Proper development of relay somatic sensory neurons and D2/D4 interneurons requires homeobox genes Rnx/Tlx-3 and Tlx-1

Trigeminal nuclei and the dorsal spinal cord are first-order relay stations for processing somatic sensory information such as touch, pain, and temperature. The origins and development of these neurons are poorly understood. Here we show that relay somatic sensory neurons and D2/D4 dorsal interneurons likely derive from Mash1-positive neural precursors, and depend on two related homeobox genes, Rnx and Tlx-1, for proper formation. Rnx and Tlx-1 maintain expression of Drg11, a homeobox gene critical for the development of pain circuitry, and are essential for the ingrowth of trkA+ nociceptive/thermoceptive sensory afferents to their central targets. We showed previously that Rnx is necessary for proper formation of the nucleus of solitary tract, the target for visceral sensory afferents. Together, our studies demonstrate a central role for Rnx and Tlx-1 in the development of two major classes of relay sensory neurons, somatic and visceral.     

The generation of granule cells during the development and evolution of the cerebellum

The cerebellum coordinates vestibular input into the hindbrain to control balance and movement, and its anatomical complexity is increasingly viewed as a high-throughput processing center for sensory and cognitive functions. Cerebellum development however is relatively simple, and arises from a specialized structure in the anterior hindbrain called the rhombic lip, which along with the ventricular zone of the rostral-most dorsal hindbrain region, give rise to the distinct cell types that constitute the cerebellum. Granule cells, being the most numerous cell types, arise from the rhombic lip and form a dense and distinct layer of the cerebellar cortex. In this short review, we describe the various strategies used by amniotes and anamniotes to generate and diversify granule cell types during cerebellar development.     

Morphology and physiology of the cerebellar vestibulolateral lobe pathways linked to oculomotor function in the goldfish

Intracellular recording and single-cell labeling were combined to investigate the oculomotor circuitry of the goldfish cerebellar vestibulolateral lobe, consisting of the eminentia granularis (Egr) and caudal lobe. Purkinje cells exhibiting highly conserved vertebrate electrophysiological and morphological properties provide the direct output from the caudal lobe to the vestibular nuclei. Biocytin labeling of the Egr distinguished numerous hindbrain precerebellar sources that could be divided into either putative mechano- or vestibulosensitive nuclei based on cellular location and axon trajectories. Precerebellar neurons in a hindbrain nucleus, called Area II, were electrophysiologically characterized after antidromic activation from the Egr (>50% bilateral) and their morphology analyzed after intracellular biocytin labeling (n = 28). Bipolar spindle-shaped somas ranged widely in size with comparably scaled dendritic arbors exhibiting largely closed field configuration. Area II neurons (85%) projected to the ipsilateral Egr with most (93%) sending a collateral through the cerebellar commissure to the contralateral Egr; however, 15% projected to the contralateral Egr by crossing in the ventral hindbrain. Axon terminals in the vestibular nucleus were the only collaterals within the hindbrain. Every Area II neuron received a disynaptic EPSP after contralateral horizontal canal nerve stimulation and a disynaptic IPSP, preceded by a small EPSP (>50%), after ipsilateral activation. Vestibular synaptic potentials were of varying shape/amplitude, unrelated to neuron location in the nucleus, and thus likely a correlate of somadendritic size. The exceptional separation of eye position and eye velocity signals into two separate hindbrain nuclei represents an ideal model for understanding the precerebellar projection to the vestibulocerebellum.     

Localization of NADPH-diaphorase containing neurons in the spinal dorsal horn and spinal sensory ganglia of the turtle Chrysemys d’orbigny

 NADPH-diaphorase positive (NDP) neurons and nerve fibers were found in the spinal dorsal horn (DH) and sensory ganglia of the turtle Chrysemys d’orbigny. Three well-defined types of NDP neurons were found in the DH: (a) elongated nerve cells with two radially arranged dendritic branches, (b) neurons with rostro-caudal dendritic branches, (c) bitufted neurons with two, practically symmetric branches that project to the ipsilateral and contralateral dorsal horns. A combination of the techniques that reveal NADPH-diaphorase activity with the horseradish peroxidase transganglionic labeling of the dorsal root collaterals, suggested that NDP neurons of the DH are second-order cells of the spinal sensory pathway. NDP neurons were also found in the spinal sensory ganglia at all metameric levels. Our findings indicate that the DH of turtles, like that of mammals, contains both the enzymatic machinery and the neural connections required to postulate the participation of nitric oxide in ”plastic phenomena” such as hyperalgesia and central sensitization. Two other alternatives or complementary hypotheses are discussed: (a) NDP neurons in the DH and sensory ganglia may represent specific cell populations involved in the processing of sensory visceral information; (b) NADPH-diaphorase reactivity may indicate sustained levels of neuronal activity. Received: 12 February 1996 / Accepted: 2 August 1996     

