Brainstem projections to spinal motoneurons: an update

1. The existence of direct projections to spinal motoneurons and interneurons from the raphe pallidus and obscurus, the adjoining ventral medial reticular formation and the locus coeruleus and subcoeruleus is now well substantiated by various anatomical techniques. 2. The spinal projections from the raphe nuclei and the adjoining medial reticular formation contain serotonergic and non-serotonergic fibres. These projections also contain various peptides, several of which are contained within the serotonergic fibres. Whether still other transmitter substances (e.g. acetylcholine) are present in the various descending brainstem projections to motoneurons remains to be determined. 3. The spinal projections from the locus coeruleus and subcoeruleus are mainly noradrenergic, but there also exists a non-noradrenergic spinal projection. 4. Pharmacological, physiological and behavioural studies indicate an overall facilitatory action of noradrenaline and serotonin (including several peptides) on motoneurons. This may lead to an enhanced susceptibility for excitatory inputs from other sources. 5. The brainstem areas in question receive an important projection from several components of the limbic system. This suggests that the emotional brain can exert a powerful influence on all regions of the spinal cord and may thus control both its sensory input and motor output.     

Cross-correlation of augmenting expiratory neurons of the Bötzinger complex in the cat

Ipsilateral and contralateral pairs of augmenting expiratory neurons were recorded simultaneously from the Bötzinger complex using glass-coated tungsten microelectrodes in pentobarbitone-anaesthetized cats. The neurons were identified both by firing pattern and by antidromic activation from the contralateral site of the dorsal respiratory group. Cross-correlation histograms of the extracellularly recorded action potentials were calculated in order to detect short time-scale synchronizations of firing indicative of synaptic connections between the neurons. The cross-correlation histograms for 40 ipsilateral pairs of neurons less than 1 mm apart showed eight (20%) narrow troughs (mean half-amplitude width ±SD, 1.1±0.37 ms) at short latencies (mean latency±SD, 1.0±0.35 ms) suggestive of monosynaptic inhibition. These included two cross-correlation histograms which showed troughs on both sides of time zero, indicating a mutual inhibition. For another four pairs of neurons (10%), a central broad peak suggestive of common activation due to either excitation or release from inhibition was evident. Contralateral pairs of expiratory neurons of the Bötzinger complex were examined in a similar manner. The cross-correlation histograms for 43 pairs of neurons showed five (12%) narrow troughs (mean half-amplitude width±SD, 1.2±0.67 ms) at short latencies (mean latency±SD, 2.7±1.47 ms) suggestive of monosynaptic inhibition. These included one cross-correlation histogram which showed troughs (one not statistically significant) on both sides of time zero, indicating a mutual inhibition. For another two pairs of neurons (4.6%) a central, broad peak suggestive of common activation due to either excitation or release from inhibition was evident. We conclude that inhibitory interconnections exist between augmenting expiratory neurons of the Bötzinger complex ipsilaterally and contralaterally. These connections may synchronize the expiratory burst of activity within this population and assist in the patterning of the burst.     

Topographic principles in the spinal projections of serotonergic and non-serotonergic brainstem neurons in the rat

The spinal projections from the raphe-associated brainstem areas containing serotonergic neurons were studied with aldehyde-induced fluorescence in combination with the retrograde fluorescent tracer True Blue in the rat. This technique makes it possible to determine simultaneously the projections of individual neurons and to detect whether serotonin is present in the same neurons. After tracer injections into the spinal cord retrogradely labeled serotonergic and non-serotonergic neurons were found in the medullary raphe nuclei and adjacent regions and to a lesser extent in association with the dorsal and median raphe nuclei in the mesencephalon. Large True Blue injections that covered one side of the spinal cord at mid-cervical level labeled about 60% of the ipsilaterally situated serotonergic neurons in the medullary raphe regions while the corresponding figure contralaterally was about 25%. On both sides a larger number of labeled non-serotonergic neurons were found; these were sometimes located dorsal to, but often intermingled with, the serotonergic cells. While the serotonergic projection from the mesencephalon could not be labeled from injections below cervical levels, the labeling in more caudal brainstem regions exhibited only minor variations depending on the rostrocaudal level of the spinal segment injected. Furthermore, quantitative data from injections at different levels indicate that the majority of the spinal-projecting neurons traverse most of the length of the cord. Summarizing the results obtained from small injections restricted to subregions of the cord we feel that it is possible to distinguish three fairly distinct pathways for spinal projections from the medullary raphe and adjacent regions: The dorsal pathway originates mainly from cells in the caudal pons and rostral medulla oblongata (rostral part of nucleus raphe magnus, nucleus raphe magnus proper, nucleus reticularis gigantocellularis pars alpha and nucleus paragigantocellularis). This pathway, which contains a large non-serotonergic component, descends through the dorsal part of the lateral funiculus and terminates mainly in the dorsal horn at all spinal cord levels. The intermediate pathway is largely serotonergic with its cell bodies located within the arcuate cell group (situated just ventral and lateral to the pyramids very close to the ventral surface of the brainstem) and in the nucleus raphe obscurus and pallidus and terminates in the intermediate grey at thoracolumbar and upper sacral levels.(ABSTRACT TRUNCATED AT 400 WORDS)     

