Distribution of phenylethanolamine N-methyltransferase cell bodies, axons, and terminals in monkey brainstem: an immunohistochemical mapping study.

Adrenaline (epinephrine) is an important candidate transmitter in descending spinal control systems. To date intrinsic spinal adrenergic neurons have not been reported; thus adrenergic input is presumably derived from brainstem sites. In this regard, the localization of adrenergic neurons in the brainstem is an important consideration. Maps of adrenergic cell bodies and to a lesser extent axons and terminal fields have been made in various species, but not in monkeys. Thus, the present study concerns the organization of adrenergic systems in the brainstem of a monkey (Macaca fascicularis) immunohistochemically mapped by means of an antibody to the enzyme phenylethanolamine N-methyltransferase (PNMT). PNMT-immunostained cell bodies are distributed throughout the medulla in two principal locations. One concentration of labeled cells is in the dorsomedial medulla and includes the nucleus of the solitary tract (NTS), the dorsal motor nucleus of the vagus (X), and an area ventral to X in a region of the reticular formation (RF) known as the central nucleus dorsalis (CnD) of the medulla. A few scattered cells are observed in the periventricular gray just ventral to the IVth ventricle and on midline in the raphe. The second major concentration of PNMT-immunostained cells is located in the ventrolateral RF, lateral and dorsolateral to the inferior olive (IO), including some cells in the rostral part of the lateral reticular nucleus (LRN). Terminal fields are located in the NTS, X, area postrema (AP), and the floor of the IVth ventricle in the medulla and pons. A light terminal field is also observed in the raphe, particularly raphe pallidus (RP). A heavy terminal field is present in locus coeruleus (LC). Fibers labeled for PNMT form two major fiber tracts. One is in the dorsomedial RF extending as a well-organized bundle through the medulla, pons, and midbrain. A second tract is located on the ventrolateral edge of the medulla and caudal pons. Fibers in this tract appear to descend to the spinal cord. A comparison with maps of other catecholamine neurons in primates is discussed, confirming that the distribution of the adrenergic system in monkeys is similar to that described in the human.

Nerve fibre morphometry: a comparison of techniques.

This study compared different approaches to measuring nerve axon and fibre diameters and areas from transverse sections. A mock photomicrograph and mock tissue section, each with 100 identical, circular 'fibres' was constructed. Three measurement protocols were investigated: (A) circular approximation from minimum diameter; (B) circular approximation from the mean of orthogonal diameters; and (C) calculation of diameter and area from a digitized circumference. For each protocol, all 100 fibres on the photomicrograph were repeatedly measured using a digitizing tablet. Similarly, the fibres on the mock tissue section were measured using a digitizing tablet and microscope with camera lucida. The variance for these data was calculated. Protocols were compared on the basis of variability and the amount of digitizing time required. For diameter measurements, protocol B showed significantly lower variability than A or C (P less than 0.05), with only a modest increase in digitizing time over A. For area measurements, protocols B and C showed significantly lower variability than A (P less than 0.05), again with a modest increase in digitizing time. Measurements made using the microscope and camera lucida showed significantly lower variability than those made from the photomicrograph, but took more time. These data suggest that for diameter measurements, a mean of orthogonal diameters approach is best, and that for area measurements, a traced circumference approach is best as it is more flexible than the orthogonal diameter approach. While the microscope and camera lucida setup is more time-consuming to use, it eliminates the need for photomicrograph production.

Intracellular labeling of afferents to the lateral superior olive in the bat, Eptesicus fuscus.

An in vitro tissue slice preparation of the bat brain stem was used to label intracellularly individual axons projecting to the lateral superior olive from two different sources: the medial nucleus of the trapezoid body (MNTB) and the anteroventral cochlear nucleus (AVCN). The tracing of individually labeled MNTB axons into the lateral superior olive reaffirms the long accepted indirect route by which information from the contralateral ear reaches the lateral superior olive. While the MNTB appears to relay input from the contralateral AVCN, information from the ipsilateral ear reaches the lateral superior olive via a direct projection from the ipsilateral AVCN. Axons from the contralateral and ipsilateral pathways have different distribution patterns upon the fusiform cells of the lateral superior olive. Axon terminals of MNTB principal cells have a perisomatic and proximal dendritic distribution pattern. Axon terminal varicosities from the ipsilateral anteroventral cochlear nucleus are distributed primarily to more distal dendrites.

