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A central objective in deciphering the nervous system in health and disease is to define the connections of neurons. The propensity of neurotropic viruses to spread among synaptically-linked neurons makes them ideal for mapping neural circuits. So far, several classes of viral neuronal tracers have become available and provide a powerful toolbox for delineating neural networks. In this paper, we review the recent developments of neurotropic viral tracers and highlight their unique properties in revealing patterns of neuronal connections.
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The laterodorsal tegmentum (LDT) is a brain structure involved in distinct behaviors including arousal, reward, and innate fear. How environmental stimuli and top-down control from high-order sensory and limbic cortical areas converge and coordinate in this region to modulate diverse behavioral outputs remains unclear. Using a modified rabies virus, we applied monosynaptic retrograde tracing to the whole brain to examine the LDT cell type specific upstream nuclei. The LDT received very strong midbrain and hindbrain afferents and moderate cortical and hypothalamic innervation but weak connections to the thalamus. The main projection neurons from cortical areas were restricted to the limbic lobe, including the ventral orbital cortex (VO), prelimbic, and cingulate cortices. Although different cell populations received qualitatively similar inputs, primarily via afferents from the periaqueductal gray area, superior colliculus, and the LDT itself, parvalbumin-positive (PV) GABAergic cells received preferential projections from local LDT neurons. With regard to the different subtypes of GABAergic cells, a considerable number of nuclei, including those of the ventral tegmental area, central amygdaloid nucleus, and VO, made significantly greater inputs to somatostatin-positive cells than to PV cells. Diverse inputs to the LDT on a system-wide level were revealed.
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The basolateral amygdala (BLA) receives dense projections from cholinergic neurons of the basal forebrain. Acetylcholine can contributes to amygdala-dependent behaviors: formation and extinction of fear memory and appetitive instrumental learning. However, the cholinergic mechanism at the circuit level has not been defined yet. We demonstrated that cholinergic-induced di-synaptic inhibition of BLA pyramidal neurons exhibits a retrograde form of short-term synaptic inhibition, depolarization-induced suppression of inhibition (DSI). Activation of nicotinic receptors was sufficient to evoke action potentials in cholecystokinin (CCK)-positive inhibitory neurons, which strongly inhibit pyramidal neurons through their perisomatic synapses. Our cell type-specific monosynaptic retrograde tracing also revealed that CCK neurons are innervated by basal forebrain cholinergic neurons. Therefore, our data indicated that CCK inhibitory neurons mediate the cholinergic-induced di-synaptic inhibition of BLA pyramidal neurons.
Subject(s)
Acetylcholine , Action Potentials , Basal Forebrain , Basolateral Nuclear Complex , Cholecystokinin , Cholinergic Neurons , Conditioning, Operant , Iontophoresis , Memory , Neurons , Pyramidal Cells , Receptors, Nicotinic , SynapsesABSTRACT
Spinal cord injury is one of the important field of neuroscience, and neural tract tracing techniques are important research tools. The neural fiber types in the spinal cord are complex, and there are some difference in structures between human beings and animals. This article reviewed different types of neural fibers closely related with spinal cord injury and regeneration, the difference between human beings and animals, and the application of neural tract tracing techniques in the study of spinal cord injury and regeneration.
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[Objective]To evaluate the axonal transport function of the regenerate nerve of buccal branch,marginal mandibular branch and cervical branch after chemically extracted acellular facial nerve allograft to the whole facial nerve defect in rat using fluoro-gold(FG),quantum dot(QD) and fast blue(FB).[Method]The extracranial section of facial nerve and branches were dissected under light microscope,and 10-mm-long segments of facial nerve gap were made on the left side.The defect was repaired by acellular facial nerve allografts with epineural suture method.Two months later,fluoro-gold(FG),quantum dot(QD) and fast blue(FB) were injected into distal bridged side of buccal branch,marginal mandibular branch and cervical branch.After three days,the slices of brain stem were collected and the facial motoneuron was observed under fluorescences microscope.[Result]There were three kinds of different fluorescences in the frozen sections of the facial nerve originated from the brainstem nuclei under the fluorescence microscope.Fluoro-gold(FG),quantum dot(QD) and fast blue(FB) labeled neurons were shown yellow,red and blue colors,respectively.[Conclusion]According to estimation of axonal branching by triple retrograde labeling,it was found that the regenerate facial nerve repaired the gap,and nerve continuity obtained restoration.The evaluation method of this study is simple,reliable and easy.It is an ideal evaluation method.