Vasoactive intestinal polypeptide (VIP)-containing neurons in the spinal cord of the rat and their projections

The distribution of vasoactive intestinal polypeptide (VIP)-like immunoreactive structures in the rat spinal cord and their projections were investigated by means of an immunofluorescent method. In the normal rat, a small number of VIP-positive fibers were observed in the superficial layer of the dorsal horn and in the lateral funiculus. With colchicine pretreatment, VIP-positive neurons were demonstrated in the lateral spinal nucleus (lsn) and in the lamina X (Rexed). Transections of the spinal cord at various levels revealed that some of the VIP neurons in the lsn might project to supraspinal areas via lateral funiculus.     

The perikaryal projections of rabbit spinal ganglion neurons

Summary Shape, length and width of the perikaryal projections of spinal ganglion neurons from adult rabbits fixed in situ by perfusion have been evaluated by means of serial section electron microscopy. The results thus obtained have been compared with those obtained by enzymatic removal of ganglionic connective tissue and satellite cells followed by direct observation of the true neuronal surface under the scanning electron microscope. The comparison has shown that the perikaryal projections exhibit a similar shape and similar size with both techniques.     

Morphology of the decrementing expiratory neurons in the brainstem of the rat

In anesthetized and artificially-ventilated rats, the morphological properties of decrementing expiratory (E-DEC) neurons were studied using intracellular recording and labeling with Neurobiotin. Sixteen E-DEC neurons were successfully labeled; ten of which were cranial motoneurons located in the facial (FN) and ambiguus (NA) nuclei. Two interneurons were labeled in the B?tzinger complex (BOT) and the ventral respiratory group (VRG) rostral to the obex, and the remaining four in the VRG caudal to the obex. All the interneurons had extensive intramedullary collaterals within the ventrolateral medulla. Terminal-like boutons were distributed ventral to the NA at the level of the BOT, both ventral to and within the NA at the level rostral to the obex and largely within the cell column tentatively designed as the ambiguous-retroambiguus complex (NA/NRA) caudal to the obex. The four interneurons in the NA/NRA had axons projecting to the spinal cord as well. The extensive intramedullary projections suggest that these E-DEC interneurons of the BOT and the VRG play a significant role in respiration. The simultaneous projections from the caudal E-DEC neurons to both the spinal cord and the NA suggest that these neurons also play integrative roles in non-respiratory behaviors including vocalization, swallowing and defecation.     

Collateral projections of trigeminal ganglion neurons to both the principal sensory trigeminal and the spinal trigeminal nuclei in the rat

