Neuro-tracing approach to study kidney innervation: a technical note

Neuro-tracing approach is a great option to study innervation of the visceral organs including the kidneys. Important factors contributing to the success of this technique include the choice of a neuro-tracer, and delivery methods to result in successful labeling of peripheral sensory and motor ganglia. The neuro-tracer is usually applied directly to the kidney accessed via a surgical opening of the abdominal wall under deep anesthesia. A series of local microinjections of the dye are performed followed by a wound closure, and recovery period from the surgery. An extra care should be taken to prevent neuro-tracer spillage and accidental labeling of the surrounding organs during injections of the dye. Retrograde neuro-tracers like Fast Blue do not cross synapses, therefore, only neuronal bodies located within dorsal root ganglion neurons and major peripheral ganglia will be labeled by this approach. Retrogradely labeled peripheral neurons could be freshly isolated and dissociated for electrophysiological recordings and biochemical analyses (gene and protein expression), whereas the whole fixed ganglia could be sectioned to undergo immunohisto- and immunocytochemical targeted staining.

Protective Effect of Paeoniflorin against Optic Nerve Crush / 华中科技大学学报（医学）（英德文版）

In order to evaluate the efficacy of traditional paeonia extract paeoniflorin against optic nerve crush, 16 Brown Norway rats were divided into two groups at random, with 8 rats in each group. In paeoniaflorin-treated group, 2 mg paeoniaflorin (total volum: 1 mL) was injected into rat's peritoneum one time a day for a period of 8 days. Rats in untreated group were given a single dose of vehicle. The optic nerve was crushed by a special forceps for 30 s in the left eye and a sham proce-dure was performed in the right eye on the 2nd day after the first injection. The retrograde fluorogold labeling of ganglion cells was conducted 5 days after optic nerve crush. The whole retina was flat-mounted thereafter. The ganglion cells that survived the crush were counted under fluorescent microscope by using an automatic counting software. As compared with the contralateral eye, the survival rate of ganglion cells in the left eye increased from 40.22% to 64.53% with a significant dif- ference found between them (t=2.55, P=0.023). The results suggested that the paeonia extract paeoniflorin possessed a protective effect against optic nerve crush.

A Study of Laryngeal Innervation by HRP Retrograde Labeling and Immunohistochemical Methods / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: Classical concept of the laryngeal neuroanatomy has been questioned by some authors. Double innervation of some intrinsic laryngeal muscles has been suggested but controversy still exists. This study investigates the possibility of double innervation of thyroarytenoid and interarytenoid muscle and also the proportion of motor component in the external and internal branch of the superior laryngeal nerve. MATERIALS AND METHODS: Horse radish peroxidase (HRP) retrograde labeling in dogs, and choline acetyltransferase (ChAT) immunohistochemistry in feline and human laryngeal nerves were used. RESULTS: Evidences suggesting the existence of double innervation in thyroarytenoid and interarytenoid muscle and the existence of sensory component in external branch and motor component in internal branch of the superior laryngeal nerve have been observed. CONCLUSION: This new concept of the laryngeal neuroanatomy may be helpeful for understanding some of the neurologic diseases of the larynx such as spasmodic disphonia or variability of the vocal cord position in vocal cord paralysis. Further neuroanatomic and physiologic study is needed.

SOURCES OF THE SUPERIOR COLLICULUS AFFERENTS IN THE ALBINO RAT: AN HRP-LABELING STUDY / 解剖学报

HRP solution (33%) was injected into the left superior colliculus of 21 adult albino rats. 1-2 days after injection, the animal was sacrificed. Brains were frozen sectioned and processed according to. DAB- or TMB-method. The labeled neurones were examined in the right superior colliculus and other areas of the brain. The resu ts are as follows:1. No matter where the HRP was injected (either in the rostral or caudal part of the superior colliculus or thepart between them) HRP labeled neurones were always observed on the opposite superior colliculus if the injection sites reached layers deeper than the stratum oPticum. The locations of the labeled neurones corresponded roughly to the sites of HRP injections. However, no HRP labeled neurones were observed when the core of HRP deposites was restricted to layers superficial to the stratum griseum intermediale.2. Of all labeled neurones, 53.6% were located in the stratum griseum intermediale, 16.5% in the stratum opticum. The rest were in deeper layers. In no case were HRP labeled neurones observed in the stratum zonale or stratum griseum superficiale.3. Labeled neurones could be classified morphologically into vertical fusiform, horizontal fusiform and multipolar neurones.4. In addition to the visual cortex, labeled neurones were also found in the inferior colliculi, paralemniscal nuclei, dorsal nuclei of the lateral lemniscus, substantia nigrae and lateral tegmental nuclei bilaterally. Labeled neurones were also found in the ipsilateral ventral nucleus of the lateral geniculate body, contralatera pretectal area and reticular formation.

