Differences in the organization of actin in the growth cones compared with the neurites of cultured neurons from chick embryos.

Sensory neurons from chick embryos were cultured on substrata that support neurite growth, and were fixed and prepared for both cytochemical localization of actin and electron microscopic observation of actin filaments in whole-mounted specimens. Samples of cells were treated with the detergent Triton X-100 before, during, or after fixation with glutaraldehyde to determine the organization of actin in simpler preparations of extracted cytoskeletons. Antibodies to actin and a fluorescent derivative of phallacidin bound strongly to the leading margins of growth cones, but in neurites the binding of these markers for actin was very weak. This was true in all cases of Triton X-100 treatment, even when cells were extracted for 4 min before fixation. In whole-mounted cytoskeletons there were bundles and networks of 6-7-nm filaments in leading edges of growth cones but very few 6-7-n filaments were present among the microtubules and neurofilaments in the cytoskeletons of neurites. These filaments, which are prominent in growth cones, were identified as actin because they were stabilized against detergent extraction by the presence of phallacidin or the heavy meromyosin and S1 fragments of myosin. In addition, heavy meromyosin and S1 decorated these filaments as expected for binding to F-actin. Microtubules extended into growth cone margins and terminated within the network of actin filaments and bundles. Interactions between microtubule ends and these actin filaments may account for the frequently observed alignment of microtubules with filopodia at the growth cone margins.

Golf: an olfactory neuron specific-G protein involved in odorant signal transduction.

Biochemical and electrophysiological studies suggest that odorants induce responses in olfactory sensory neurons via an adenylate cyclase cascade mediated by a G protein. An olfactory-specific guanosine triphosphate (GTP)-binding protein alpha subunit has now been characterized and evidence is presented suggesting that this G protein, termed Golf, mediates olfaction. Messenger RNA that encodes Golf alpha is expressed in olfactory neuroephithelium but not in six other tissues tested. Moreover, within the olfactory epithelium, Golf alpha appears to be expressed only by the sensory neurons. Specific antisera were used to localize Golf alpha protein to the sensory apparatus of the receptor neurons. Golf alpha shares extensive amino acid identity (88 percent) with the stimulatory G protein, Gs alpha. The expression of Golf alpha in S49 cyc- kin- cells, a line deficient in endogenous stimulatory G proteins, demonstrates its capacity to stimulate adenylate cyclase in a heterologous system.

Quantification of cutaneous sensory nerves and their substance P content in psoriasis.

The aim of the present study was to extend our previous hypothesis that the inflammatory reaction in psoriasis is neurogenic, and that substance P mediates the inflammation. For this purpose, the pattern of neurofilament-positive sensory nerve fibers was studied and the lengths and substance P content of these fibers measured morphometrically in dermal and epidermal compartments of the psoriatic lesion, psoriatic but lesion-free skin, and control skin. The epidermis and dermis of the psoriatic lesions were significantly more densely innervated with neurofilament-positive fibers than either lesion-free psoriatic or control skin. Although substance P is known to be rapidly degraded in tissues, and its actual concentrations in the sections were unknown, there was an increase in substance P containing nerves in the psoriatic lesion, the increase being significant in the epidermal nerve fibers. No significant differences in the measured parameters were obtained between lesion-free psoriatic and control skin. These results indicate that there is an altered pattern of sensory nerves in a psoriatic plaque and that substance P may be an important mediator in the inflammatory processes that contribute either to the initiation or maintenance of a psoriatic lesion.

Immunohistochemical localization of serotonin and choline acetyltransferase in sensory neurones of the locust.

Sensory neuronal cell bodies in the leg of locust, Schistocerca gregaria, were visualized with antibodies to locust choline acetyltransferase and with antibodies to serotonin by the avidin-biotin peroxidase technique. Two groups of sensory cells react with the antibody to choline acetyltransferase: One group is associated with external mechanoreceptors (i.e., hair-plate hairs and campaniform sensilla) and the other with internal proprioceptors (i.e., chordotonal organs and multiterminal receptors). Sensory cells which react with the antibody to serotonin are associated only with internal proprioceptors being found in both chordotonal organs and multiterminal receptors. In the metathoracic femoral chordotonal organ indirect evidence suggests that some sensory cells are reactive to both antibodies. Some multiterminal receptors react with anti-choline-acetyltransferase, while others react with antiserotonin. These results support the conclusion that most insect sensory neurones are cholinergic but some are serotoninergic.

