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Nerve terminals in the human periodontal ligament were investigated by immunohistochemistry for neurofilament protein (NFP) and glia-specific S-100 protein. The human periodontal ligament was found to be innervated by NFP-immunoreactive nerve fibers, and contained free and specialized nerve endings; essentially all these nerve fibers were associated with S-100 protein-immunoreactive elements. Free nerve endings showing tree-like ramifications were frequently observed along the whole length of the tooth. The specialized nerve endings consisting of thick nervous elements were classified into four types as follows: Ruffini-like endings which were composed of expanded nerve terminals arranged in a dendritic fashion were found mainly around the root apex; coiled nerve endings were located in the mid-region of the periodontal ligament; spindle-shaped types, and expanded nerve endings, both rarely found near the root apex. Immunostaining for S-100 protein showed that no lamellated nerve terminals were found in the human periodontal ligament. The region-specific distribution of various nerve terminals demonstrated in this study seems to be suited to effectively receive mechanical stimuli to the teeth from various directions.

By means of immunochemical method the presence of vasopressin in the melanocytes of the man's brain arteries was found. The terminals of the cholinergic axons come to an end on the surface of these cells. It is suggested that the melanocytes take part in the regulation of the brain arteries mobility.

The electron microscopic investigation was performed to analyze somatostatin-contained nerve terminals in the median eminence of 21 days old malnourished rats' hypothalamus. In nerve terminals of malnourished animals in compared with controls ones there was found the increased density of granular vesicles (11.62 +/- 0.40 and 8.56 +/- 0.39 in 1 micron2, respectively) and decreased density of electron lucent vesicles with 120-160 nm diameter (1.66 +/- 0.18 and 3.43 +/- 0.26 in 1 micro2, respectively). The revealed increase in density of granular vesicles in axon terminals with positive immunohistochemical reaction to somatostatin in malnourished rats was explained by slow somatostatin release.

In this study we utilized the technique of simultaneous immunofluorescent double-labeling to investigate possible coexistence of the putative neurotransmitter thyrotropin-releasing hormone (TRH) with serotonin (5-HT) and with substance P (SP) in the intermediolateral cell column (IML) of rat spinal cord. We observed fibers and terminals immunoreactive for both TRH and 5-HT or TRH and SP in IML. In addition, this technique was used in animals in which we retrogradely labeled, with fluorescent tracer dyes, preganglionic sympathetic neurons within IML from either the adrenal medulla or the proximal cut end of the cervical sympathetic trunk. In these animals, fibers and terminals containing these combinations of neurotransmitters appeared to oppose identified preganglionic sympathetic neurons in IML. These data represent the first direct immunohistochemical demonstration of fibers and terminals in spinal cord which display coexistence of TRH- with either 5-HT- or SP-immunoreactivity. In addition, the proximity of TRH-immunoreactive fibers and terminals to sympathetic preganglionic neurons in IML support a role for TRH in the regulation of central sympathetic outflow.

Intraventricular injections of 6-hydroxydopamine (6-OHDA) caused a significant decrease in the tyrosine hydroxylase of striatal synaptosomes of rats, without influencing GABA levels. Significant decreases in the muscarinic and nicotinic receptors also occurred in this preparation after 3 days. The receptors were measured by specific binding studies with N-methylatropine and alpha-bungarotoxin respectively. These results indicate that muscarinic and nicotinic receptors are located on the dopaminergic nerve terminals of corpus striatum.

A newly developed synaptosomal model was used to evaluate the in vivo effects of the GABA-elevating drugs aminooxyacetic acid (AOAA, 30 mg/kg i.p.) and valproic acid (VPA, 200 mg/kg i.p.) on GABA levels in nerve endings of 11 brain regions in rats as a function of time after administration. The data obtained were compared with the magnitude and time course of the effects of both drugs in rats on body temperature, pain response and against seizures induced by electroshock, pentylenetetrazol and 3-mercaptopropionic acid. Following AOAA, maximum increases in synaptosomal GABA levels of brain regions were observed 6 hr after administration. At this time, GABA was significantly elevated up to 300% over control values in synaptosomal fractions from all 11 regions. However, the hypothermic and antinociceptive effects of the drug as well as its anticonvulsant action against electroshock and pentylenetetrazol induced seizures were maximal 1 hr after injection and had vanished after 6 hr, i.e. at the time of maximum GABA increases in synaptosomes. The only pharmacological effect of AOAA which paralleled the time course of the synaptosomal GABA elevation was the attenuation of seizures induced by 3-mercaptopropionic acid. Following VPA, the effect on synaptosomal GABA levels was much more rapid in onset and significant increases were already determined 5 to 30 min after administration. Significant increases of up to 80% over control values were found in synaptosomal fractions from olfactory bulb, frontal cortex, hippocampus, hypothalamus, tectum, substantia nigra and cerebellum. In contrast to AOAA, the time course of the synaptosomal GABA increases, at least in some regions, was similar to the time course of VPA's antinociceptice effects and its anticonvulsant effects in the three seizure models studied. The data may suggest that AOAA and VPA increase different pools of GABA within nerve terminals, only one of which is involved in GABA-mediated neurotransmission.