Employing a combination of fluorescent retro grade double labelling and immunofluorescence histo chemistry for substance P (SP) and calcitonin gene-relat ed peptide (CGRP), we examined collateral projections from single neurons in the trigeminal ganglion (TG) of the rat to both the principal sensory trigeminal nucleus (Vp) and the oral, interpolar or caudal subnuclei of the spinal trigeminal nucleus (Vo, Vi or Vc). In the rats that were unilaterally injected with fast blue (FB) into the Vp and with diamidino yellow (DY) into the Vo, Vi or Vc, neurons labelled with FB and/or DY were observed in the TG ipsilateral to the injections. Of the labelled TG neurons, about 2% were double labelled with both trac ers in the rats that were injected with FB into the Vp and with DY into the Vo or Vi, and about 10% were double labelled in the rats that were injected with FB into the Vp and with DY into the Vc. The results indicate that TG neurons sending their axons to the Vp project, by way of axon collaterals, to the Vc more frequently than to the Vo or Vi.Some of the TG neurons double labelled with FB and DY exhibited SP-or CGRP-like immunoreactivity (LI): Of the TG neurons that were double labelled with FB injected into the Vp and with DY injected into the Vo, Vi or Vc, about 38%, 49% and 42%, respectively, displayed SP-LI, and about 54%, 58% and 59%, respectively, showed CGRP-LI. Some of the SP-or CGRP-LI TG neurons that were double labelled with FB and DY were assumed to mediate pain signals to both the Vp and the spinal trigeminal nucleus (Vo, Vi and/or Vc) by way of axon collaterals.Yun-Qing Li is on leave from the Department of Anatomy, The Fourth Military Medical University, Xian, People's Republic of China     

Brainstem projections from the lateral hypothalamic area in the rat, as studied with autoradiography

Descending projections from the lateral hypothalamic area to the brainstem were studied, using [3H]-amino acid autoradiography, in the rat. Two main ipsilateral paths were reorganized. One is the periventricular fiber system projecting to the midbrain central gray. The other is a fiber system which eventually descends the central tegmental field, projecting strongly to the dorsal raphe nucleus, medial and lateral parabrachial nuclei, nucleus reticularis parvocellularis, solitary nuclei and dorsal motor nucleus of the vagus nerve. Sparse projections were observed to the nuclei raphe magnus, obscurus and pallidus, group B3 (or the ventrolateral subpial group) and spinal trigeminal nucleus.     

Axonal projections and conduction properties of olfactory peduncle neurons in the rat

Summary Electrophysiological methods were employed to study the axonal properties of the neurons of anterior olfactory nucleus (AON), transition zone (TZ), and rostral prepyriform cortex (RPPC) and their projections towards the ipsilateral and contralateral olfactory bulb (IOB, COB) in the rat.Of 91 antidromically driven cells, 39 (43%) and 32 (35%) responded to IOB and COB stimulation, respectively; 20 (22%) were discharged from both bulbs. Collision tests performed on the latter group indicated that these neurons have a short main axon which divides near the soma, projecting one branch to the COB and a thinner one toward the IOB. Mean conduction velocities of axons projecting to the IOB and the COB were 0.4 m/s and 0.7 m/s, respectively, the faster conducting axons having shorter refractory periods.Of the 38 neurons tested, 92% showed decreases in threshold and latency (up to 20% of control antidromic latency) after a test volley that was preceded by a conditioning pulse at intervals of 20–215 ms. Latency decreases were greater for slowly conducting axons than for the faster ones. These after-effects of impulse activity in OB afferent axons were attributed to the presence of a supernormal period of increased conduction velocity and excitability similar to that found in the olfactory nerve (Bliss and Rosenberg, 1974).This work was supported by a grant, Convenio de Cooperatión Científico-Tecnológica, Argentino-Norteamericano, No. 7363/ 75, provided by the Consejo Nacional de Investigaciones Científicas y Técnicas de Argentina     

Axonal projections from Bötzinger expiratory neurons to contralateral ventral and dorsal respiratory groups in the cat