Distribution of substance P-like immunoreactive retinal ganglion cells and their pattern of termination in the optic tectum of chick (Gallus gallus).

Substance P-like immunoreactive (SP-LI) neurons were identified within the inner nuclear layer and ganglion cell layer of the chick retina. The SP-LI cells in the inner nuclear layer consisted of several subtypes of neurons, differing in soma size and dendritic arborization. In the ganglion cell layer a population of moderately labelled SP-LI neurons was also present. About 6-9 microns in diameter and spaced 50-80 microns apart, they formed a regular array across the entire retina, with a density of about 400 cells/mm2 in the superior temporal retina, declining to less than 100 cells/mm2 in the peripheral retina. The total number of SP-LI cells in the ganglion cell layer was approximately 75,000. Individual axons could be followed toward the optic nerve head. Lesions near the optic nerve head resulted in axotomy of ganglion cells within a limited portion of the retina. Two days of postaxotomy there were numerous SP-LI swellings in the proximal segments of axotomized axons. SP-LI neurons in the axotomized zone were larger, more numerous, and showed increased staining of their processes. Fourteen days following a retinal lesion, there was depletion of all SP-LI cells in the ganglion cell layer within the axotomized zone, but the SP-LI neurons in the inner nuclear layer were not noticeably affected. Following a localized injection of rhodamine-coupled latex beads into the optic tectum, a population of retinal ganglion cells (RGCs) in the contralateral retina was retrogradely labelled. Many of these cells also exhibited SP-like immunoreactivity. Examination of the optic tectum indicated the presence of SP-LI fibres in laminae 2-13 (nomenclature of Cajal: Histologie du Systeme Nerveux. Vol. 2. Paris: Maloine, '11), with immunoreactive terminal regions present mainly in laminae 2-4, 7, and 9-13. SP-LI cell bodies were found predominantly in laminae 10-12 and 13. Fourteen days following a retinal lesion, SP-LI processes and terminals were depleted from laminae 2 and 3. Immunoreactive cells and processes in the remaining laminae of the optic tectum were not noticeably altered. The present report confirms the existence of SP-LI retinal ganglion cells in the chick retina and demonstrates their contribution to lamina specific SP-LI arborization in the optic tectum.

The sympathetic and sensory components of the caudal lumbar sympathetic trunk in the cat.

The cell bodies of the lumbar sensory and sympathetic pre- and postganglionic neurons that project in the caudal lumbar sympathetic trunk of the cat have been labeled retrogradely with horseradish peroxidase applied to the central end of their cut axons. The application was made just proximal to the segmental ganglion that sends its gray rami to the L7 spinal nerve, and so identified the sympathetic outflow concerned primarily with the vasculature of the hindlimb and tail. The numbers, segmental distribution, location, and size of the labeled somata have been determined quantitatively. Labeled cell bodies were found ipsilaterally, but the segmental distributions of the different cell types were not matched. Afferent cell bodies lay in dorsal root ganglia L1-L5 (maximum L4), preganglionic cell bodies in spinal segments T10-L5 (maximum L2/3), and postganglionic cell bodies in ganglia L2-L5 (maximum L5). Both numbers and dimensions of labeled dorsal root ganglion cells were variable between experiments (maximum about 1,000); the majority were small relative to the entire population of sensory neurons. Labeled preganglionic cell bodies were located right across the intermediate region of the spinal cord, extending from the lateral part of the dorsolateral funiculus to the central canal. The highest density of labeled neurons lay at the border between the white and gray matter (corresponding to the intermediolateral cell column) with smaller proportions medially in L1-L2, and laterally in caudal L4-L5. Medial preganglionic neurons were generally larger than those lying in lateral positions. From the data, it is estimated that about 650 afferent, about 4,500 preganglionic, and some 2,500 postganglionic neurons project in each lumbar sympathetic trunk distal to the ganglion L5 in the cat.