A monoclonal antibody against choline acetyltransferase (ChAT), the acetylcholine-synthesizing enzyme, was used to study cholinergic synapses on identified (Golgi stained) granule cells in the rat fascia dentata. Choline acetyltransferase immunocytochemistry was applied to 40-microns Vibratome sections cut perpendicular to the longitudinal axis of the hippocampus. Light microscopy revealed fine varicose ChAT-immunoreactive axons in all layers of the fascia dentata, i.e., in the stratum moleculare, the stratum granulosum, and the subgranular polymorph zone. Most fibers were observed in the vicinity of granule cell bodies where they ran mainly parallel to the granular layer. Next, the immunostained Vibratome sections were sandwiched between small pieces of Parafilm and piled to form a block that was covered with agar and Golgi stained. After that, the sections were separated by cutting away the agar and removing the Parafilm. Sections containing well-impregnated granule cells were gold-toned (Fairén et al., '77), embedded in Araldite, and subjected to ultrathin sectioning for electron microscopy. A total of 14 gold-toned granule cells were examined in the electron microscope for synaptic contacts with cholinergic afferents. Choline acetyltransferase-immunoreactive axon terminals were observed that established symmetric synaptic contacts with the cell bodies and dendritic shafts of the gold-toned identified granule cells. Two types of contact were observed on spines arising from gold-toned granule cell dendrites. Immunoreactive terminals established asymmetric synaptic contacts with the head of small spines and symmetric contacts with the stalk of large, complex spines. The boutons forming asymmetric synaptic contacts with the cup-shaped spine head of the complex spines were not found to be immunoreactive. Our results demonstrate that cholinergic fibers to the rat fascia dentata establish characteristic types of synaptic contact with different postsynaptic elements of granule cells, suggesting a complex function of this afferent system.

Neuropeptide Y-containing nerve cell bodies and processes were identified by electron microscopic immunocytochemistry in the guinea pig small intestine. Labeled nerve processes were numerous in the myenteric plexus. However, a few immunoreactive nerve fibers were found in all layers of the small intestine. Some of the immunoreactive nerve processes were found in close apposition to the epithelial cells of the crypts of Lieberkühn and to endothelial and smooth muscle cells. The neuropeptide Y-containing nerve cell bodies were preferentially located in the submucous ganglia. In the myenteric plexus many synaptic connections were observed between the neuropeptide Y-immunoreactive nerve fibers and unlabeled nerve cell bodies and other nerve fibers. These findings provide a morphologic basis for the possibility that neuropeptide Y may act as a transmitter and exert postsynaptic effects on intrinsic neurons, in addition to participating in the regulation of smooth muscle activity and epithelial cell functions.

The auditory and vestibular sense-organs of the rabbit were examined for the presence of substance P(SP) by an immunohistochemical technique using a monoclonal SP-antibody. Between 30 and 40 per cent of the cells of the vestibular ganglia and about 50 per cent of the spiral ganglion cells innervating the extreme basal part of the cochlea showed SP-like immunoreactivity. The neural elements in the vestibular sensory epithelia, notably the calyx-formed nerve terminals of type I sensory cells of the maculae, also showed strong SP-like immunoreactivity. The findings suggest the possibility of a modulator or transmitter role for SP in the inner ear function.

A combined morphological and physiological analysis of the properties of the adrenergic vasoconstrictor innervation was carried out in the splanchnic vasculature in the rat. Three generations of vessels were studied: (1) the abdominal aorta and the caval vein; (2) the superior mesenteric artery and vein; (3) 200 microns branches of the superior mesenteric artery and their parallelling veins. The adrenergic innervation was visualized by the Hillarp-Falck fluorescence technique, and by the immunohistochemical localization of immunoreactivity to tyrosine hydroxylase and dopamine-beta-hydroxylase. Determination of responses to applied noradrenaline, to transmural nerve stimulation and to direct activation of the muscle was made on ring preparations mounted in a myograph. All vessels were found to be innervated at the adventitio-medial border by noradrenergic nerves with varying density; the small arteries had the highest nerve density, the abdominal aorta was least innervated. When related to the maximal isometric response to applied noradrenaline, the maximal neurogenic response parallelled the density of the adrenergic innervation. Variations in frequency for half-maximal response among the vessels could not, however, be ascribed to innervation properties only. The constriction under isotonic conditions amounted to 20% of the initial circumference in the aorta, and to 30% in the small arteries and veins. The relation between response to applied noradrenaline and to nerve stimulation was similar for isometric and isotonic responses. In the smaller veins, a beta-receptor-mediated decline in the response to applied noradrenaline was seen. This decline was much less pronounced in neurogenic responses. The results indicate a gradation from proximal to peripheral arteries towards denser innervation and greater neurogenic responses. On the venous side only minor differences were found.