Summary We studied projection patterns of the augmenting expiratory neurons of the Bötzinger complex (BÖT) in the contralateral brainstem. Three experimental approaches were used: 1) electrophysiological analysis using antidromic microstimulation, and morphological analyses using 2) intraaxonal injection of HRP, and 3) application of the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L). Taken together, the three methods revealed morphological details of the axonal arborizations of the expiratory neurons in the BÖT and the ventral respiratory group (VRG). The majority of augmenting expiratory neurons of the BÖT had axonal collaterals in the contralateral brainstem. The stem axons to the contralateral side crossed the midline almost at the level of the cell somata. They descended dorsomedial to the ventral spinocerebellar tract and gave off collateral branches directed dorsomedially. Terminal boutons were distributed abundantly in the caudal part of the BÖT and in the more caudally situated VRG. Axon collaterals sometimes ran to the dorsal respiratory group (DRG) and distributed terminal boutons there. Together with the fact of extensive ipsilateral arborizations shown previously, the present results indicate that the augmenting expiratory neurons of the BÖT have wide bilateral influence on the BÖT, VRG, DRG, and spinal cord.Abbreviations VII facial nucleus - XII hypoglossal nucleus - AMB nucleus ambiguus - AP area postrema - CX external cuneate nucleus - D descending vestibular nucleus - DX dorsal-motor nucleus of the vagus - M medial vestibular nucleus - NTS nucleus of the solitary tract - R nucleus of Roller - S solitary tract - RFN retrofacial nucleus This paper is dedicated to Professor Hajime Mannen on the occasion of his 65th birthdaySupported by grants-in-aid for Scientific Research nos. 60304044, 62570068, 62770043, and 63570027 from the Japan Ministry of Education, Science and Culture     

Association of spinal lamina I projections with brainstem catecholamine neurons in the monkey

In addition to giving primary projections to the parabrachial and periaqueductal gray regions, ascending lamina I projections course through and terminate in brainstem regions known to contain catecholaminergic cells. For this reason, double-labeling experiments were designed for analysis with light and electron microscopy. The lamina I projections in the Cynomolgus monkey were anterogradely labeled with Phaseolus vulgaris leucoagglutinin (PHA-L) and catecholamine-containing neurons were labeled immunocytochemically for tyrosine hydroxylase (TH). Light level double-labeling experiments revealed that the terminations of the lamina I ascending projections through the medulla and pons strongly overlap with the localization of catecholamine cells in: the entire rostrocaudal extent of the ventrolateral medulla (A1 caudally, C1 rostrally); the solitary nucleus and the dorsomedial medullary reticular formation (A2 caudally, C2 rostrally); the ventrolateral pons (A5); the locus coeruleus (A6); and the subcoerulear region, the Kölliker-Fuse nucleus, and the medial and lateral parabrachial nuclei (A7). At the light microscopic level, close appositions between PHA-L-labeled lamina I terminal varicosities and TH-positive dendrites and somata were observed, particularly in the A1, A5 and the A7 cell groups on the contralateral side. At the electron microscopic level, examples of lamina I terminals were found synapsing on cells of the ventrolateral catecholamine cell groups in preliminary studies. The afferent input relayed by these lamina I projections could provide information about pain, temperature, and metabolic state as described previously. Lamina I input could impact interactions of the catecholamine system with higher brain centers modulating complex autonomic, endocrine, sensory, motor, limbic and cortical functions such as memory and learning. Nociceptive lamina I input to catecholamine cell regions with projections back to the spinal cord could form a feedback loop for control of spinal sensory, autonomic and motor activity.     

Age-related decrease in the overall extent of perikaryal projections in rabbit spinal ganglion neurons

The overall extent of the perikaryal projections of sensory neurons from spinal ganglia of young adult and aged rabbits was estimated by a stereological method using the electron microscope. The extent of perikaryal projections was significantly smaller in the aged animals. This age-related decrease did not seem to depend on factors intrinsic to the neuron, but on the absence of a satellite cell covering over extensive portions of the nerve cell body surface. This decrease may influence the organization of the subplasmalemmal cytoskeleton, metabolic exchange between the nerve cell body and its environment and perhaps also neuronal metabolism.     

Bötzinger-complex expiratory neurons monosynaptically inhibit phrenic motoneurons in the decerebrate rat