Withdrawal of collaterals of sympathetic axons to the rat eye during postnatal development: the role of function.

Neurones in the superior cervical ganglion (SCG) of the rat can be retrogradely labelled by injection of Fast blue (FB) into the anterior eye chamber with the maximum number being labelled during the second postnatal week. In adult rats, however, many fewer neurones can be so labelled. In the present study, we have investigated whether this reduction may be due to the withdrawal of collaterals of neurones which project to the posterior eye compartments. For these experiments, we have used either one (FB) or two (FB and horseradish peroxidase (HRP)) retrogradely transported markers injected with a micropipette into the anterior or posterior (vitreous) parts of the eye in rats aged 14, 31 and greater than 50 days. Using FB, we have shown that in 14-day-old rats, 40% of the neurones which project to the eye have collaterals in anterior and posterior compartments while only 1% of neurones maintain dual projections in animals aged more than 7 weeks. Furthermore, there is a reduction in the total number of neurones projecting to the eye between 14 and 31 days postbirth. The presence of significantly greater numbers of cells with collaterals to both eye compartments in the young versus the adult rats has been confirmed using both FB and HRP as retrograde markers. The importance of function on the rearrangement of these projections was investigated by either rearing animals in the dark from the time of birth, or by sectioning the preganglionic nerve trunk to the SCG, soon after birth.(ABSTRACT TRUNCATED AT 250 WORDS)

Adjustment of the myelin sheath to axonal atrophy in the rat spinal root by the formation of infolded myelin loops.

Atrophy of the L4 dorsal and ventral spinal roots was experimentally induced by unilateral sciatic neurectomy in groups of young (2 and 4 months) and older (12 months) albino rats. During the 4 months following neurectomy, the occurrence of infolded myelin loops (IMLs) was quantitatively examined in transverse sections prepared using perfusion fixation with glutaraldehyde and embedding in epoxy resin. The number of IMLs was higher on the operated side and increased with the time of survival and the age of the animals. The formation of IMLs is a characteristic early response of a large-caliber myelin sheath to axonal atrophy, probably reflecting the presence of redundant myelin.

The fiber caliber of 5-HT immunoreactive axons in the dorsolateral funiculus of the spinal cord of the rat and cat.

Although there is considerable evidence that the analgesic action of electrical brain stimulation is mediated in part by serotonergic (5-HT) axons in the dorsolateral funiculus (DLF) of the spinal cord, studies in the rat have questioned the existence of this pathway. In this study, we used antisera directed against a conjugate of 5-HT and bovine serum albumin (BSA) to identify immunoreactive 5-HT axons in the DLF of the rat and cat. Both light and electron-microscopic studies were performed so that the fiber caliber of the labeled axons could also be determined. We found a rich complement of immunoreactive 5-HT axons in the DLF of both rat and cat. Although these could be seen without difficulty in the normal cat, in the rat it was necessary to make a lesion of the DLF to build up the staining rostrally. Ultrastructural analysis established that almost all of the labeled axons (in rat and cat) were unmyelinated. We conclude that there are indeed 5-HT immunoreactive axons in the DLF of the rat and cat. These presumably derive from neurons of the medullary nucleus raphe magnus (NRM), which have been implicated in the descending controls exerted by opiates and electrical brain stimulation. The results suggest that previous physiological studies of the properties of the opiate-responsive, spinally projecting NRM neurons were not made from those that are 5-HT containing.

Lectin and neuropeptide labeling of separate populations of dorsal root ganglion neurons and associated "nociceptor" thin axons in rat testis and cornea whole-mount preparations.