Anatomical studies on the nucleus reticularis tegmenti pontis in the pigmented rat. I. Cytoarchitecture, topography, and cerebral cortical afferents.

The nucleus reticularis tegmenti pontis (NRTP) is a precerebellar reticular nucleus that has been found to be related to cerebropontocerebellar pathways and, more recently, to eye movements. The present study investigates the cytoarchitecture, the topography, and the cerebral cortical projections to the NRTP in the pigmented rat. The cytoarchitecture and topography of the NRTP was determined by examination of Nissl-stained material sectioned in the transverse and sagittal planes. Two cytoarchitectonically distinct portions of the NRTP are apparent; a central subdivision (NRTPc) composed of large multipolar, small spherical, and fusiform neurons, and a pericentral subdivision (NRTPp) composed of loosely packed small fusiform and spherical neurons. The NRTPc is located dorsal to the medial lemniscus and pyramidal tracts over the caudal two-thirds of the pons. It extends caudodorsally to the region just rostral and ventral to the abducens nucleus. The NRTPp is adjacent to the lateral margins of the NRTPc, rostrally, and lies ventral to the caudal portions of the NRTPc. Large injections of horseradish peroxidase (HRP) were made into the cerebellum in order to determine the degree to which each subdivision of the NRTP contributes to the cerebellar projection. A high percentage of NRTPc neurons and a lower percentage of NRTPp neurons were labeled. These differences in labeling density and neuronal morphology noted above confirm the appropriateness of subdividing the NRTP into central and pericentral subdivisions. The cerebral cortical afferents to the NRTP were examined by placing small iontophoretic injections of HRP into the NRTPc and NRTPp. A systematic examination of all cortical areas revealed that the HRP-labeled neurons are entirely localized within pyramidal layer V of three major cortical areas: the ipsilateral prefrontal cortex (Brodmann areas 8, 8a, 11, and 32); the ipsilateral motor and somatosensory cortices (Brodmann areas 2, 4, 6, and 10), and the bilateral cingular cortex (Brodmann areas 24a, 24b, 29c, and 29d). By far, the heaviest cortical labeling with HRP injections into the medial NRTPc is within the cingular cortex that may, in the rat, be homologous to the frontal eye field of the cat and monkey. In contrast, injections involving the lateral NRTPc or the NRTPp produced labeling within wide regions of the cortex with the greatest number in the somatomotor cortex.(ABSTRACT TRUNCATED AT 400 WORDS)

Anatomical studies on the nucleus reticularis tegmenti pontis in the pigmented rat. II. Subcortical afferents demonstrated by the retrograde transport of horseradish peroxidase.

The subcortical nuclear groups projecting to the nucleus reticularis tegmenti pontis (NRTP) were studied in pigmented rats with the aid of the retrograde horseradish peroxidase (HRP) technique. Small iontophoretic injections of HRP were placed in the medial regions of the NRTP, an area that has been shown in several species to be involved in eye movements. Other large injections in the NRTP or small injections placed just outside the nucleus were used to clarify the projections to the NRTP. Results indicate that the NRTP receives afferents from visual relay nuclei, including the nucleus of optic tract, the superior colliculus, and the ventral lateral geniculate nucleus; oculomotor-associated structures including the zona incerta, the H1 and H2 fields of Forel, the nucleus subparafasciculus, the interstitial nucleus of Cajal, the visual tegmental relay zone of the ventral tegmental area of Tsai, the mesencephalic, pontine, and medullary reticular formations, the nucleus of the posterior commissure, and a portion of the periaqueductal gray termed the supra-oculomotor periaqueductal gray; cerebellar and pontomedullary nuclei, including the superior, lateral, and medial vestibular nuclei, the deep cerebellar nuclei, and NRTP interneurons, and nuclei related to limbic functions including the lateral habenula, the mammillary nuclei, the hypothalamic nuclei, the preoptic nuclei, and the nucleus of diagonal band of Broca. A surprisingly large number of afferents to the medial regions of the NRTP arise from visual- or eye-movement-related nuclei. The projection from the nucleus of the optic tract (NOT) confirms previous anatomical and physiological studies on the pathways involved in horizontal optokinetic nystagmus, but the number of NOT afferents is small in relation to other areas potentially related to visuomotor pathways such as the zona incerta, ventral lateral geniculate nucleus, fields of Forel, perirubral area, and subparafasciculus. The NRTP may also relay information related to vertical visuomotor reflexes (e.g., vertical optokinetic nystagmus) given the strong projections from the medial terminal nucleus of the accessory optic system, visual tegmental relay zone, supra-oculomotor periaqueductal gray, interstitial n. of Cajal, and midbrain reticular formation. The presence of significant NRTP projections from the superior colliculus and the mesencephalic and pontine reticular formations suggests that these nuclei may provide the pathways for the noted saccade-related activity of NRTP neurons. In addition, projections from the vestibular nuclei were found that provide the anatomical basis for head velocity signals recorded in NRTP neurons.(ABSTRACT TRUNCATED AT 400 WORDS)