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Immunofluorescence histochemistry combined with retrograde tracing technique was employed to observe the effects of masseteric nerve transection on the expression of Trk ( tropomyosin-related kinase) receptor proteins, namely TrkA, TrkB and TrkC in the trigeminal mesencephalic nucleus ( Me5 ) of the rat. At 7 and 14 days following transection of masseteric nerve through which Fluorogold (FG) was applied to identify the Me5 neurons innervating masseter, brain sections were immunohistochemically processed to detect the three Trk isoforms in FG-labeled Me5 neurons. With the percentage of double-labeled neurons to the total number of FG-labeled neurons as the index,we demonstrated ( 1 ) a significant increase in the percentage of TrkA-immunoreactive (IR) Me5 neurons at both 7 and 14 days after nerve transection, (2) no significant, but gradual, increase in the percentage of TrkB-IR Me5 neruons with longer survival time post transection and ( 3 ) little change of TrkC expression. The current findings indicate that axotomy differently affected the expression of the individual Trk receptors and these expression patterns may reflect an adaptation of the Me5 neurons to the peripheral nerve injury.
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Objective In order to make further investigation of the functional meaning of Ca~(2+)dependent phospholipids binding protein AnnexinⅡ,we tested the effects of AnnexinⅡ combined with embryonic neural stem cells(NSCs) transplantion on the repair of spinal cord injury(SCI). Methods The spinal cord of the adult rats was transected completely between T_9-T_(10).AnnexinⅡ and NSCs were injected at the transected site.The lesion area of the spinal cord,growth of axon,and survival number and migration of the transplanted NSCs were measured.The survival number was shown by prelabeling the NSCs with Hoechest 33342 and the growth of axon traversing the transected area was shown by fluorogold(FG) retrograde tracing.The AnnexinⅡ injection+NSCs implantation groups were compared with groups that respectively received 1.NSCs implantation alone;2.sham-operation+NSCs implantation and 3.Vehicle injection of culture medium. Results Our results demonstrated that AnnexinⅡ treatment in vivo could significantly reduce the lesion area(P
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This study was performed to investigate origins of the dorsal root ganglion cells containing calcitonin gene -related peptide (CGRP) which innervate the calcaneal tendon in the rat. We used the horseradish peroxidase (HRP) or fluoro -gold (FG) to trace retrogradely somatic afferents in dorsal root ganglion cells after unilateral injections into the rat calcaneal tendon. HRP or fluoro -gold labeled DRG cells for the calcaneal tendon were seen generaaly in lumbosacral (L1 to S1) DRGs ipsilaterally. In lumbosacral DRGs, the largest number of labeled cells were found in the L6 DRG. Many DRG cell bodies contained the CGRP throughout the L1~S1. A plenty of HRP -or FG -labeled cells innervating the calcaneal tendon were also identified to contain the CGRP in L1~S1 DRGs. These FG +/- CGRP DRG cells innervating the calcaneal tendon were primarily found in the L6 DRG. These results suggest that the main sensory DRG for the calcaneal tendon is the L6. This fact may be available in diagnosis and treatment of neurogenic pain in the calcaneal tendon.
Subject(s)
Animals , Rats , Calcitonin , Diagnosis , Diagnosis-Related Groups , Ganglia, Spinal , Horseradish Peroxidase , Immunohistochemistry , Spinal Nerve Roots , TendonsABSTRACT
PURPOSE: This study was undertaken to determine the origins of dorsal root ganglion (DRG) cells containing calcitonin gene-related peptide (CGRP) which innervate the quadriceps femoris tendon in the rat. MATERIALS AND METHODS: DRG cells containing CGRP, which innervate the quadriceps femoris tendon, from 25 rats (Sprague-Dawley, 200-250 g) were examined using the retrograde tracing technique (neural tracers: horseradish peroxidase and fluorogold) combined with immunohistochemistry. RESULTS: Injection of horseradish peroxidase (HRP) or fluoro-gold (FG) into the quadriceps femoris tendon resulted in the ipsilaterally labelling of cells between L1 and L6 DRGs. However, a large number of the labelled cells innervating the quadriceps femoris tendon were found in the L3 and L4 DRGs. Many DRG cells were immunostained with CGRP antibody in the L1-6 DRGs. The number of CGRP immunoreactive cells in the lumbar DRGs was larger than in the sacral DRG. FG labelled cells containing CGRP immunoreactivity (FG+CGRP cells) were found in the lumbosacral DRGs. Many FG+CGRP cells innervating the quadriceps femoris tendon were located in the L3 and L4 DRGs. CONCLUSION: These results show that the main DRG origin for the sensory innervation of the quadriceps femoris tendon is L3 or L4. The neurogenic pain of the quadriceps femoris tendon may originate from this region, and suggests that this may be important for the release of neurogenic pain.