The location of the perikarya of LH-RH neurons in the rat hypothalamus and their pathways to the median eminence were studied by immunohistochemistry and radioimmunoassay after placing stereotaxic electrolytic lesions in several parts of the hypothalamus. The principal location of the cell somata was found to be in the ventral part of the medial preoptic area; their pathways were classified into a main baso-lateral pathway and an accessory descending pathway branching off from the former. The main pathway was found to cross in the vicinity of the corresponding neuronal perikarya. The central median eminence and the dorsal and ventral walls of the tubero-infundibular sulcus of the caudal part of the median eminence are innervated mainly by the baso-lateral pathway. On the other hand, the rostral and most caudal protions of the median eminence are innervated principally by the descending pathway and have a subsidiary dual innervation. The projection of LH-RH neurons to the OVLT is believed to originate from perikarya adjacent to this circumventricular organ.

The ultrastructure, afferent input, and sites of termination of neurons containing neuropeptide Y-like immunoreactivity (NPY-LI) were examined in the adult rat nucleus accumbens by using the peroxidase-antiperoxidase (PAP) method. The NPY-LI was seen in sparsely distributed, spindle-shaped perikarya having cross-sectional diameters of 15-20 microns. These perikarya exhibited highly invaginated nuclear membranes and thin rims of cytoplasm containing Golgi lamellae, dense-core vesicles, and other organelles. A few large, principally aspiny, dendrites also showed NPY-LI. The dendrites received synaptic input from unlabeled terminals forming both symmetric and asymmetric junctions. Immunolabeling for NPY was evident in other processes that were not clearly differentiated as dendrites or axons. These were seen primarily near glial processes and the basal laminae of blood vessels. A few myelinated and many unmyelinated axons and axon terminals also were labeled for NPY. These terminals contained numerous, small (40-60 nm), clear and one or more large (80-100 nm) dense core vesicles. Forty-seven percent (27 out of 57) of the terminals containing NPY-LI formed symmetric junctions with unlabeled dendrites or dendritic spines. The remainder lacked recognizable densities within single planes of section. The neurons exhibiting NPY-LI in the nucleus accumbens were characterized further with respect to their afferent input from terminals labeled for the GABA-synthesizing enzyme, glutamic acid decarboxylase (GAD). Immunogold labeling of a rabbit antiserum against NPY and PAP labeling for a sheep antiserum to GAD were sequentially applied to the same sections. The GAD-labeled terminals formed symmetric junctions primarily with the more numerous unlabeled dendrites. However, a few synaptic junctions also were detected between the GAD-labeled terminals and dendrites showing immunogold labeling for NPY. We conclude (1) that in the rat nucleus accumbens, NPY-LI is found principally in neurons of the aspiny type and (2) that the output from these presumably intrinsic neurons to other neighboring neurons or blood vessels is at least partially modulated by GABA.

The distribution and properties of receptors for corticotropin-releasing factor (CRF) were analyzed in the brain of cynomolgus monkeys. Binding of [125I]tyrosine-labeled ovine CRF to frontal cortex and amygdala membrane-rich fractions was saturable, specific, and time- and temperature-dependent, reaching equilibrium in 30 min at 23 degrees C. Scatchard analysis of the binding data indicated one class of high-affinity sites with a Kd of 1 nM and a concentration of 125 fmol/mg (approximately equal to 30% of the receptor number in monkey anterior pituitary membranes). As in the rat pituitary and brain, CRF receptors in monkey cerebral cortex and amygdala were coupled to adenylate cyclase. Autoradiographic analysis of specific CRF binding in brain sections revealed that the receptors were widely distributed in the cerebral cortex and limbic system. Receptor density was highest in the pars tuberalis of the pituitary and throughout the cerebral cortex, specifically in the prefrontal, frontal, orbital, cingulate, insular, and temporal areas, and in the cerebellar cortex. A very high binding density was also present in the hippocampus, mainly in the dentate gyrus, and in the arcuate nucleus and nucleus tuberis lateralis. A high binding density was present in the amygdaloid complex and mamillary bodies, olfactory tubercle, and medial portion of the dorsomedial nucleus of the thalamus. A moderate binding density was found in the nucleus accumbens, claustrum, caudate-putamen, paraventricular and posterior lateral nuclei of the thalamus, inferior colliculus, and dorsal parabrachial nucleus. A low binding density was present in the superior colliculus, locus coeruleus, substantia gelatinosa, preoptic area, septal area, and bed nucleus of the stria terminalis. These data demonstrate that receptors for CRF are present within the primate brain at areas related to the central control of visceral function and behavior, suggesting that brain CRF may serve as a neurotransmitter in the coordination of endocrine and neural mechanisms involved in the response to stress.