We examined respiratory neurons in the Bötzinger complex of the medulla oblongata in 18 vagotomized, paralyzed, ventilated, and decerebrated rats and tested the hypothesis that bulbospinal expiratory neurons in this region monosynaptically inhibit phrenic motoneurons. First, we surveyed the types of respiratory neurons found in the Bötzinger complex; only 11 of the 98 (～11%) examined were bulbospinal, and all discharged only during late expiration (E2), usually with an augmenting discharge frequency (AUG). Then, we examined the spinal projections of 34 E2-AUG neurons using antidromic activation and found that all projected as far as the C4 or C5 segments of the spinal cord but no further caudally. Most (30, ～88%) had only unilateral projections, the majority (25, ～83%) ipsilateral, but 4 neurons (～12%) had bilateral projections. Their axons could be antidromically activated at low currents (less than 10 μA) in the dorsal-lateral part of the spinal cord at the C2–3 border; 0.5–1.2 mm (mean±SD 0.84±0.23 mm) below the dorsal surface and 0.7–1.5 mm (1.19±0.25 mm) lateral from the midline. We sought evidence for connections from bulbospinal E2-AUG neurons to 118 phrenic motoneurons by computing spike-triggered averages (STAs) of their intracellular potentials triggered by the action potentials of 38 unilaterally-projecting E2-AUG neurons. Resting phrenic motoneuron membrane potentials ranged from –40 to –75 mV (–56±8 mV) and fluctuations with the respiratory cycle from 7 to 20 mV (14±4 mV). Of the 118 STAs computed, hyperpolarizations were evident in 18 (～15%) STAs, evoked by 11 of 38 (～29%) E2-AUG neurons. Their amplitudes varied from 35 to 550 μV (105±113 μV), 10–90% fall times from 0.4 to 0.9 ms (0.63±0.17 ms), and half-amplitude widths from 1.3 to 3.2 ms (2.0±0.52 ms). Most (16/95, ～17%) of the STAs that displayed hyperpolarizations were associated with ipsilateral trigger neurons but some (2/23, ～9%) resulted from contralateral trigger neurons. We conclude that Bötzinger-complex, expiratory neurons project to the C4 and/or C5 segments of the cervical spinal cord but no further caudal. Their axons are located dorsolaterally in the upper cervical segments of the spinal cord, and they monosynaptically inhibit phrenic motoneurons during the late part of expiration.     

Cortical projections of monoamine oxidase-containing neurons from the posterior hypothalamus in the rat

The dipeptide, L-phenylalanyl-L-glutamate (PG), augments the specific binding of the excitatory amino acid receptor antagonist, [3H]2-amino-7-phosphonoheptanoic acid (APH), to rat forebrain membranes by 5-fold at 100 microM with an EC50 of 4.9 microM. The increase in the specific binding of [3H]AHP induced by PG results exclusively from an increase in Bmax. In contrast, PG inhibits the specific binding of [3H]kainic acid to forebrain membranes with a Ki of 6.8 microM. Of several related peptides examined, active ones affected the two receptor sites in a reciprocal fashion. The results suggest an allosteric interaction between [3H]APH and kainate receptors modulated by glutamate-containing peptides.     

Perikaryal projections of developing spinal ganglion neurons in the chick demonstrated by scanning electron microscopy

 The perikaryal projections of sensory ganglion neurons in chick embryos were observed by scanning electron microscopy after removal of the connective tissues and satellite cells by enzymatic digestion treatment. The perikaryal projections were seen not only on the surface of the perikarya but also on the surface of the stem processes. The projections were up to 3 μm in length, and their transverse diameters ranged between 0.12 and 0.24 μm from incubation (embryonic) day 10 to posthatching day 2. On days 6 and 8 of incubation, thicker projections with transverse diameters of 0.24–0.9 μm were observed transiently in addition to those described above, and some of them looked like vestiges of neuronal processes during development. The thin projections emerging mainly in the later developmental stages increased in number as spindle-shaped bipolar neurons differentiated into (pseudo)unipolar cells. Morphometric analysis revealed that the density of perikaryal projections correlated well with the shape and size of each neuron; thin perikaryal projections were more numerous on those of mature pseudounipolar neurons than on the surface of premature ganglion neurons, and they increased in number as the individual ganglion cell bodies grew larger. The neuronal shape- and size-dependent increase in perikaryal projections during development may support the hypothesis that perikaryal projections are structural devices for increasing neuronal surface areas and possibly the efficiency of metabolic activities. Accepted: 18 June 1996     

Distribution of neurons of origin of zinc-containing projections in the amygdala of the rat

