皮下注射(85 mg·kg-1 ·d-1,共2d)制作兔急性心肌缺血模型,通过全细胞膜片钳技术研究对照组、心肌缺血组及氟伐他汀(10 mg· kg-1·d-1,共7d)干预组的SCG神经元延迟整流钾通道特性的改变.结果 对照组、缺血组和氟伐他汀干预组的兔SCG神经元的延迟整流钾通道最大峰值电流幅值分别为(80.80±22.80) pA/pF、(154.93±31.56) pA/pF、(119.82±24.82)pA/pF,3者的半数激活电压分别为(9.86±0.57)mV、(12.36±1.09) mV和(9.52±0.49) mV,斜率因子分别为(19.66±0.75)mV、(27.33±1.64) mV和(17.76±0.61) mV.急性心肌缺血使SCG神经元延迟整流钾通道电流(IK)幅度增大,氟伐他汀的干预可一定程度上使这种增大的电流有所减小.同时,氟伐他汀干预可使心肌缺血组移动的激活曲线向对照组方向移动.结论 心肌缺血情况下SCG神经元延迟整流钾通道特性产生的变化可一定程度上通过氟伐他汀的干预得到修复,这可能是氟伐他汀治疗心脏疾病的机制之一.     

家兔颈上神经节神经细胞分离培养的形态观察

本文通过神经细胞分离培养技术,观察了家兔颈上神经节神经细胞的形态学特点。指出,细胞分离培养的家兔颈上神经节神经细胞可呈单极、双极和多极性;双核神经细胞除核外,其形态特征与单核神经细胞相同,在新生家兔颈上神经节中存在着与脊神经节中神经母细胞相似的泪滴状神经细胞。双核神经细胞核与泪滴状神经细胞的形态学演变过程尚须进一步研究。     

家兔颈上神经节双核神经细胞的实验观察

本文通过组织培养和电子显微镜观察了家兔颈上神经节双核神经细胞在NGF阳激光作用下超微结构的改变。得出:1)在组织培养条件下,交感神经节单、双核神经细胞体的超微结构对NGF的反应相同。2)激光促进交感神经节细胞再生合成代谢形态学改变的生物学效应需要NGF的协助;其促进机理尚须进一步作生理生化研究。     

颈上神经节摘除术及节内注射术

颈上神经节是交感干头端的一个交感神经节,它接受来自脊髓侧角的节前纤维,然后发出节后纤维支配头颈部各器官,它的头端发出颈内动脉神经,作为交感干的延续伴随颈内动脉通过颈动脉管进入颅腔,从而支配颅内各脑区〔1〕。因而颈上神经节的位置及功能十分重要,可通过它来探讨头颈部及颅内各器官的交感神经供给情况。我们采用颈上神经节摘除术和节内注射荧光金束路追踪技术,成功地开展了交感神经纤维对松果体和颌下淋巴结的神经支配研究。现将该技术介绍于下:操作步骤:1.颈正中纵行切开动物皮肤2.钝性分离胸锁乳头肌,暴露颈动脉鞘3.将颈动脉鞘轻…     

急性分离脑部神经元的膜片钳技术研究概况

膜片钳是研究离子通道的重要手段,而急性分离技术是标本制备的主要方法。我们对以往的相关方法进行综述,试图寻找一种有效、简洁、可行的标本制备技术。     

大鼠颈上神经节脑内移植存活实验研究

本实验用幼年、年青和成年SD大鼠颈上神经节(SCG)分别移植于48只经6—OH-DA损毁黑质的同种成年鼠尾状核,存活1—2个月后,用尼氏染色和甲醛—戊二醛诱发单胺类荧光法对移植物进行了观察。结果18例移植幼鼠SCG者,移植物的节神经元几乎全部丧失;17例移植年青的SCG者,移植物中见有存活神经元的有9例,占52.94%;13例移植成年鼠SCG者,移植物中有存活神经元的有8例,占61.54%。存活神经元数量较少,且多分布在移植物的周围区,其胞质尼氏染色较深,荧光显微镜观察,显示指示CA存在的特异绿色荧光。在存活神经元存在的区域及该移植物与宿主脑组织之间的界面有大量毛细血管,并常有胶质细胞增生。本文对影响移植节神经元存活的因素进行了讨论。     