By the use of well-characterized antibodies against porcine dynorphin-A(1-8), an endogenous opioid peptide, and the use of a modified immunofluorescence microscopic technique, dynorphin-A(1-8) stained perikarya, nerve fibres, and nerve terminals were visualized in the rat duodenum. Dynorphin-A(1-8) immunoreactive perikarya were revealed with certainty only in the myenteric plexus, while dynorphinergic nerve fibres could bee seen in the myenteric plexus and circular muscle layer, but not in the longitudinal muscle layer and submucous plexus. Dynorphin-A(1-8) immunofluorescent nerve endings were in close contacts with submucosal blood vessels, probably arterioles, and Brunner's gland cells. These findings suggest that the opioid peptide dynorphin-A(1-8) might be synthetized within myenteric plexus perikarya of the rat duodenum and that it might modulate the peristaltic activity, intestinal blood pressure, and production of mucopeptides synthetized within Brunner's gland cells.

The present paper is concerned with the arrangement of axons and synaptic terminals immunostained for calcitonin gene-related peptide (CGRP), a primary afferent marker, in the primate (Macaca fascicularis) dorsal horn. The CGRP axons and terminals are uniformly distributed in laminae I and II outer (o) but they are concentrated laterally and distributed intermittently in the reticulated region of lamina V. A prominent bundle of labeled axons is seen in the sacral cord dorsal to the central canal. Emphasis is given to the relation of CGRP-immunoreactive terminals to other terminals, both labeled and unlabeled, in laminae I and IIo. In this regard, adjacent CGRP-immunoreactive terminals are often united by puncta adhaerentia. Of particular interest is the observation that CGRP-immunoreactive terminals can be found presynaptic to other terminals which sometimes resemble central primary afferent endings. In addition CGRP-immunoreactive terminals end on other CGRP terminals. Both findings suggest that primary afferent terminals interact synaptically with other primary afferent terminals.

The fast axonal transport of proteins was studied in the cat sciatic nerve after injection of [3H]leucine into the spinal ganglion or the ventral horn of the seventh lumbar segment. The amount of transported proteins after ganglion injection was linearly related to the amount of label present at the ganglion. At variable intervals after ganglion or spinal cord injection, the sciatic nerves were sectioned in some experiments. The transport of proteins continued in the peripheral nerve stump in a wavelike manner, but the advancing wave leaves a labeled trail behind. A fraction of this trail corresponds to proteins moving at slower velocities than the velocity of proteins in the wave front. Another fraction of the trail corresponds to molecules retained by the axons. Each nerve segment of 5 mm in length retains 1.5% of the transported proteins, and the profile of retained proteins along the sciatic nerves follows a single exponential function. From the proportion of retained proteins, the concentration of transported proteins at the terminals of branching axons as a function of the branching ratio was estimated. In the case of motor axons innervating the soleus muscle of the cat, the concentration of recently transported proteins at the nerve terminals would be approximately 0.83% of the proteins leaving the spinal cord. This low concentration of transported proteins at the nerve terminals may explain the lability of neuromuscular synapses when axonal transport is decreased or interrupted.

The coexistence of met5- and leu5-enkephalin-like immunoreactivities with catecholamines in the rat adrenal medulla was studied with combined fluorescence microscopy and immunocytochemistry. Both met5- and leu5-enkephalin-like immunoreactivities were localized in few heavily stained adrenaline cells and in a population of nerves innervating adrenaline cells and as well as ganglion cells among the adrenaline cells. Only occasionally single noradrenaline cells exhibited light immunostaining for both enkephalins but no positive fibers could be found around the noradrenaline cells. In electron microscope the immunoreaction was seen in the granules of the adrenaline cells and in the large synaptic vesicles of the nerve terminals around the adrenaline cells. The present findings suggest that enkephalin-like immunoreactivity coexists mainly with adrenaline in rat adrenal medulla and that the enkephalin immunoreactive terminals regulate secretion of adrenaline from rat adrenal medulla.