THE CGRP IMMUNOREACTIVE NEURONS INNERVATE BOTH THE CAPSULES OF CERVICAL ZYGAPOPHYSEALJOINTS AND FORE LIMB / 解剖学报

Objective To understand the base of the neuroanatomy underlying the nociceptive syndrome occurring in cervical zygapophyseal joints,in which pain in upper limb was accompanied. Methods The capsules of the C4-T1 zygapophyseal joints of SD rats were immunostained with anti-calcitonin-gene-related peptide(CGRP) antibody,and a modified tricetry labeling was done as following:the fluorescents,fast blue(FB) and nucleus yellow(NY),were injected,respectively,into the right radial nerve and capsules of the zygapophyseal joints of neck,then, the ipsilateral dorsal root ganglia(DRG) were sectioned and the retrogradely labeled sensory neurons in the DRG were observed and photographed,finally,the sections were further immunostained with anti-CGRP antibody,eventually,the neurons in both of the photos labeled by fluorescents and anti-CGRP antibody were comparatively observed and analyzed. Results The nervers fibers containing CGRP distributed throughout the capsules of the zygapophyseal joints from C4-T1 in the rat.Most of these fibers coursed straightly and individually,and some of them anastomosed to each other to form the networks of the nerve fibers.The neurons retrograde labeled by fluorescents were observed in each of the DRG from C5-T1.These neurons shared a labeling populations of FB single(76%),NY single(16%) and FB/YN double labeling(8%) respectively.Moreover, about 40% fluorescents labeled neurons were immunoreacted to CGRP.Conclusion The peripheral processes from a population of sensory neurons that contain CGRP in the cervical DRG simultaneously innervated both the capsules of cervical zygapophyseal joints and the fore limb with the radial nerve.The present results suggest that the nociceptive syndrome of cervical zygapophyseal joints with pain in human upper limb might be the referred pain occurring in the level of the DRG.

SPINAL PROJECTIONS TO THE PARABRACHIAL NUCLEUS IN THE RAT——HRP AND FLUOROGOLD RETROGRADE TRACT TRACING STUDY / 解剖学报

By introducing a mixture of WGA-HRP and HRP or fluorogold into the parabrachial nucleus, the cell origin of the spinoparabrachial projections in the rat have been carefully examined. The labelled neurons were found in bilateral spinal gray matter, lateral spinal nucleus and lateral cervical nucleus with contralateral predominance. They were mainly located in lamina I, lamina II, lamina IV, lamina V and lamina VII of the gray matter and also in the gray matter commissure posterior to the central canal. Comparing the distribution patterns of the projection neurons in the cervical, thoracic, lumbar and sacral segments, we did not find any distinct differences. The fact that the parabrachial nucleus receives a wide extensive projections form the spinal segments suggests that the spino-parabrachial pathways are possibly involved in the transmission of multiple sensory inputs.