大鼠三叉神经脊束核尾侧亚核和脊髓向丘脑和臂旁核的强啡肽能和一氧化氮能投射

本文用荧光金逆行追踪与免疫荧光组化染色相结合的方法，对大鼠三叉神经脊束核尾侧亚核和颈髓背角浅层向丘脑腹基底复合体和臂旁核的强啡肽能和NO能投射进行了研究．强啡肽原前体样阳性胞体主要位于尾侧亚核和颈髓背角的Ⅰ层和Ⅱ层外侧部；NOS样阳性胞体主要位于尾侧亚核和颈够背角Ⅱ层，Ⅰ层较少。将荧光金注入丘脑腹基底复合体后，荧光金逆标神经元主要见于对侧尾侧亚核、颈髓背角的Ⅰ层和外侧网状核，Ⅱ层偶见；将荧光金注入臂旁核后，逆标神经元主要见于同侧尾侧亚核和颈髓背角的Ⅰ、Ⅱ层，少量位于外侧网状核。尾侧亚核向丘脑瓜基底复合体投射神经元的16．6％，向臂旁核投射神经元的24．8％呈强啡肽原前体样阳性；颈髓背角浅层向丘脑腹基底复合体投射神经元的19．2％，向臂旁核投射神经元的272％呈强啡肽原前体样阳性。向丘脑腹基底复合体和臂旁核投射的强啡肽原前体／荧光金双标神经元分别占尾侧亚核浅层内强啡肽原前体样阳性神经元总数的7％和18％，分别占颈髓背角浅层内强啡肽原前体样阳性神经元总数的8．1％和21．9％。这些双标神经元多呈大梭形及中等大圆形和梨形。由昆侧亚核向丘脑腹基底复合体投射神经元的5．1％呈NOS阳性，向臂旁核投射神经元的11．8％呈NOS阳性。由颈髓背角浅层向丘脑版?     

The bHLH factor Olig3 coordinates the specification of dorsal neurons in the spinal cord

Neurons of the dorsal horn integrate and relay sensory information and arise during development in the dorsal spinal cord, the alar plate. Class A and B neurons emerge in the dorsal and ventral alar plate, differ in their dependence on roof plate signals for specification, and settle in the deep and superficial dorsal horn, respectively. We show here that the basic helix-loop-helix (bHLH) gene Olig3 is expressed in progenitor cells that generate class A (dI1-dI3) neurons and that Olig3 is an important factor in the development of these neuronal cell types. In Olig3 mutant mice, the development of class A neurons is impaired; dI1 neurons are generated in reduced numbers, whereas dI2 and dI3 neurons are misspecified and assume the identity of class B neurons. Conversely, Olig3 represses the emergence of class B neurons in the chick spinal cord. We conclude that Olig3 expression distinguishes the two major classes of progenitors in the dorsal spinal cord and determines the distinct specification program of class A neurons.     

内脏大神经初级传入神经元向中枢投射的光镜和电镜研究——CT-HRP跨神经节运送法

用猫20只,于内脏大神经注射CT—HRP,对初级传入在中枢内的投射进行了光、电镜观察。光镜下,内脏初级传入神经元的中枢投射纤维主要经Lissauer's束到脊髓后角边缘,部分可能终止于Ⅰ层,大部分分为内、外侧束包绕灰质后角边缘由浅部板层向深部板层进入Ⅴ、Ⅶ、Ⅹ层。内侧束可能有部分上升到薄束核。电镜观察,在Ⅰ、Ⅴ、Ⅶ、Ⅹ层看到标记的轴突末梢且可直接与交感节前神经元形成突触。本文还对内脏传入通路进行了讨论并认为交感传入纤维可能经脊髓后角神经元中继上传至孤束核。     

Parasagittal compartmentation of cerebellar mossy fibers as revealed by the patterned expression of vesicular glutamate transporters VGLUT1 and VGLUT2

The cerebellum receives sensory signals from spinocerebellar (lower limbs) and dorsal column nuclei (upper limbs) mossy fibers. In the cerebellum, mossy fibers terminate in bands that are topographically aligned with stripes of Purkinje cells. While much is known about the molecular heterogeneity of Purkinje cell stripes, little is known about whether mossy fiber compartments have distinct molecular profiles. Here, we show that the vesicular glutamate transporters VGLUT1 and VGLUT2, which mediate glutamate uptake into synaptic vesicles of excitatory neurons, are expressed in complementary bands of mossy fibers in the adult mouse cerebellum. Using a combination of immunohistochemistry and anterograde tracing, we found heavy VGLUT2 and weak VGLUT1 expression in bands of spinocerebellar mossy fibers. The adjacent bands, which are in part comprised of dorsal column nuclei mossy fibers, strongly express VGLUT1 and weakly express VGLUT2. Simultaneous injections of fluorescent tracers into the dorsal column nuclei and lower thoracic–upper lumbar spinal cord revealed that upper and lower limb sensory pathways innervate adjacent VGLUT1/VGLUT2 parasagittal bands. In summary, we demonstrate that VGLUT1 and VGLUT2 are differentially expressed by dorsal column nuclei and spinocerebellar mossy fibers, which project to complementary cerebellar bands and respect common compartmental boundaries in the adult mouse cerebellum.     