To determine whether vascular postsynaptic alpha 2-adrenoceptors are innervated by sympathetic nerves in man, normal volunteers were infused for 90 min with angiotensin II (A II) and the alpha 2-adrenoceptor agonist alpha-methylnoradrenaline (MNA) in a double-blind cross-over study. Whilst systolic blood pressure returned to baseline within 5 min of terminating the A II infusion it remained elevated 40 min after stopping MNA. The prolongation of the pressor response to MNA, a substrate for neuronal uptake, was probably due to activation of alpha 2-adrenoceptors by MNA re-released from contiguous sympathetic nerve endings. The proximity of alpha 2-adrenoceptors to sympathetic nerve terminals suggests that they could contribute to blood pressure regulation in man.

Anterograde labelling following focal injections of Phaseolus vulgaris leucoagglutinin was used to identify the threedimensional cortical distribution of thalamocortical (TC) fibers from the ventrolateral nucleus of the thalamus of the cat. The labelled TC fibers were distributed usually in layers I and III of the motor cortex and the terminals in layer III tended to aggregate into patches about 1-1.5 mm wide in a mediolateral direction. These patches were arranged in longitudinal strips about 2-5 mm long in a rostrocaudal direction and were separated by gaps of terminal free area.

Five patients with idiopathic orthostatic hypotension and defective vasoconstrictor responses to the Valsalva maneuver and to application of ice to the forehead were found to have absent vasoconstriction in the forearm in response to intra-arterial administration of tyramine and enhanced vasoconstrictor responses in response to intra-arterial administration of norepinephrine. These findings strongly suggested norepinephrine depletion from the nerve endings and inability of the sympathetic nerve endings to take up norepinephrine. The depletion of norepinephrine from sympathetic nerve endings was confirmed in four patients by demonstrating histochemically the absence of catecholamine-specific fluorescence in sympathetic vasomotor nerves from deltoid muscle. It is suggested that depletion of norepinephrine from nerve endings was responsible for autonomic dysfunction and orthostatic hypotension in these patients.

In the mesencephalon of the chicken, the two large groups of CA terminals and fibers appeared to correspond to the dorsal and ventral bundles of the CA containing fibers in the rat. These ran rostrally in the medial forebrain bundle and appeared to terminate within the thalamus, hypothalamus, preoptic region and corpus striatum. 5-HT containing terminals and fibers also were observed in the mesencephalon and appeared to run rostrally in the medial part of the medial forebrain bundle. In comparison with the CA terminals, however, 5-HT terminals were for the greater part, difficult to observe in many areas of the brain. Based on the observation concerning postnatal ontogeny of monoamine-containing neurons, the chicken brain appears to acquire behavioral and functional maturity at an earlier stage after birth than does that of the rat. Effect of reserpine, nialnmide, L-Dopa and 5-HTP on the monoamine-containing terminals and fibers are briefly discussed.

To determine differential tissue antigens in the same section immunocytochemically using the electron microscope, the neurohypophysis was examined following the application of a freeze-drying tissue preparation and staining with the protein A-colloidal gold-antibody complex method (Hisano S, Adachi T, Daikoku S: J Histochem Cytochem 32:705, 1984). At the light microscopic level, colocalized immunostaining for methionine-enkephalin (ENK) and oxytocin (OXT) was found in the rat neurohypophysis under different physiological states. Small pieces of the neurohypophysial tissue were frozen and dried. The dried tissue was fixed with paraformaldehyde vapor and embedded. The ultrathin sections were stained with the antibody for ENK coupled with protein A-small colloidal gold, and antibody for OXT or vasopressin (VP) conjugated with protein A-large colloidal gold. The ultrastructures of the nerve terminals were well preserved and showed many membrane-limited secretory granules. It was possible to identify both OXT- and VP-containing nerve terminals as their secretory granules were differentially labeled with protein A-colloidal gold anti-OXT or anti-VP complex, respectively. The secretory granules, which were labeled with large gold particles for OXT, also carry small gold particles. It is evident that ENK coexists with OXT in the same granules.

The possible existence of tryptamine-containing neurons originating in the midbrain raphe is suggested by several reports of tryptamine-mediated responses to electrical stimulation of the raphe nuclei. To assess this hypothesis, we have investigated the effects of electrolytic lesions of the median and dorsal raphe nuclei on striatal, hypothalamic, and hippocampal concentrations of tryptamine, 5-hydroxytryptamine (5-HT), and 5-hydroxyindoleacetic acid. In addition, the rat striatal tryptophan concentrations were also determined. No changes in the concentrations of tryptamine were observed at 1 or 2 weeks after lesioning the dorsal and median raphe nuclei, at which time the other 5-hydroxyindoles were markedly reduced; furthermore, no reductions were observed in tryptamine concentrations in the striatum, hypothalamus, or hippocampus of rats pretreated with a monoamine oxidase inhibitor. The only change observed in these rats was a limited increase in striatal tryptamine and tryptophan observed at 1 day after lesioning. The results indicate that tryptamine concentration is independent of the integrity of 5-HT-containing neurons of the midbrain raphe nuclei. Furthermore, if tryptamine-containing neurons that have terminal projections to the striatum, hypothalamus, and hippocampus exist, their cell bodies are located in regions outside the dorsal and median raphe nuclei. Another possibility could be that tryptamine is located in glial cells.