组胺在豚鼠颈上神经节中的分布

目的 检测组胺在外周交感神经元中的分布。方法 应用原位杂交组织化学技术检测组氨酸脱羧酶(HDC)mRNA的表达,用荧光双标免疫组织化学技术观察组胺和酪氨酸羟化酶(TH)在神经细胞中的共存。结果豚鼠颈上神经节中有较多的神经元呈．HDC阳性,HDC mRNA阳性信号在大、小细胞的细胞浆内均有分布,在细胞核内未检测到阳性信号;组胺的阳性细胞和TH的阳性细胞在豚鼠颈上神经节中均有大量分布,在大型和小型神经元内均发现有双标阳性信号;6．羟多巴胺(6-OHDA)化学损毁颈上神经节的交感神经细胞后阳性细胞大量减少。结论 提示在颈上神经节细胞的胞浆内有组胺的合成,组胺与单胺类神经递质在外周交感神经元中共存。     

Sodium and calcium currents of acutely isolated adult rat superior cervical ganglion neurons

Neurons enzymatically isolated from the adult rat superior cervical ganglion (SCG) were investigated using the whole-cell variant of the patch-clamp technique. Currentclamp studies revealed the following mean passive and active membrane properties: resting membrane potential, –54.9 mV; input resistance, 349 M; action potential (AP) threshold, –29.8 mV; AP overshoot, 53.3 mV; AP maximum rate of rise, 166.4 V/s; and AP duration, 3.2 ms. Chemosensitivity to acetylcholine remained intact following enzymatic dispersion. Voltage-clamp studies of a transient tetrodotoxin-sensitive Na⁺ current revealed activation and inactivation processes which could be fit to modified Boltzmann equations. Na⁺ current activation parameters for the half activation potential (Vh) and slope factor (K) were –23.3 mV and 5.3 mV, respectively. Inactivation parameters forVh andK were –59.3 mV and 7.6 mV, respectively. Voltage-clamp studies also revealed a high voltageactivated sustained inward current which was eliminated upon removal of external Ca²⁺, greatly reduced by 500 M Cd²⁺, and supported by Ba²⁺ or Sr²⁺. Tail current analysis of this Ca²⁺ current revealed a sigmoidal activation. A low voltage-activated transient Ca²⁺ current was not observed. We conclude that isolated SCG neurons retain the properties of neurons in intact ganglia and provide several advantages over conventional preparations for the study of voltagegated membrane currents.     

NOS-positive preganglionic neurons innervate a subpopulation of postganglionic neurons in superior cervical ganglion in rats

To determine the postganglionic targets of NOS-containing preganglionic neurons, we studied the association of NADPH-diaphorase positive preganglionic fibers and retrogradely labeled postganglionic neurons in the superior cervical ganglion (SCG) in rats. Wheat germ agglutinin-horseradish peroxidase solution was applied to the anterior chamber of the eye, middle cerebral artery, subcutaneous layer of the facial skin, or submucosal layer of the inside of the lip. Two days after tracer application, the rats were perfused with fixative solution. Serial sections of the SCG were stained histochemically for NADPH-diaphorase followed by diaminobenzidine reaction. More than 80% of the labeled postganglionic neurons innervating the structures in the subcutaneous or submucosal layer showed close association with NADPH-diaphorase positive preganglionic nerve terminals; approximately one-third of these labeled neurons were encircled by dense baskets of pericellular terminals. On the other hand, most of the postganglionic neurons innervating the iris (69%) or the cerebral artery (90%) did not show a distinct association with NADPH-diaphorase positive terminals. These results suggest that one of the principal roles of the NOS-containing preganglionic neurons may be in controlling the postganglionic neurons which innervate the structures in the subcutaneous or submucosal layer.     