在体和离体的鸡背根节神经细胞5-羟色胺的免疫组织化学研究

用PAP免疫组织化学方法研究了鸡胚和雏鸡背根节内5-HT神经细胞的形态与胚胎发育,并在离体细胞培养条件下研究了靶周围组织(皮肤)对其胚胎发育的影响。在鸡腰骶部背根节,5-HT免疫反应阳性细胞最先出现于鸡胚13d(E_(13)),占0.4%;孵出后2d(AH_2)占6.2%。阳性细胞主要为大的A类细胞和极少的小B类细胞。免疫反应阳性的周围神经末梢位于皮肤和跟腱中。取自E_9鸡胚的背根节细胞培养7d后有部分出现5-HT阳性免疫反应,取自E_6鸡胚的背根节细胞培养10d仍为阴性反应;然而,当取自E_6鸡胚的背根节细胞与皮肤组织联合培养10d后,则出现5-HT阳性免疫反应细胞。本文还对脊髓5-HT细胞的出现进行了讨论。     

Differential spinal projections from the forelimb areas of the rostral and caudal subregions of primary motor cortex in the cat

We used anterograde transport of WGA-HRP to examine the topography of corticospinal projections from the forelimb areas within the rostral and caudal motor cortex subregions in the cat. We compared the pattern of these projections with those from the somatic sensory cortex. The principal finding of this study was that the laminar distribution of projections to the contralateral gray matter from the two motor cortex subregions was different. The rostral motor cortex projected preferentially to laminae VI–VIII, whereas caudal motor cortex projected primarily to laminae IV–VI. Confirming earlier findings, somatic sensory cortex projected predominantly to laminae I–VI inclusive. We found that only rostral motor cortex projected to territories in the rostral cervical cord containing propriospinal neurons of cervical spinal segments C3-4 and, in the cervical enlargement, to portions presumed to contain Ia inhibitory interneurons. We generated contour maps of labeling probability on averaged segmental distributions of anterograde labeling for all analyzed sections using the same algorithm. For rostral motor cortex, heaviest label in the dorsal part of lamina VII in the contralateral cord was consistently located in separate medial and lateral zones. In contrast, no consistent differences in the mediolateral location of label was noted for caudal motor cortex. To summarize, laminae I–III received input only from the somatic sensory cortex, while laminae IV–V received input from both somatic sensory and caudal motor cortex. Lamina VI received input from all cortical fields examined. Laminae VII–IX received input selectively from the rostral motor cortex. For motor cortex, our findings suggest that projections from the two subregions comprise separate descending pathways that could play distinct functional roles in movement control and sensorimotor integration.     

Activity during mastication of periodontal mechanosensitive neurons of the trigeminal subnucleus oralis of the rabbit

1. The activity of mechanosensitive neurons was examined before and during mastication. One hundred and seventy-eight neurons were recorded in the rostral parts of the trigeminal sensory nuclei of 20 rabbits anesthetized with urethan. Twenty-eight neurons received inputs from the periodontal mechanoreceptors, all on the ipsilateral side. Nineteen had receptive fields that were restricted to one tooth; 2 could be activated from more than 1 tooth, and 6 included parts of the mucosa. Only the latter were spontaneously active. 2. All periodontal neurons with a mandibular input responded to graded electrical stimulation of the inferior alveolar nerve at minimum latencies of less than or equal to 3.4 ms, and approximately half had inputs from the sensorimotor cortex. 3. Almost all periodontal units recorded were found to lie in, or just outside, the dorsal part of the most rostral subdivision of the spinal trigeminal nucleus (subnucleus oralis, pars gamma). None projected to the ipsi- or contralateral thalamus. 4. All periodontal neurons fired during mastication. Those without mucosal receptive fields fired during jaw closure, with almost all activity confined to the slow-closing phase when pressure is applied to the teeth. Injections of local anesthetic showed that input from mucosal fields was responsible for activating neurons in other phases of the cycle. 5. Possible roles in the control of mastication for these periodontal interneurons were discussed.     