Substance P (SP) immunoreactive nerve endings in the laryngeal mucosa were studied by PAP immunohistochemistry with light and electron microscopy. SP immunoreactive sensory endings were observed in the epithelium as intra-epithelial free nerve endings and taste bud-like structures. A small number of autonomic SP immunoreactive nerve fibers were observed running parallel to arterioles which were over 30 micron in diameter and terminated in glandular cells. Contrary to findings by silver impregnation, intraepithelial free nerve endings were more frequently observed on the lower surface of the vocal cord. The taste bud-like structures were classified into two different types: simple terminations and reticular terminations, according to the mode of the SP immunoreactive nerve fiber. Immature or degenerated taste bud-like structures in the larynx were assumed to be mechanical receptors because these receptors lacked outer taste pores and taste hairs.

Rabbits were immunized with nerve ending fraction prepared from guinea pig brain. Serum antibodies to total gangliosides were followed by enzyme-linked immunosorbent assay; their titers were highest at 2 to 3 weeks after immunization and some rabbits showed a response to reinjections. Specific reactivities of the antibodies against each molecular species of gangliosides were analyzed by liposome lysis assay and liposome lysis inhibition assay. Antibody responses were detected against GM1, GD1b, GM3 and GM2, but not against GD1a and GT1b. Natural and immune antibodies to the asialo glycolipids and to galactosylceramide were also observed.

The aim of the presented paper was to compare the Falck-Hillarp histofluoresce method for monoamines with its cryostat modification and the glyoxylic acid SPG method. The sensitivity and specificity of these methods and also the occurence of unspecific fluorescence is discussed.

The central arborizations and endings of type I spiral ganglion neurons were labeled with intracellular injections of horseradish peroxidase (HRP) after their characteristic frequency (CF) and spontaneous discharge rate (SR) were physiologically determined. A fiber-by-fiber analysis was conducted and the morphological data compared with the fiber's response properties. The total number of branch points was correlated with total fiber length, a relationship that remained relatively constant when analyzing the ascending and descending branches together or separately. On the other hand, the ascending branches of four out of five fibers having CFs below 0.5 kHz bifurcated and gave rise to a pair of terminal endbulbs of Held. Low- and medium-SR fibers gave rise to more endings than did high-SR fibers, especially on the ascending branch. This difference was accounted for by small endings, a category composed of terminal boutons, string endings, and small complex endings. The categories of modified endbulbs, and endbulbs of Held did not vary in number with respect to fiber SR. The mean area of each ending type within the small ending category was statistically smaller for low- and medium-SR fibers than for high-SR fibers, whereas the mean area of modified endbulbs and endbulbs of Held was not correlated with fiber SR. Total ending area per fiber appeared independent of either CF or SR. These results are discussed in relation to issues of conservation of axon arborizations and terminals, and convergence of input from the different SR groups.

The distribution of gamma aminobutyric acid (GABA)-containing neurons and nerve terminals was examined in the rat septal area by using specific antibodies to the enzyme glutamic acid decarboxylase (GAD) in combination with the avidin-biotin immunoperoxidase method. Whereas only a few GAD positive neurons were present in the septum of normal rats, the septal area of rats treated with colchicine, an inhibitor of fast axonal transport, showed numerous GAD-immunoreactive neurons. These neurons were evenly filled with GAD-immunoreactive material throughout the cytoplasm of the soma and proximal parts of the dendrites. Although GAD-positive neurons were present in most parts of the septal area, their density differed greatly in the different septal subnuclei. Both the diagonal band of Broca (vertical and horizontal parts) and the lateral septum were rich in GAD positive cell bodies, whereas the medial septal nucleus and the intermediate parts of the lateral septum contained relatively few. Within the lateral septum itself a larger number of labeled cell bodies was present in its ventral subdivision. The anterior hippocampal rudiment (taenia tecta) contained numerous GAD-positive neurons, while the septal component of the island of Calleja (insula magna) was devoid of them. GAD-immuno-positive neurons found within the septum ranged from small (15 microns) to large (30-35 micron). They were round or multipolar in the diagonal band, medium-sized multipolar in the lateral septum, and pyramidal, round or fusiform in the anterior hippocampal rudiment. GAD-immunoreactive nerve terminals are present in most subdivisions of the septal nuclei, with the exception of myelinated fiber tracts, and throughout all rostrocaudal levels of the septum. However, the density of the innervation is not the same within all individual nuclei. The lateral septum (dorsal and ventral parts) contained high density innervation but the diagonal band of Broca had a lower density of GAD-positive terminals. The lateral border of the islands of Calleja was rich in thick GAD-positive processes that appeared to be continuous with GAD-immunoreactive processes of the substantia inominata. The inner portion of the molecular layer adjacent to the granule cells of the anterior hippocampal rudiment contained a rich GAD-positive terminal field.