Physiological, morphological, and cytochemical characteristics of a layer 1 neuron in cat striate cortex.

We have recorded from a small neuron in layer 1 of the striate visual cortex in a 34-day-old kitten. It had a simple, orientation-selective receptive field that was nondirectional and showed length summation. The neuron was injected intracellularly with horseradish peroxidase. Computer-aided reconstruction revealed that it had a dense axonal plexus confined to layer 1, elongated in the anteroposterior dimension. By means of an antibody directed against a GABA-like antigen, and postembedding immunocytohemistry, the neuron was found to be strongly immunoreactive. The main input to soma and dendrites of the neuron was from synapses that were not GABA-L-immunoreactive, and probably originated from pyramidal cells. The axon of the cell formed synapses on dendritic shafts and spines, whose most likely sources were the apical tufts of pyramidal cell dendrites. These data suggest that such neurons may be involved in local circuits that contribute to the formation of pyramidal cell receptive fields.

Brainstem terminations of extraocular muscle primary afferent neurons in the monkey.

The central terminations of afferent nerve fibers from the extraocular muscles of the monkey were investigated by means of transganglionic transport of wheat germ agglutinin-conjugated horseradish peroxidase (WGA/HRP). Following injections of selected extraocular muscles with WGA/HRP, terminal labeling was apparent in the ipsilateral trigeminal sensory and cuneate nuclei. The density of trigeminal projections varied markedly from one rostrocaudal level to the next, being heaviest within the ventrolateral portion of pars interpolaris of the spinal trigeminal nucleus. A second extraocular muscle afferent representation was noted in ventrolateral portions of the cuneate nucleus. This projection was restricted to rostral portions of pars triangularis of the cuneate nucleus, partially overlapping the afferent termination from dorsal neck muscles. It is likely that some of the problems encountered in formulating conclusions regarding the functional role of extraocular muscle proprioception are due to a lack of detailed information of the central termination pattern of muscle afferents. Taken together, the present findings should provide a basis for further anatomical and physiological studies designed to elucidate the role played by extraocular muscle proprioceptors in vision and oculomotor control.

Somatotopic organization of the primary sensory trigeminal neurons in the hagfish, Eptatretus burgeri.

Primary sensory trigeminal projections were investigated in the hagfish following application of horseradish peroxidase (HRP) to the sensory branches. In our control preparations we were able to distinguish five sensory ganglia and their respective nerves. HRP application confirmed the almost exclusive relation of each of these nerves to their respective ganglia, with very little overlap. In normal frontal sections of the medulla oblongata, five columns of fibers surrounded by neuronal cell bodies could be clearly distinguished, but the number is probably fortuitous, for there was no one-on-one relationship with the five trigeminal ganglia. From their peripheral connections, we surmised that columns 1 and 3 handle general cutaneous sensation, columns 2, 4, and 5 handle taste sensation, and column 5 handles general mucous cutaneous sensation conveyed by utricular ganglion cells. Dorsally located columns received projections from nerves with dorsal peripheral connections, and more ventrally located columns received projections from nerves with ventral peripheral connections. This relation is the reverse of that seen in other vertebrates.