Two types of acid-sensing ion channel currents in rat hippocampal neurons

Two types of acid-sensing ion channel (ASIC)-like currents in cultured rat hippocampal neurons were recorded and their characteristics were studied by using a whole-cell recording technique. The results revealed that the ASIC-like currents, induced by a quick drop of the extracellular pH, decayed with different time constants (τ) of 229 ± 16 (Type I) and 1209 ± 56 ms (Type II). The ASIC-like currents displayed different sensitivities to extracellular proton (pH_0.5 was 6.17 ± 0.04 for Type I and 5.70 ± 0.07 for Type II) and amiloride, a specific ASIC channel blocker (IC₅₀ was 1.19 ± 0.37 μM for Type I and 0.14 ± 0.02 μM for Type II). Among all the 360 recorded neurons, ASIC-like currents were induced in 314 neurons (87.2%). In the neurons expressing ASICs, Type I currents were evoked from 269 neurons (85.7%) and Type II currents were induced only from 45 neurons (14.3%). As these ASIC-like currents presented various electrophysiological and pharmacological properties, further experiments should be conducted to decipher the complex subunit composition of ASICs in the hippocampus.     

Sympathetic neurons in lower cervical ganglia send axons through the superior cervical ganglion

In addition to the preganglionic axons which innervate the superior cervical ganglia, the cervical sympathetic trunks of the rat have been shown to contain axons of ganglionic neurons. Following the application of horseradish peroxidase to the cut cervical sympathetic trunk just caudal to the superior cervical ganglion, a population of approximately 300 labeled neurons was found in the inferior and middle cervical ganglia. The labeled neurons were localized primarily in the more rostral regions of these ganglia. The axons of most of these neurons entered the superior cervical ganglion, passed through it, and left via the external carotid nerve.The relevance of these observations to physiological studies on the cervical sympathetic nervous system is discussed     

Expression of preproenkephalin mRNA in rat superior cervical ganglion during postnatal development

The number of principal neurons in the rat superior cervical ganglion (SCG) exhibiting enkephalin-peptide immunoreactivity is reported to be limited. To better determine the degree of enkephalinergic phenotype in sympathetic neurons, sections of SCGs from rats aged newborn to adult were processed for in situ hybridization histochemistry, using a [³⁵S]cRNA probe directed against preproenkephalin (PPENK). > 50% of principal ganglion neurons express mRNA for PPENK in adults. Striking variability in labeling intensity is observed. PPENK mRNA is detected in developing ganglia beginning at postnatal days 4–7. Both the number of cells and intensity of labeling increases with postnatal development. These results indicate that expression of PPENK mRNA is more widespread than expression of enkephalin peptides and develops postnatally.     

Immunohistochemical evidence from co-localization and denervation studies for four types of substance P-containing nervous structures in the rat superior cervical ganglion

Four types of substance P-immunoreactive structures have been distinguished in the rat superior cervical ganglion by double-immunofluorescence microscopy: (1) A major population of mainly varicose fibres enmeshed singly-scattered neuronal perikarya, some of which contained vasoactive intestinal polypeptide-immunoreactivity. These substance P-immunoreactive fibres did not contain colocalized calcitonin gene-related peptide (CGRP) and were absent after transection of the cervical sympathetic trunk. (2) A rather small substance P-immunoreactive fibre population with colocalized calcitonin gene-related peptide-immunoreactivity was distributed in a patchy manner and disappeared after cutting the postganglionic branches. (3) Most of the intraganglionic small intensely fluorescent (SIF) cell clusters were intensely substance P-immunoreactive. SIF cells were not visibly changed in number and fluorescence intensity by either surgical procedure. (4) Immunoreactivity was not visible in principal ganglionic neurons of control ganglia, but occurred in cell bodies after pre- as well as after postganglionic nerve transection. Some of the substance P-immunolabelled perikarya in addition revealed immunostaining to antisera against the catecholamine-synthesizin enyzme tyrosine hydroxylase or against the neuropeptides leu-enkephalin and vasoactive intestinal polypeptide, respectively. The results strongly suggest that, in addition to a substance P-containing preganglionic input (1), and a supply by substance P-containing sensory axon collaterals (2), the superior cervical ganglion of the rat gives origin to a paraganglionic (3) and a postganglionic (4) substance P-immunoreactive intrinsic system, the latter becoming visible only after disconnection of the sympathetic pathway.     