As part of a program to explore patterns of innervation by nociceptor-related thin sensory axons in a variety of peripheral regions, we have labeled calcitonin gene-related peptide immunoreactive (CGRP-IR) nerve fibers in whole mounts of rat testicular tunica vasculosa and cornea. Efforts were undertaken to visualize the numerically significant fluoride-resistant acid phosphatase (FRAP)-containing axon population, whose peripheral endings have heretofore remained undemonstrable due to technical limitations of currently available acid phosphatase methods. Various histochemical markers that colocalize with FRAP in dorsal root ganglion (DRG) and spinal cord were examined, and a plant lectin, Griffonia simplicifolia I-B4, has been identified that not only selectively labels FRAP(+) sensory ganglion cells and central terminals in spinal cord, but also differentially stains a large number of thin axons in testicular and corneal whole mounts. Slender lectin-labeled fibers are abundant in cornea, and are distributed throughout tunica vasculosa preparations unrelated to blood vessels. CGRP-IR axons, in contrast, maintain close adherence to vascular patterns and are more coarse and varicose in appearance. Lectin staining therefore provides the first practical and specific method for visualization of peripheral FRAP(+) axons consisting principally of sensory C fibers but possibly including a small number of unmyelinated autonomic axons. It should now be feasible, using individual whole-mount preparations from various peripheral nociceptor-innervated tissues, to examine the distributions of both peptidergic and FRAP(+) fibers, which together comprise the vast majority of thin sensory axons. It may then be possible to correlate the observed anatomical patterns with knowledge regarding properties of corresponding physiologically characterized receptive fields.

Axon shape as a basis for multinode functional units in a hierarchical neural model.

The ability of animals to perform fixed action patterns and to access information by categories suggests that there are several types of hierarchical organization in the nervous system. This paper employs data about axon shape and neurotransmitter effect to demonstrate the emergence of hierarchical structure in a neural model. Two dimensions of neural classification, axon shape and neurotransmitter effect, are used to generate a five-node-type neural model. Neurons are classified as interneurons, relay cells, and monoamine transmitters on the basis of axon shape; the transmitter classifications include excitatory, inhibitory, and parameter-changing. The five types of nodes in the model correspond to all the biologically observed combinations: excitatory and inhibitory short-range, excitatory and inhibitory long-range-directional, and long-lasting long-range-diffuse nodes. The emergence of multinode functional units (MFUs) from the five-node-type model is mathematically demonstrated. These units correspond to cortical columns anatomically defined by the axon fields of relay cells, and are called columnar multinode functional units (CMFUs). CMFUs may, in turn, be part of larger functional groups designated coherent populations, which consist of widely distributed CMFUs in retinotopically equivalent locations. The existence of coherent populations imposes a three-level hierarchical structure on the model. To represent this hierarchical structure, a new type of CMFU node, which has a set of vector-valued inputs and outputs, is introduced. Each CMFU node contains a system of short-range nodes which supplies it with vector-valued inputs. Sets of long-range-diffuse nodes are also treated as vector-valued nodes whose outputs control the size and number of coherent populations. The role of coherent populations and hierarchical organization in the nervous system is discussed for such cognitive tasks as visual perception, attention and learning. Physiological and behavioral evidence are cited which support the existence of a similar three-level hierarchy in vertebrate brains.

[Neurotransmitters of giant neurons in the African giant snail, Achatina fulica Férussac. IV. Supplemental results]. / Neurotransmetteurs des neurones géants chez l'Escargot géant africain, Achatina fulica Férussac. IV. Résultats complémentaires.

Following the previous works, we identified recently the twelve giant neurones in the ganglia of an African giant snail (Achatina fulica Férussac), by the pharmacological study of their sensitivities to putative neurotransmitters and derivatives, and by the morphological investigation of their axonal pathways due to the intracellular injection of Lucifer Yellow. The neurones studied were: TAN-2, TAN-3, BAPN, LPPN, LBPN and LAPN in the right parietal ganglion; RPeNLN and LPeNLN in the pedal ganglia; and d-LBAN, d-LBMN, d-LBCN and d-LBPN in the left buccal ganglion.