Neurofilament phosphorylation in axons and perikarya: immunofluorescence study of the rat spinal cord and dorsal root ganglia with monoclonal antibodies.

Rat dorsal root ganglia and spinal cord were stained with 12 monoclonal antibodies reacting with phosphorylated epitopes of two neurofilament proteins (NF 150K and NF 200K). Three monoclonal antibodies were axon-specific in both locations; neuronal perikarya were not stained. Nine monoclonal antibodies stained a subpopulation of neurofilament-positive sensory neurons, as indicated by double labeling experiments with polyclonal antibodies reacting with phosphorylated and dephosphorylated forms of the neurofilament protein triplet. Of these nine antibodies, two stained motor neuron perikarya in the spinal cord, while the remaining seven antibodies were axon-specific in this location. Subpopulations of stained and unstained motor neurons were not observed. With all 12 antibodies, the staining pattern in the lumbar dorsal root ganglia and spinal cord remained unchanged following sciatic nerve crush and ligature. The findings suggest that, in the neurofilament, some phosphorylated epitopes are axon specific, while other phosphorylated epitopes are present in both axons and perikarya. Furthermore, they suggest that differences exist between neuronal populations as to the presence of phosphorylated epitopes in perikaryal neurofilaments. It remains to be seen whether phosphorylation events in perikarya and axons have similar or different effects on neurofilament structure and function.

The motor nuclei and sensory neurons of the IIIrd, IVth, and VIth cranial nerves in the monitor lizard, Varanus exanthematicus.

The motor nuclei of the oculomotor, trochlear, and abducens nerves of the reptile Varanus exanthematicus and the neurons that subserve the sensory innervation of the extraocular muscles were identified and localized by retrograde and anterograde transport of horseradish peroxidase (HRP). The highly differentiated oculomotor nuclear complex, located dorsomedially in the tegmentum of the midbrain, consists of the accessory oculomotor nucleus and the dorsomedial, dorsolateral, intermediate, and ventral subnuclei. The accessory oculomotor nucleus projects ipsilaterally to the ciliary ganglion. The dorsomedial, dorsolateral, and intermediate subnuclei distribute their axons to the ipsilateral orbit, whereas the ventral subnucleus, which innervates the superior rectus muscle, has a bilateral, though predominantly contralateral projection. The trochlear nucleus, which rostrally overlaps the oculomotor nuclear complex, is for the greater part a comma-shaped cell group situated lateral, dorsal, and medial to the medial longitudinal fasciculus. Following HRP application to the trochlear nerve, almost all retrogradely labeled cells were found in the contralateral nucleus. The nuclear complex of the abducens nerve consists of the principal and accessory abducens nuclei, both of which project ipsilaterally. The principal abducens nucleus is located just beneath the fourth ventricle laterally adjacent to the medial longitudinal fasciculus and innervates the posterior rectus muscle. The accessory abducens nucleus has a ventrolateral position in the brainstem in close approximation to the ophthalmic fibers of the descending trigeminal tract. It innervates the retractor bulbi and bursalis muscles. The fibers arising in the accessory abducens muscles form a loop in or just beneath the principal abducens nucleus before they join the abducens nerve root. The afferent fibers conveying sensory information from the extraocular muscles course in the oculomotor nerve and have their perikarya in the ipsilateral trigeminal ganglion, almost exclusively in its ophthalmic portion.

Neurotransmitter specificity of cells and fibers in the medial preoptic nucleus: an immunohistochemical study in the rat.