The present study describes the distribution of neurons of origin of zinc-containing pathways in the amygdaloid complex of the rat, using the selenium method for simultaneous retrograde labeling of all zinc-containing neurons. With this method, vesicular ionic zinc is precipitated intravitally with selenium compounds and transported retrogradely to the parent neurons, where it can be visualized by silver amplification. Neurons labeled retrogradely with silver-amplified precipitate were observed in all amygdaloid nuclei except for the lateral olfactory tract nucleus, the accessory olfactory tract nucleus and the central nucleus. Very few labeled cell bodies were seen in the anterior amygdaloid area and the medial nucleus. The amygdalo-hippocampal area and the amygdalo-piriform transition area both showed a substantial number of labeled somata throughout their rostrocaudal extent. In the anterior cortical nucleus, very few labeled cell bodies were found in the rostral pole, whereas they were abundant in the caudal quarter of the nucleus. In the posterolateral cortical nucleus, the number of labeled cell bodies increased gradually; there were none in the rostral pole, but most of the neurons in the caudal part were labeled. The posteromedial cortical nucleus contained a great number of labeled somata, but with some variation in the rostrocaudal extent of the nucleus. Considerable numbers of labeled neurons were observed throughout the lateral nucleus. In the basolateral nucleus, a small number of labeled cell bodies was present in the rostral half, but a gradual increase was observed in the caudal direction. Finally, in the basomedial nucleus, very few labeled cell bodies were present in the rostral two-thirds, whilst a considerable number was encountered in the caudal one-third. Possible functional implications of neuronal zinc are considered. The distribution of neurons of origin of zinc-containing projections has been compared with previously described intrinsic connections of the rat amygdala, and tracts that may possibly be zinc-containing are outlined and discussed. It is concluded that in all probability a substantial proportion of the intrinsic connectivity of the rat amygdaloid complex is zinc-containing.The authors thank Ms. M. Sørensen, Mrs. A. Lyhr, Mr. A. Meier and Mrs. K. Wiedemann for excellent technical help.     

GABA-immunoreactive neurons in the dorsal horn of the rat spinal cord

An antiserum to GABA was used on semithin resin-embedded sections of rat dorsal horn. Immunoreactive neurons were evenly distributed throughout laminae I-III and constituted between 24 and 33% of the total neuronal population within three laminae. Fifty Golgi-stained cells in lamina II were tested with the antiserum. Most of the islet cells examined were immunoreactive, although some small islet cells were not. None of the 14 stalked cells tested was immunoreactive. These results provide further evidence that the stalked and islet cells of lamina II form two distinct functional classes and suggest that the islet cells function as inhibitory interneurons.     

手阳明大肠经和手厥阴心包经相关的脊髓前角运动神经元投射支配特性

目的绘制支配手阳明大肠经和手厥阴心包经相关的脊髓运动神经元在脊髓的空间定位和树突投射的特异性分布。方法利用霍乱毒素B单位结合辣根过氧化物酶(CB-HRP)可被外周神经末梢吸收和运输的特性,在大鼠前肢手阳明大肠经和手厥阴心包经路径上的肌肉的每一穴位点分别注射5μL的CB-HRP(1%)溶液,经过3~4 d的药物吸收后,通过灌流固定-切片-TMB组织化学反应程序,显示出支配这些肌肉的运动神经元胞体及突起,并观察手阳明大肠经和手厥阴心包经相关的运动神经元在脊髓的空间的分布特性。结果大鼠手阳明大肠经以及手厥阴心包经相关的运动神经元在颈3-颈7节段脊髓腹角分别形成首尾走向、定位明确的运动神经元柱状联结;同一条经对应的运动神经元之间存在丰富的树突-树突和树突-胞体定向投射,形成一个由运动神经元胞体及其树突构成的细胞柱状实体。结论同经相关的运动神经元之间构成一个紧密连接的神经元柱,柱间的神经元的树突-树突和树突-胞体定向投射可能是循经感传反射活动的结构基础。     

胚胎大鼠脊髓运动神经元的体外培养及鉴定

本实验建立了体外胚胎大鼠脊髓运动神经元的分离培养,并进行鉴定。取孕15d SD大鼠胚胎的脊髓腹侧组织,用胰蛋白酶和胶原酶消化分离成单细胞悬液,进行原代细胞培养,应用神经元特异性的烯醇化酶(NSE)抗体和抗神经微丝抗体-SMI32单克隆抗体对培养细胞进行鉴定,同时行尼氏染色。结果分离培养的细胞鉴定为脊髓运动神经元(SMN),SMN在体外适宜的条件下可存活4周左右,可作为神经科学研究的一种手段。