Subject(s)
Animals , Rats , Calcitonin Gene-Related Peptide , Diagnosis-Related Groups , Ganglia, Spinal , Horseradish Peroxidase , Immunohistochemistry , Quadriceps Muscle , Spinal Nerve Roots , TendonsABSTRACT
The origin of neuropeptide Y(NPY)-like immunoreactive(IR)fibers in the lateral and basolateral amygdaloid nuclei was examined by using indirect immunofluorescence and retrograde tracing method in the rat.1njection of a retrograde tracer.fluorogold(FG),into the lateral and basolateral amygdaloid nuclei,labeled many neurons in the ipsilateral anterior amygdaloid area,and simultaneous treatment with antiserum against NPY stained some of these neurons.Destruction of the anterior amygdaloid area caused an ipsilateral decrease of NPY-IR fibers in the lateral and basolateral amygdaloid nuclei.These findings indicate that NPY-IR neurons in the anterior amygdaloid area project ipsilaterally to the lateral and basolateral amygdaloid nuclei.In addition,the present study also shows that NPY-IR neurons located in the lateral and basolateral amygdaloid nuclei are intrinsic to the amygdaloid complex,since after destruction of the anterior amygdaloid area,some of NPY-IR fibers still can be found in lateral and baso lateral nuclei,and transection of the two major amygdalofugal system,stria terminalis and ventral amygdalofugal pathway,failed to cause the accumulation of NPY-like immunoreactivity in the axons proximal to the section.
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Objective To explore the fluorescent distribution, and the relation between fluorescent intensity and time after injecting rear-thigh muscles with DiI. Methods Sixty-five new born mice were equally and randomly divided into 13 groups. Fluorescent distribution and intensity were investigated at 2, 4, 6, 8, 12, 24 h, and 2, 4, 7, 14, 28 d and 2, 3 months after injecting the left posterior limb rear thigh muscles with 1.5 ?l 4 mg/ml DiI for one mouse. Results The labeling neurons were scattered from L2 level to S2 level and associated dorsal root ganglion (DRG), but the most were located at L4 to L6 section. The faint red fluorescence neurons were observed at dorsal root ganglion and cornu anterius medullae spinalis 6 h post-injection. The labeling neurons increased up to the 4th day. The fluorescent intensity enhanced gradually from 6 h to 24 h, then kept the intensity for 3 months. Conclusion It is a quick, precise, persistent method to trance and label the dorsal root ganglions sensory neuron and spinal motoneuron by injecting rear-thigh muscles with DiI, and the rate of labeling neurons can be improved by prolonging the tracing time properly. It is also provide basic data for clinical or experimental neuron label and location.
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The WGA-HRP retrograde tracing combined with immunohistochemical method was used to study the sources of CCK positive fibers in the male rat posterior pituitary. The CCK positive neurons projecting to the posterior pituitary localized mainly in the paraventricular nucleus, periventricular areas,and medial preoptic area. A few double labeled neurons were demonstrated in the subependymal area of the interventricular foramen and the floor of the 3rd ventricle. The CCK positive neurons projecting to the posterior pituitary, which weakly stained, were found in the bed nucleus of the stria terminalis, lateral hypothalamic area and the dorsal part of the supraoptic nucleus, too. Our result showed much more distribution of CCK-ir neurons projecting to the posterior pituitary in the hypothalamus than Palkovits′conclusion that the CCK positive fibers in posterior pituitary almost all originated from the magnocellular paraventricular nuclei. Some double labeled neurons in periventricular area were closely approximte to 3rd venticular cavity which suggested that these neurons may monitor the changes in the cerebrospinal fluid.
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Objective To estimate the effect of the NO on the medial amygdaloid nucleus(Me), we studied the origins of the NOS positive terminals in the Me. Methods Noergic afferent projection to Me was identified by a combined NADPH-d histochemical staning and retrograde CTb immunocytochemical method after microinjecting CTb into Me. Results The double labeled of neurons (NOS and CTb) were located in dorsal raphe nucleus, locus ceruleus, basolateral amygdaloid nucleus, parabrachial nucleus, ventrolateral part of periaqueductal gray.Conclution The NADPH-d positive terminals in the Me originates from the aforementioned nucleus, and may relate to the function of the Me.