The medial preoptic nucleus (MPN) is a sexually dimorphic complex with three major subdivisions. The cell-dense central (MPNc) and medial (MPNm) subdivisions are larger in male rats, while the cell-sparse lateral subdivision (MPNl) occupies a majority of the nucleus in females. In the present study we evaluated the distribution of possible monoaminergic and peptidergic cells and fibers within the MPN, as well as in adjacent regions of the medial preoptic area of the adult male rat. For this, we used an indirect immunohistochemical method with antisera to serotonin (5HT), dopamine beta-hydroxylase (DBH), tyrosine hydroxylase (TH), neuropeptide Y (NPY), cholecystokinin (CCK), vasoactive intestinal polypeptide (VIP), substance P (SP), neurotensin (NT), corticotropin-releasing factor (CRF), luteotropin-releasing hormone (LRH), somatostatin (SS), thyrotropin-releasing hormone (TRH), oxytocin (OXY), vasopressin (VAS), adrenocorticotropic hormone (1-24; ACTH), alpha-melanocyte-stimulating hormone (alpha-MSH), leucine-enkephalin (L-ENK), and calcitonin gene-related peptide (CGRP). The results suggest that cell bodies and/or fibers crossreacting with all of these putative neurotransmitters are differentially distributed within the MPN. Within the MPNm, the densest plexuses of fibers were stained with antisera to SP and NPY, while moderate densities of fibers were stained with anti-DBH, SS, CCK, CGRP, ACTH, and alpha-MSH, and only a few fibers were stained with anti-5HT, TH, NT, VAS, and L-ENK. Moderate numbers of SP- and L-ENK-immunoreactive cell bodies, and a few SS-, NT-, CRF-, and TRH-stained cell bodies were also found within the MPNm. The MPNc contained a dense plexus of CCK-immunoreactive fibers, as well as a few CRF-immunoreactive fibers. Both fiber types were localized almost exclusively to this subdivision, while most of the others studied here appeared to avoid it selectively. This suggests that there are relatively few inputs to the MPNc, and that they tend to avoid other parts of the nucleus, although moderate densities of DBH- and NPY-immunoreactive fibers were found in both the MPNm and MPNc. The MPNc contained several CCK-immunoreactive cell bodies as well as a moderate number of TRH-stained cell bodies. Both cell types were nearly completely localized to the MPNc. The major inputs to the MPNl studied here appear to be stained with antisera to 5HT and L-ENK, although moderate numbers of NT- and CRF- immunoreactive fibers were also found in this part of the nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)

Olfactory marker protein is present in olfactory receptor cells of human fetuses.

Light microscopic observations indicated that cellular differentiation of the primary olfactory pathway in human fetuses was relatively advanced by the end of the first trimester. However, immunohistochemical staining showed that olfactory marker protein was present in the receptor cells only at about 28 weeks post-conception. At about 32 weeks, modest amounts of the protein could be found in the peripheral olfactory nerve layer and a few glomeruli, with increased staining observed in subsequent weeks. The results show conclusively that human fetal olfactory neurons synthesize olfactory marker protein during the later half of gestation.

Acetylcholinesterase and luteinizing hormone-releasing hormone distinguish separate populations of terminal nerve neurons.

The terminal nerve is composed of a morphologically heterogeneous population of unipolar, bipolar and multipolar neurons located in the nasal and intracranial cavities of vertebrates. The question has arisen as to whether these neurons are neurochemically heterogeneous and therefore possibly functionally different as well. Among the substances localized in the terminal nerve are acetylcholinesterase and luteinizing hormone-releasing hormone-like immunoreactive material. We have developed a double-label procedure, combining immunocytochemistry and enzyme histochemistry to determine whether these two substances are localized within different populations of terminal nerve neurons. Compatibility of the two procedures was accomplished by modifications of the fixative and primary antibody solutions. In the immunocytochemical step, the avidin-biotin-peroxidase complex coupled to a new chromogen, Chromo-red, produced a bright red reaction product in neurons containing luteinizing hormone-releasing hormone-like material. This reaction product was easily differentiated from the black silver-intensified acetylcholinesterase label. In both neonatal and adult preparations, a large population of terminal neurons contained the acetylcholinesterase label only, whereas a smaller population contained both acetylcholinesterase and luteinizing hormone-releasing hormone-like material. The acetylcholinesterase-containing population of neurons was concentrated peripherally and included multipolar neurons. In contrast neurons with the two substances co-localized were unipolar or bipolar and were concentrated centrally. The simultaneous visualization of acetylcholinesterase and luteinizing hormone-releasing hormone-like material in the same tissue section enable the differentiation of two separate neurochemically defined populations of terminal neurons. The distribution of these two neuronal types was the same in neonatal and adult animals. These data provide support for a functional diversity of terminal neurons.

neuropeptides in long ascending spinal tract cells in the rat: evidence for parallel processing of ascending information.