Intradural connections between adjacent cervical spinal roots.

It is not always possible to localize the level of cervical pathology accurately on the basis of clinical signs and symptoms. Intradural intersegmental connections between sensory rootlets occur frequently in the cervical region and have been shown to be clinically and surgically significant. Similar connections between motor rootlets also have been noticed, but their incidence was not reported. Fifty-four human cervical spines were dissected to investigate the incidence of both types of connections. Fifty-three of the 54 specimens had posterior rootlet connections, and nine of the 54 had anterior connections. The preponderant pattern (85%) was for a peripheral dorsal or ventral rootlet to join the central portion of the next rostral or caudal root, and for the two to pass together into the spinal cord. Six distinct patterns were recognized, and a classification system is proposed. These connections may provide a pathway for overlap of sensory dermatomes and motor innervation of the neck and upper extremity. Our observations imply that when a cervical nerve root is injured, small segments of an adjacent root may be equally affected, and the process may be clinically localized one segment higher or lower than it actually is.

An economic anterograde axonal tracing method using Phaseolus vulgaris agglutinin (PHA) P-form.

Phaseolus vulgaris leucoagglutinin (PHA-L) has been demonstrated to be an excellent neuroanatomical anterograde tracer. So far there are relatively few applications of this technique, mainly due to the cost of the lectin. Instead of PHA-L, we have successfully used PHA-P, which is a crude and inexpensive form of PHA-L. The sensitivity of the present method, examined in the rat striatonigral pathway, is as high as that of the conventional method. Furthermore, a large amount of PHA-P, injected into the cat eye, demonstrated the retinofugal projection in detail.

[Synaptogenesis and axon collaterals coming from the white matter in the upper cervical cord of the 10 day (stage 36) chick embryo--Golgi and electron microscopic studies].

Our previous study with 3H-thymidine autoradiography showed that neurons of the zona spongiosa, the nucleus proprius of the dorsal horn, the zona intermedia and the ventralhorn differentiated earlier than those of the substantia gelatinosa and the neck and the base of the dorsal horn, and that neurons of the substantia gelatinosa which were the last to differentiate reached their final position at stage 36 (Fig. 1). In the upper cervical cord of chick embryos at stage 36 when all spinal neurons finished cell migration and the cytoarchitecture similar to that of the cat spinal cord (Rexed, 1952) could be recognized (cf. Figs. 1, 3B), we studied the distribution of synapses by the electron microphotomontage (Fig. 3 A) and the morphology of axon collaterals coming from the white matter by the Golgi method (Fig. 4), in order to examine i) which spinal neurons have synaptic contacts at this stage and ii) what part of the axon collateral makes synaptic contacts. In the white matter, synapses were numerous around the gray matter and they were few in the peripheral part along the external surface of the cord. The paucity of synapses in the peripheral part was explained by a finding that dendrites reaching the external surface of the cord were few in number at this stage (cf. Fig. 3 C). In the gray matter, synapses were more numerous and denser in the zona intermedia and the ventral horn than in the dorsal horn.(ABSTRACT TRUNCATED AT 250 WORDS)

Neural anatomy of the human anterior cruciate ligament.

The histology of the anterior cruciate ligament was studied by a modified technique of the Gairns gold chloride stain for neural elements. Three morphological types of mechanoreceptors and free nerve-endings were identified: two of the slow-adapting Ruffini type and the third, a rapidly adapting Pacinian corpuscle. Rapidly adapting receptors signal motion and slow-adapting receptors subserve speed and acceleration. Free nerve-endings, which are responsible for pain, were also identified within the ligament. These neural elements comprise 1 per cent of the area of the anterior cruciate ligament.

Reticulo-spinal neurons participating in the control of synergic eye and head movements during orienting in the cat. II. Morphological properties as revealed by intra-axonal injections of horseradish peroxidase.