Glutamate has been shown to be a neurotransmitter in the central nervous system of vertebrates, and it has been hypothesized that glutamate is functional as a neurotransmitter in the spinal cord dorsal horn. A monoclonal antibody to fixative-modified glutamate was used in this study to examine the light microscopic and ultrastructural profiles of glutamate-like immunoreactivity in the superficial dorsal horn of the rat spinal cord. Glutamate-like immunoreactivity was observed in neurons, fibers, and terminals of both laminae I and II. Marginal zone immunoreactive neurons ranged from 10 to 30 micron in diameter and received many nonimmunoreactive somatic synapses. In substantia gelatinosa, immunoreactive neurons were observed in both inner and outer layers, ranged 5 to 10 micron in diameter, and received few nonimmunoreactive somatic synapses. Glutamate-like immunoreactive dendrites were observed in both laminae and were contacted primarily by nonimmunoreactive synaptic terminals that generally contained small clear vesicles. Both myelinated and unmyelinated immunoreactive axons were observed in Lissauer's tract. Immunoreactive terminals contained small (40 nm) clear vesicles and generally formed simple synaptic contacts with nonimmunoreactive dendrites in laminae I and II. The results of this study corroborate the importance of glutamate as a neurotransmitter in spinal sensory mechanisms.

The spinal trigeminal subnucleus caudalis processes nociceptive input from the head. However, physiological and behavioral studies in monkeys and humans indicate that painful stimuli from the central face and oral cavity also project through trigeminal nuclei rostral to the spinal subnucleus caudalis. Both enkephalin (ENK) and serotonin (5-HT) are present in rostral trigeminal nuclei and these regions receive inputs from the raphe complex. Thus, it appears that elements of pain-modulating circuitry proposed by Basbaum and Fields (Annu. Rev. Neurosci., 7:309-338, 1984) for the spinal and medullary dorsal horn may also exist in this region. In order to begin an exploration of this circuitry, the present study combines the techniques of retrograde transport of HRP from the ventral posteromedial thalamic nucleus (VPM) of the cat's thalamus to label trigeminothalamic relay cells. Secondarily, immunocytochemical techniques are employed to define the distribution patterns of ENK and 5-HT cells and terminals in relationship to both labeled and nonlabeled neurons in each of the subnuclei of the spinal trigeminal nucleus. Trigeminothalamic relay cells were observed in laminae I and II, the magnocellular region, and the interstitial nucleus (IN) of subnucleus caudalis (Vc). ENK was found in axodendritic and axosomatic terminals, together with a population of small fusiform neurons in all these same areas except the magnocellular region. ENK axosomatic contacts innervated approximately 30% of labeled relay cells, chiefly in lamina I and the IN, or small unlabeled neurons in the same area. Serotonin activity occurred principally in lamina I and the IN and was confined almost exclusively to axodendritic terminals. Examination of subnucleus interpolaris (Vi) revealed relay cells distributed throughout the length of the nucleus and increasing in numbers at rostral levels. A rostral extension of the IN was found just ventrolateral to the main body of Vi and contained numerous labeled cells. The distribution of ENK activity was restricted to the ventral part of Vi and the IN and occurred in axodendritic and axosomatic terminals. These latter elements innervated 30-40% of labeled relay cells in Vi, particularly those located in the IN. Cells containing ENK generally resembled the fusiform cells found in Vc and were distributed in ventral Vi and the IN. Some ENK cells were larger, displayed several dendrites, and occurred only in the ventral Vi. Serotonin within Vi and Vc was confined principally to axodendritic terminals.(ABSTRACT TRUNCATED AT 400 WORDS)

The synaptic relationships between substance P-containing terminals and dopaminergic neurons (immunoreactive for tyrosine hydroxylase) in the substantia nigra were studied at both light and electron microscopic levels using a pre-embedding double-labeling immunocytochemical method. Many substance P-containing terminals were found to form synapses directly with dendrites and somata of nigral dopaminergic neurons. Since most of the substance P-containing axon terminals arise from the striatum, this result suggests that striatal substance P neurons can have monosynaptic influence on nigral dopaminergic neurons.

gamma-Aminobutyric acid (GABA)-like immunoreactivity in the chick vestibular endorgans was examined using an antiserum against GABA coupled with glutaraldehyde to bovine serum albumin. GABA-like immunoreactivity was confined to the cytoplasm of the hair cells in both cristae and maculae. GABA-like immunoreactive cells were evenly distributed throughout the sensory epithelia, and no difference existed between type I and type II hair cells. The results provide evidence that GABA-like immunoreactivity is localized to sensory cells and raises the possibility that GABA may serve as an afferent neurotransmitter in the chick vestibular end organs.