Amitriptyline modulates calcium currents and intracellular calcium concentration in mouse trigeminal ganglion neurons

Migraine is increasingly recognized as a channelopathy, and abnormalities of voltage-activated ionic channels could represent the molecular basis for the altered neuronal functioning. The high-voltage-activated (HVA) Ca²⁺ channels in the trigeminovascular system play a role in the pathophysiology of migraine. In the present study, effects of amitriptyline (AMT), a commonly used migraine prophylactic drug, on the HVA calcium currents (I_Ca) were examined in mouse trigeminal ganglion neurons using whole-cell patch clamp technique. AMT produced concentration- and use-dependent inhibition of HVA I_Ca. Bath application of GÖ-6983 (a selective protein kinase C inhibitor) or H89 (a protein kinase A inhibitor) did not reduce the AMT-induced inhibition of HVA I_Ca. A similar inhibition was observed when calcium imaging was used to directly monitor the effects of AMT on KCl-induced increments of intracellular Ca²⁺ concentration ([Ca²⁺]_i). By blocking HVA Ca²⁺ channels and Ca²⁺ entry into cells, AMT could prevent the release of neurotransmitters and help restore the neuronal threshold for excitation. Our findings suggest interesting therapeutic mechanisms for AMT in migraine prevention.     

Eugenol modifies the excitability of rat sciatic nerve and superior cervical ganglion neurons

Eugenol is a phenylpropene obtained from the essential oils of plants such as clove and basil which has ample use in dentistry. Eugenol possesses analgesic effects that may be related to the inhibition of voltage-dependent Na⁺ channels and/or to the activation of TRPV1 receptors or both. In the present study, electrophysiological parameters were taken from the compound action potentials of the isolated rat sciatic nerve and from neurons of the superior cervical ganglion (SCG) impaled with sharp microelectrodes under current-clamp conditions. In the isolated rat sciatic nerve, eugenol inhibited the compound action potential in a concentration-dependent manner. Action potentials recorded from SCG neurons were inhibited by eugenol with an IC₅₀ of 0.31 mM. At high concentrations (2 mM), during brief applications, eugenol caused significant action potential blockade while it did not interfere with the resting membrane potential or the membrane input resistance. Surprisingly, however, at low eugenol concentrations (0.6 mM), during long time applications, a reversible reduction (by about 50%) in the input membrane resistance was observed, suggesting the possible involvement of a secondary delayed effect of eugenol to reduce neuronal excitability.     

Temperature sensitivity of Ca currents in chick sensory neurones

We have investigated the effects of temperature on the Ca currents of chick sensory neurones. Raising the temperature from 17 to 37°C, caused low-threshold (LVA, T) and high-threshold (HVA, L and N) Ca currents to show a marked amplitude increase and a drastic acceleration of their activation-inactivation gatings. Compared to HVA channels, the LVA type showed a weaker temperature sensitivity. Its averageQ ₁₀ values were closer to those of other voltage-operated ion channels: 1.7 (permeability), 1.9 (activation) and 2.2 (inactivation). Alternatively, the activation kinetics and peak permeability of HVA Ca channels showed maximalQ ₁₀ values of about 5 and 2.8, respectively. HVA channel deactivation was less sensitive to temperature (Q ₁₀ 1.8). Inactivation of these channels was slow and monoexponential between 17 and 22°C, but faster and double exponential above 30°C, uncovering a fast temperature-sensitive decaying phase. The size and rate of decay of this component decreased with increasing membrane depolarizations and persisted at holding potentials positive to −80 mV, suggesting the involvement of temperature-sensitive Ca-mediated processes in the mechanism of HVA channel inactivation. Our data are consistent with the view that heating from 17 to 37°C causes both an increased probability of Ca channels to open and a drastic acceleration of their activation-inactivation kinetics.