A study has been made of the involvement of spinal peptidergic neurons in ascending tracts at lumbar-sacral levels in rats, by combining the retrograde transport of a protein-gold complex with immunocytochemistry. Ten neuropeptides have been considered for their presence in the cells of origin of the following six ascending tracts, including some involved in pain transmission: the spinosolitary tract, the medial and lateral spinoreticular tracts, the spinomesencephalic tract, the spinothalamic tract and the postsynaptic dorsal column tract. Although there was overlap in the distribution of several of the types of peptidergic cells and some ascending tract cells only a very small percentage of long ascending tract cells were found to contain neuropeptides. Most (90%) of those peptidergic ascending tract cells, however, were clearly congregated in two distinct spinal regions: the lateral spinal nucleus and the region surrounding the central canal (including lamina X). Ascending tract cells in both of these regions contained a wide variety of neuropeptides. Immunoreactivities for a total of seven different peptides were seen. The lateral spinal nucleus had the highest percentage of neuropeptide containing ascending tract cells; cells of all the four populations of peptidergic neurons lying in this region were involved in supraspinal projections; they stained for vasoactive intestinal polypeptide, bombesin, substance P or dynorphin and their axons projected in the spinomesencephalic, spinoreticular and spinosolitary tracts. The region surrounding the central canal contained bombesin-, enkephalin-, cholecystokinin- and somatostatin-immunoreactive ascending tract cells; these cells were found at the origin of the spinothalamic, spinomesencephalic, spinoreticular and spinosolitary tracts. In this region only the cells staining for substance P were not involved in supraspinal projections. The peptidergic ascending tract cells in other spinal regions were few; they were found in either lamina I or lateral part of lamina V. Ascending tract lamina I cells reacted for dynorphin or vasoactive intestinal polypeptide and their axons projected in the spinosolitary and spinomesencephalic tracts. Ascending tract lamina V cells reacted for somatostatin and were found at the origin of the medial component of the spinoreticular tract. It is proposed that peptidergic ascending tract cells form minor but distinct subgroups within each ascending tract. Each of the ascending tracts are divisible into peptide- and nonpeptide-containing groups of cells which convey information in a parallel fashion.(ABSTRACT TRUNCATED AT 400 WORDS)

The effect of unilateral dorsal root ganglionectomies or ventral rhizotomies on alpha 2-adrenoceptor binding to, and the substance P, enkephalin, and neurotensin content of, the cat lumbar spinal cord.

The density of alpha 2-adrenoceptor binding sites and the content of substance P, enkephalins, and neurotensin were determined in quadrants of the lumbar spinal enlargement of control cats and of cats upon which either unilateral dorsal root ganglionectomies or unilateral ventral rhizotomies had been performed. The performance of unilateral dorsal root ganglionectomies resulted in a significant decrease (45-55%) of substance P content in the ipsilateral dorsal horn 7 and 21 days postoperatively. The concentration of alpha 2-adrenoceptor binding sites ([3H]rauwolscine Bmax) in the ipsilateral dorsal horn was consistently and significantly decreased at these same postganglionectomy times (20% reduced relative to the contralateral dorsal horn). Enkephalin content 7 and 21 days after ganglionectomies was not significantly different from control, whereas the neurotensin content of the ipsilateral dorsal horn was significantly increased in the 21-day survival cats. The performance of unilateral ventral rhizotomies did not produce any statistically significant changes in the density of alpha 2-adrenoceptor binding sites or in the substance P or enkephalin content of any spinal quadrant. The neurotensin content of both the ipsilateral dorsal and ipsilateral ventral quadrants of the ventral rhizotomized cats was significantly increased. The significant decrease of alpha 2-adrenoceptor binding site concentration in the ipsilateral dorsal horn after unilateral dorsal root ganglionectomies suggests that approximately 20% of the alpha 2-adrenoceptors present within the cat lumbar spinal dorsal gray are located on the axons or terminals of primary sensory afferents. Consistent with this interpretation of the ganglionectomy results, we found significant levels of saturable [3H]rauwolscine binding to homogenates of the cat L4-Sl spinal dorsal root ganglia. Because alpha 2-adrenoceptor binding sites in the ipsilateral ventral lumbar spinal gray were not significantly reduced after unilateral ventral rhizotomies, our results provide no evidence for the location of alpha 2-adrenoceptor on lumbar spinal motoneurons.