Nerve injury increases an excitatory action of neuropeptide Y and Y2-agonists on dorsal root ganglion neurons

Damage to sensory nerves invokes the expression of neuropeptide Y in the cell bodies of sensory neurons in dorsal root ganglia. We therefore compared the action of this peptide on control dorsal root ganglia neurons with its action on neurons from animals in which the sciatic nerve had been cut. Neuropeptide Y (0.1-1.0 microM) increased the excitability of 24% of control neurons and its effect was stronger and more cells (56%) were affected after axotomy. Increased excitability was mediated via a Y2-receptor and resulted from attenuation of Ca2+-sensitive K+-conductance(s) secondary to suppression of N-type Ca2+ channel current. Y1-agonists potentiated L-type Ca2+ channel current in control neurons without altering excitability. This Y1-effect was attenuated whereas effects mediated via Y2-receptors were enhanced after axotomy. No evidence was found for involvement of Y4- or Y5-receptor subtypes in the actions of neuropeptide Y either on control or on axotomized dorsal root ganglion neurons. It is concluded that neuropeptide Y increases the excitability of sensory neurons by interacting with a Y2-receptor and thereby decreasing N-type Ca2+ channel current and Ca2+-sensitive K+-conductance(s). When peripheral nerves are damaged, dorsal root ganglion neurons start to express neuropeptide Y and its excitatory Y2-excitatory effects are enhanced. The peptide may therefore contribute to the generation of aberrant sensory activity and perhaps to the etiology of injury-induced neuropathic pain.     

Complementary immunohistochemical distribution of the neurofilament triplet and novel intermediate filament proteins in the autonomic and sensory nervous system of the guinea-pig

We have previously established that immunoreactivity for the triplet of polypeptides that comprise the class IV intermediate filament proteins (NFP-triplet) is localized in specific subpopulations of neurons in guinea-pig sensory and autonomic ganglia. Antibodies to novel neurofilament proteins, including a polyclonal antibody to a 57 kDa neuronal intermediate filament polypeptide (NIF57kD) and a monoclonal antibody (CH1) to a 150 kDa intermediate filament, or associated, protein were used in combination with antibodies to the NFP-triplet for double-labelling immunohistochemistry. The results show that different subpopulations of neurons in the guinea-pig dorsal root ganglia, coeliac ganglion and enteric ganglia can be distinguished by their complementary immunoreactivity for these proteins. In dorsal root ganglia, larger neurons are intensely immunoreactive for the NFP-triplet while immunoreactivity with CH1 and NIF57kD antibodies is restricted to the small to medium-sized neurons. In the coeliac ganglion, two regionally defined subpopulations of neurons can be distinguished by their immunoreactivity for either the NFP-triplet or NIF57kD, whereas CH1 labels all neurons with equal intensity. Three classes of morphologically distinct myenteric neuron subpopulations are also distinguished by their immunoreactivity for either the NFP-triplet, NIF57kD or CH1 antibodies. Two classes of submucous neurons are labelled both with CH1 and NIF57kD antibodies but show faint or no immunoreactivity for the NFP-triplet. It is concluded that intermediate filament protein immunoreactivity marks different subpopulations of neurons, which suggests that these proteins may have specific roles in neuronal function.     

Rat trigeminal lamina I neurons that project to thalamic or parabrachial nuclei contain the mu-opioid receptor

Ligands of the mu-opioid receptor are known to inhibit nociceptive transmission in the dorsal horn, yet the cellular site(s) of action for this inhibition remain to be fully elucidated. Neurons located in lamina I of the dorsal horn are involved in distinct aspects of nociceptive transmission. Neurons projecting to the thalamus are thought to be involved in sensory-discriminative aspects of pain perception, while neurons projecting to the parabrachial nucleus are thought to be important for emotional and/or autonomic responses to noxious stimuli. The present study examined these two populations of lamina I projection neurons in the trigeminal dorsal horn to determine if the mu-opioid receptor protein (MOR1) is differentially located in these populations of neurons. Lamina I projection neurons were identified using the retrograde tracer FluoroGold (FGold). FGold was injected into either the contralateral thalamus (ventral posterolateral (VPM)/ventral posterolateral (VPL) thalamic region) or into the ipsilateral parabrachial nuclei. The distribution of MOR1 in these neurons was determined using immunocytochemistry. The distribution of MOR1-ir within these two populations of lamina I projection neurons was examined by both confocal and electron microscopy. We found that both populations of projection neurons contained MOR1. Immunogold analyses revealed the presence of MOR1-ir at membrane sites and within the cytoplasm of these neurons. Cytoplasmic receptor labeling may represent sites of synthesis, recycling or reserve populations of receptors. MOR1 was primarily found in the somata and proximal dendrites of projection neurons. In addition, these neurons rarely received synaptic input from MOR1-containing axon terminals. These results indicate that lamina I neurons in trigeminal dorsal horn that project to the thalamic and parabrachial nuclei contain MOR1 and are likely sites of action for MOR ligands that modulate sensory and/or autonomic aspects of pain transmission in the trigeminal dorsal horn.