AFFERENT PATHWAY OF THE VAGAL NERVE FIBERS CONVEYING CARDIAC PAIN INFORMATION VIA THE SUPERIOR CERVICAL GANGLION / 解剖学报

Objective To examine the cardiac primary afferents passing through the superior cervical ganglion which the sensory neurons are located in the vagal ganglion. Methods Retrograde tracing transport combined with immunohistochemistry. Results After injecting the horseradish peroxidase(HRP) into the superior cervical ganglion in the rat, the small number of retrogradely labeled neurons consistently appeared in the upper local portion of the nodose ganglion. The same injecting of fluorogold(FG) followed by immunohistochemical staining with SP, it was found that the double\|labeled neurons with FG/SP were approximately 20% occupied the total population of the SP positive neurons in the nodose ganglion. Conclusions The present results, associated with previous reports, suggest that the pathway of the vagal afferent conveying cardiac pain information which contains SP pass through the superior cervical ganglion.\;[

THE DISTRIBUTION OF NOS-POSITIVE NEURONS IN THE PROJECTING PATHWAY TO THE SUBNUCLEUS RETICULARIS DORSALIS IN THE RAT / 解剖学报

Objective To observe the distribtuion of NOS positive neurons in the projecting pathway to the subnucleus reticularis dorsalis(SRD) in the rat. Methods We used a double staining technique combined retrograde tract tracer and NADPH d histochemistry. Results FG/NOS double labeled neurons were found in the laminae Ⅰ,Ⅹ of spinal cord,dorsal raphe nucleus(DR),linear raphe nucleus(LR),and raphe obscurus nucleus(RO).In the periaqueductal gray(PAG),there were many FG labeled or NOS labeled neurons,but no FG/NOS double labeled neurons were found.Conclusion\ NO might play a role in the pathway of afferent projections of the SRD.\;[

MORPHOLOGICAL STUDIES ON THE TECTAL CELLS PROJECTING TO NUCLEUS ROTUNDUS IN THE CHICK / 解剖学报

Objective To study the morphological characteristics of tectal cells in stratum griseum centrale(SGC) which project to nucleus rotundus(Rt) in chick. Methods Tectal cells projecting to the Rt were retrogradely labeled by using the injection or implantation of a small amount of carbocyanine fluorescent tracer(DiI) into Rt postmortemly in chicks.Results Labeled SGC cells were classified into four types according to the location of the soma and dendritic endings in the tectal layers.Type 1 cells of the SGC,whose somata were located in superficial part of the SGC,gave off dendritic endings to layer F.Type 2 cells in the SGC,whose somata were also located in superficial part of the SGC,gave off dendritic endings to layer D.Type 3 cells,whose somata were located in deep part of the SGC,gave off primary dendrites obliquely in layer H-J of SGFS.Type 4 SGC cells,whose somata were located in the deep part of the SGC,gave off dendrites horizontally and their dendrites were located within the SGC.The labeled dendrites of type I and 2 cells of the SGC formed bush-like or bifurcated endings extending horizontally in layer F and bottle brush endings vertically in layer D,respectively.The dendrites of type 3 and type 4 cells mainly formed free endings in their extending deep tectal layers.Conclusion The dendrites of superficial SGC cells(type 1 and type 2 cells) extend to retinorecipient tectal layers(layer F and layer D,respectively),having the shape of bush-like or bifurcated endings extending horizontally in layer F and bottle brush endings vertically in layer D,respectively corresponding to the shapes of terminals of optic nerves in these layers.The dendrites of deep SGC cells(type 3 and type 4 cells) do not extend to the retinorecipient tectal layers,mainly forming free endings in the tectal layers deeper than layer H.

POSTERIOR PITUITARY PROJECTING NEURONS IN THE BED NUCLEI OF THE STRIA TERMINALIS IN THE RAT / 解剖学报

After injecting the posterior pituitery of the albino rats with cholera-toxinconjugated HRP,retrogradely labeled neurons were found in the area of the bednuclei of the stria terminalis (BST),which were then analyzed,based on therecent cytoarchitectonical study of the BST.In the anterior subdivision of the BSTdendrites could be seen in the parastrial nucleus with a few labeled cells surround-ing it;cells of the magnocellular nucleus were found scattered in the subdivision;some labeled cells were also identified in the ventral nucleus.Surrounding the dor-sal half of the posterior division of the BST there were several fairly distinct labe-led cell groups,viz.,the anterior fornical nucleus,medial and lateral dorsal accessorygroups,and the subinterventricular group.Though these cell groups are contiguouswith the BST,they do not seem to belong to the BST itself,but their dendritesoften extend into the posterior division of the BST,thus sharing some commonafferents with the latter.A few less constant cells were sometimes seen scattered invarious parts of the BST.