There is some controversy concerning a possible effect of diabetes mellitus on the sympathetic nervous system in humans with spontaneous diabetes mellitus and in animals with experimental diabetes mellitus. In this study we compared the tissue norepinephrine (NE) concentration of normal and diabetic Chinese hamsters in the untreated state and after treatment with insulin. Diabetes resulted in a 128% increase in the NE concentration of the kidney in female but not male hamsters. The NE concentration was increased in the liver (133%) and in the cerebral cortex (118%) of both male and female hamsters. There was no significant increase in the NE concentration of hypothalamus, acinar pancreas, pancreatic islets, or heart of diabetic hamsters. Three days of insulin therapy reduced the elevated NE concentration in kidney, liver and cerebral cortex of diabetic hamsters to the levels found in normal hamsters. However, insulin therapy of normal hamsters did not reduce the tissue NE concentration of the kidney, liver, and cerebral cortex below the normal levels found in these animals. Insulin therapy reduced the hypothalamic concentration of NE in both diabetic and normal hamsters. The increase in kidney NE concentration in female diabetic hamsters was not due to a reduction in renal size, for the kidneys of both female and male diabetic hamsters were larger than those of normal hamsters. When synthesis of NE was inhibited with alpha-methyltyrosine, there was a comparable rate of fall in the tissue NE concentration in the four experimental groups, suggesting that the increased tissue NE concentration in the tissues of diabetic hamsters was not due to a decreased rate of disappearance of this compound.(ABSTRACT TRUNCATED AT 250 WORDS)

In the present study, the origin of calcitonin gene-related peptide (CGRP) to the dorsal horn in the rat lumbar spinal cord is investigated. CGRP immunoreactivity is examined following multiple unilateral and bilateral dorsal rhizotomies and isolated cord preparations (spinal cords are isolated by transecting the cord in two places and cutting all dorsal roots between the transections). Seven to 11 days after surgery, unilateral multiple dorsal rhizotomies result in a drastic decrease in CGRP-stained terminals on the operated side; following bilateral dorsal rhizotomies and isolated cord preparations, one or two CGRP varicosities remain in the dorsal horn in each section. The numbers of CGRP-immunostained varicosities observed in the latter two preparations are not significantly different, suggesting that few if any axons descending from the brain contribute to the CGRP terminal population in the spinal cord dorsal horn. Based on these data, we hypothesize that dorsal root ganglion cells are the only source of CGRP to the rat lumbar dorsal horn.

Luteinizing hormone-releasing hormone immunoreactivity was studied in the olfactory system of the rat in combination with acetylcholinesterase histochemistry. Neuronal perikarya containing luteinizing hormone-releasing hormone lie in the medial septal nucleus, the vertical limb of the diagonal band of Broca, the olfactory tubercule and the ganglionated plexus of the terminal nerve. Labelled fibres spread in the superficial layers of the main and accessory olfactory bulbs, some encompassing the strongly acetylcholinesterase-positive atypical glomeruli. Others are observed on the medial side of the bulb, running along the terminal nerve bundles and ganglia. These fibres join the vomeronasal nerve branches and proceed distally towards the nasal cavity. In the septal submucosa, immunoreactive fibres are partly associated with the terminal nerve network. Conspicuous endings filled with luteinizing hormone-releasing hormone are observed on blood vessels of the olfactory mucosa. Such well-differentiated terminals might be the neurosecretory afferents of a new neurohemal area. Immunoreactive terminals are also observed around the excretory ducts of the anterior medial glands. We have failed to observe any labelled fibres in the olfactory and vomeronasal epithelia. The results of the present study are discussed with respect to possible functional interpretations. It is suggested that significant amounts of luteinizing hormone-releasing hormone could be released in the submucosal capillaries in spite of the scarcity of immunoreactive fibres. Similar afferents could also modulate the secretory activity of some nasal glands. Synaptic events involving the neuropeptide might occur in the olfactory bulb, particularly in atypical glomerular areas previously characterized by their high acetylcholinesterase content. Finally, no anatomical support for a chemosensory function of fibres containing luteinizing hormone-releasing hormone has been brought out by our work.

Immunoreactive leucine-encephalin has been found in nerve terminals and cell bodies of neurons in the human brain. The most intense reaction was found in the globus pallidus, followed by the spinal nucleus of the trigeminal nerve. These observations suggest that in man endogenous opioid substances may play an important part in neurotransmission in areas of the central nervous system related to pain and locomotor activity.