Evidence for a visceral afferent origin of substance P-like immunoreactivity in lamina V of the rat thoracic spinal cord.

A visceral afferent origin of substance P-like immunoreactivity in lamina V of the lower thoracic spinal cord of the rat was investigated. In transverse sections from normal animals there was a moderately dense substance P-immunoreactive innervation of lamina V. In some sections there was a dorsoventrally orientated fibre bundle from the superficial dorsal horn entering lamina V. In parasagittal sections, substance P-immunoreactivity in lamina V was found arranged in clusters, with a periodicity in the rostrocaudal axis of 200-600 microns. In some cases these were seen to be continuous with a dorsoventrally orientated fibre bundle from the superficial dorsal horn. After section of the splanchnic nerve there was a consistent reduction in the density of the substance P-like immunoreactivity in lamina V, with fewer clusters on the operated side. Adult rats treated neonatally with capsaicin showed a substantial reduction of substance P-immunoreactivity in laminae I and II and the virtual abolition of staining in lamina V. These results provide evidence of a visceral origin for some of the substance P-like immunoreactivity in lamina V of the rat thoracic spinal cord. In addition, they confirm that most of the substance P-immunoreactivity in the dorsal horn is of primary afferent origin.

Immunohistochemical study of neuropeptides in vagal and glossopharyngeal afferent neurons in the rat.

The presence and distribution of multiple neuropeptides in vagal and glossopharyngeal afferent ganglia of the rat were studied using immunohistochemistry. Substance P-, calcitonin-gene related peptide-, cholecystokinin-, neurokinin A-, vasoactive intestinal polypeptide-, and somatostatin-immunoreactive neurons were detected in each visceral afferent ganglion. Neurotensin-immunoreactive cells were not observed. In the nodose ganglion (inferior ganglion of the vagus nerve) occasional immunoreactive cells were scattered throughout the main (caudal) portion of the ganglion with small clusters of cells seen in the rostral portion. The pattern of distribution of the various peptides in the nodose ganglion was similar, with the exception of vasoactive intestinal polypeptide-immunoreactive neurons which exhibited a more caudal distribution. The relative numbers of immunoreactive cells varied, with the greatest numbers being immunoreactive for substance P or vasoactive intestinal polypeptide, and the lowest numbers being immunoreactive for neurokinin A and somatostatin. A build-up of immunoreactivity for each of the peptides, except somatostatin and neurotensin, was detected in vagal nerve fibers of colchicine-injected ganglia. Numerous peptide-immunoreactive cells were also found in the petrosal (inferior ganglion of the glossopharyngeal nerve) and jugular (superior ganglion of the vagus nerve) ganglia. No specific intraganglionic distribution was noted although the relative numbers of cells which were immunoreactive for the different peptides varied considerably. Substance P and calcitonin-gene related peptide were found in large numbers of cells, cholecystokinin was seen in moderate numbers of cells, and neurokinin A, vasoactive intestinal polypeptide and somatostatin were seen in fewer cells. These data provide evidence for the presence and non-uniform distribution of multiple peptide neurotransmitters in vagal and glossopharyngeal afferent neurons. In general, relatively greater numbers of immunoreactive cells were located in the rostral compared with caudal nodose ganglion, and in the petrosal and jugular ganglia compared with the nodose ganglion. Thus, multiple neuropeptides may be involved as afferent neurotransmitters in the reflexes mediated by vagal and glossopharyngeal sensory nerves.