Previously we described physiological properties of pontine reticulo-spinal neurons which generate bursts and decaying tonic discharges related to eye movements and neck muscle activity during ipsiversive gaze shifts (Grantyn and Berthoz 1987). Two of these "eye-neck reticulo-spinal neurons" (EN-RSN) were labeled by intra-axonal injections of HRP. The present report provides a detailed description of their morphology with an emphasis on the topography of axon collaterals, bouton numbers, and the structure of preterminal ramifications in different target areas. The cell bodies of labeled EN-RSNs were located rostro-ventrally to the abducens nucleus. Their descending axons issued 8 and 13 collaterals (left and right EN-RSN, respectively) at different rostro-caudal levels, between the abducens nucleus and the pyramidal decussation. On the basis of the size of their cell bodies, the isodendritic type of dendritic branching and their multiple collateralization, EN-RSNs correspond to the class of "generalized" reticular neurons, often referred to as The Scheibels' neurons. Collaterals of EN-RSNs terminated in the following structures: the abducens and facial nuclei, the medial and lateral vestibular nuclei, the nn. prepositus and intercalatus, and the bulbar reticular formation. As judged from bouton numbers, the strongest connection of both neurons was with the abducens nuclei. Terminations in the rostral part of the medial vestibular and prepositus nuclei indicate that EN-RSNs may also influence oculomotor output activity through these indirect routes. In the facial nucleus, a majority of terminations was found in its medial subdivision containing motoneurons of ear muscles. However, other subdivisions were also contacted by EN-RSNs. Most terminations in the rostral bulbar reticular formation are distributed to the dorsal, gigantocellular field. Within this field, there is a substantial contribution to the zone characterized by the highest density of reticulo-spinal neurons projecting directly to neck motoneurons. Other target areas which may participate in the modulation of spinal cord activity by EN-RSNs are the ventral reticular nucleus in the caudal medulla and the lateral vestibular nucleus. EN-RSNs also establish connections with precerebellar structures: the prepositus and the paramedian reticular nuclei. The numbers of boutons on collaterals issued within 6 mm of the injection site varied between 37 and 469. The occurrence of presumed axo-somatic contacts was low (0-8.2%) and not characteristic for any particular target area. Local accumulations of boutons in the form of small and large field clusters was a common observation.(ABSTRACT TRUNCATED AT 400 WORDS)

Development and axonal outgrowth of identified motoneurons in the zebrafish.

We have observed the development of live, fluorescently labeled motoneurons in the spinal cord of embryonic and larval zebrafish. There are 2 classes of motoneurons: primary and secondary. On each side of each spinal segment there are 3 individually identifiable primary motoneurons, named CaP, MiP, and RoP. The motoneurons of the embryo and larva are similar in morphology and projection pattern to those of the adult. During initial development, axons of primary motoneurons make cell-specific, divergent pathway choices and grow without error to targets appropriate for their adult functions. We observed no period of cell death, and except for one consistently observed case, there was no remodeling of peripheral arbors. We have observed a consistent temporal sequence of axonal outgrowth within each spinal segment. The CaP motor axon is the first to leave the spinal cord, followed by the axons of the other primary motoneurons. The Mauthner growth cone enters the spinal cord after all the primary motoneurons of the trunk spinal cord have begun axonal outgrowth. Secondary motor growth cones appear only after the Mauthner growth cone has passed by. Our results suggest that this stereotyped temporal sequence of axonal outgrowth may play a role in defining the contacts between the Mauthner axon and the motoneurons; the behavior of growth cones in the periphery suggests that interactions with the environment, not timing, may determine path-finding and peripheral connectivity of the motoneurons.

Central connections of Limulus ventral photoreceptors revealed by intracellular staining.

We stained the central terminations of Limulus ventral photoreceptors by intracellular injection of cobalt chloride into the cell bodies. Axons of these photoreceptors enter the protocerebrum via the ventral optic nerve and pass to the medulla. As they reach the surface of the medullar neuropil they branch profusely in fine processes with intermittent varicosities. Each axonal arborization covers about 0.01-0.02 mm2 of this surface immediately adjacent to the medullar ganglion cell layer. Each point on the surface of the medullar neuropil receives, on the average, input from about 6 ventral photoreceptor axons.

The changing view of neural specificity.

The generation of specific patterns of neuronal connections has usually been regarded as a central problem in neurobiology. The prevailing view for many years has been that these connections are established by complementary recognition molecules on the pre- and postsynaptic cells (the chemoaffinity theory). Experimental results obtained in the past decade, however, indicate that the view that axon guidance and synaptogenesis proceed according to restrictive chemical markers is too narrow. Although a more rigid plan may prevail in some invertebrates, the formation of specific connections in vertebrates also involves competition between axon terminals, trophic feedback between pre- and postsynaptic cells, and modification of connections by functional activity.

Primary sensory ganglion cells projecting to the principal trigeminal nucleus in the mallard, Anas platyrhynchos.

The trigeminal and glossopharyngeal ganglia of the adult mallard were studied following HRP injections into the principal trigeminal nucleus (PrV). The PrV consists of the principal trigeminal nucleus proper (prV) and the principal glossopharyngeal nucleus (prIX). After an injection into the prV, the labeled cells were found in the ipsilateral trigeminal ganglion. After an injection into the prIX, labeled cells were found in the ipsilateral distal glossopharyngeal ganglion, but not in the proximal ganglion of the IX and X cranial nerve (pGIX + X). In Nissl preparations, two types of ganglion cells in the trigeminal ganglion, pGIX + X, and distal ganglion of N IX could be distinguished: larger light cells and smaller dark cells. We could not determine whether the HRP-labeled cells belonged to both types or to one of them; but because all the labeled cells were over 20 microns, we concluded that the smallest cells (10-19 microns) in the trigeminal ganglion and distal ganglion of N IX did not project to the PrV. The labeling of the cells in the distal ganglion of N IX (average 34.5 microns) was uniformly moderate. In the trigeminal ganglion there were two types of labeled cells: heavily labeled cells (average 29.1 microns) and moderately labeled cells (average 35.1 l microns). These two types of labeling (moderate and heavy) may reflect two types of primary sensory neurons: cells with ascending, nonbifurcating axons, and cells with bifurcating axons. We speculate that the former are proprioceptive neurons and the latter tactile neurons. Labeled bifurcating axons in the sensory trigeminal complex gave off collaterals to all parts of the descending trigeminal nucleus except to the caudalmost laminated spinal part.

Projections from the cochlear nucleus to the inferior colliculus in normal and neonatally cochlea-ablated gerbils.

The distribution of the projection from one cochlear nucleus (CN) within each inferior colliculus (IC) was studied in adult, normal gerbils and adult gerbils subjected to unilateral ablation of the contralateral cochlea at 2 days of age. The projection was studied by using the Fink-Heimer technique for impregnating degenerating axons and their terminal processes with silver. Following an extensive, unilateral lesion of the CN, degeneration was seen in both ICs of all animals. In normal animals, degeneration was both more widespread and heavier in the contralateral than in the ipsilateral central nucleus of IC (ICC). Degeneration was most widespread in the rostral and lateral parts of both ICCs and in the ventral part of the contralateral ICC. Degeneration was observed in 26% of the area examined in ipsilateral ICC and in 73% of the area examined in contralateral ICC. In cochlea-ablated animals there was a much greater similarity in the area of degeneration in the ICC ipsilateral (57%) and contralateral (67%) to the CN lesion. The same regional distributions of degeneration were observed as in the normal animals except that the distribution of degeneration in the ipsilateral ICC more closely resembled the normal contralateral than the normal ipsilateral profile. We conclude that the normal distribution of projections from the CN within the ipsilateral ICC is substantially modified by neonatal ablation of the contralateral cochlea.