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PubMed1981

Functional organization of the oculomotor nucleus in the baboon.

The functional organization of the oculomotor nucleus was investigated using horseradish peroxidase (HRP) histochemistry. In a series of baboons, injections of HRP were made into the skeletal muscles supplied by the oculomotor nerve (medial rectus, superior rectus, inferior rectus, inferior oblique, and the levator palpebrae superioris). After a 48-hour survival time the animals were sacrificed via perfusion-fixation and the brains treated according to the tetramethylbenzidine (TMB)-HRP method of Mesulam (1978). The inferior oblique, inferior rectus, medial rectus, and levator palpebrae superioris muscle are supplied by cells located primarily in the homolateral oculomotor nucleus. Some fibers to the levator palpebrae superioris arise from cells in both caudal central nuclei. The superior rectus muscle receives fibers from cells in the contralateral oculomotor nucleus. The results were compiled using a Lucite-plate reconstruction method that permits visualization of the three-dimensional configuration of the neuronal populations within the oculomotor nucleus. Oculomotor neurons are organized in a vertical column that may be anatomically divided into rostral, middle, and caudal thirds. A section through any of these levels may be further subdivided into dorsal, intermediate, and ventral zones. Each of the oculomotor skeletal muscles was found to have cells at almost all levels of the nucleus and in certain zones at each level. These functional cell groups intermingle with one another in the baboon and do not remain segregated into distinct subnuclei or subdivisions. Much overlap was evident between cells innervating the homolateral inferior rectus, homolateral inferior oblique, and the contralateral superior rectus muscles. There was also overlap between those cells supplying the homolateral levator palpebrae superioris and the homolateral medial rectus muscles.

Animals↗
PubMed1986

The accessory nerve nucleus in the baboon.

In the savanna baboon, Papio cynocephalus, the accessory nerve nucleus was identified by using a mixture of 20% free horseradish peroxidase (HRP) and 2.5% HRP conjugated to wheat germ agglutinin (WGA) in a 5% aqueous detergent solution (Nonidet P-40). Following surgical exposure of the appropriate nerve branch to the sternocleidomastoid or trapezius muscle, the nerve was transected, placed in an Argyle tubing collar, and bathed in 5-10 microliter of the tracer. After a 48-hour survival time and vascular perfusion-fixation, 40-micron sections of the lower medulla oblongata and the cervical spinal cord were treated according to the tetramethyl benzidine (TMB)-HRP method of Mesulam (J. Histochem. Cytochem. 26: 106-117, 1978). The accessory nucleus extends as a distinct column of neurons from lower medullary levels into the rostral part of C5. One to ten labeled cells were present in each section, and all labeled neurons were located on the side of the bathed nerve. The rostral portion of the accessory nucleus occupies a central position, its intermediate portion occupies a lateral position, and its caudal portion occupies a central position within the ventral horn. All labeled neurons were confined to Rexed's lamina IX, ranged from 15 to 75 micron in diameter, and were either distinctly round (oval) or stellate in shape. Neurons within the baboon accessory nucleus supplying the sternocleidomastoid muscle were located from lower medullary to upper C2 spinal cord levels, while those supplying the trapezius muscle extended from C2 to C5.

Accessory Nerve↗
PubMed1997

Quantitative determination of oLAb titers in various animal species.

It is generally accepted, that lipid peroxidation plays a pathogenic role in atherosclerosis. Furthermore, recent studies indicate that antibodies directed against oxidative modifications of Low Density Lipoprotein (oLAb) contribute to atherosclerotic processes and may have some function in other disorders. These antibodies have been determined predominantly in humans, because assays for oLAb measurement use species specific anti IgG conjugates. From such assay designs it is not possible to get directly comparable data from various animal species. Main advantages of comparable data between animal species are that results of animal experiments can be interpreted using human calibrators and that results of immunisations and production of monoclonal antibodies are directly comparable not only within, but also between animal species. The aim of this study was to find a modification for ELISAs for oLAb determination, which allows to measure sera of various animal species simultaneously. Microtitration plates were coated with oxidised LDL and blocked with bovine serum albumine. Human and animal sera were then pipetted into the plate in logarithmic serial dilutions and incubated for 2 h at 37 degrees C. After washing, a protein A horse-radish peroxidase conjugate (Biomakor, Israel) was added to each well in a dilution of 1:20,000. The incubation conditions had to be optimized to achieve reliable results. After another washing step, the assay was developed with TMB. Absorptions were read at 450 nm in a microplate photometer. Following the manufacturers incubation instructions, which recommended a duration of 1 h at room temperature, the system did not work optimally. No binding of protein A to IgG molecules bound to oxidised LDL could be observed, if the system was incubated at 37 degrees C. In our hands, best results were achieved for several animal species, if the conjugate was incubated for two hours at 2-4 degrees C in a refrigerator. Under these conditions, assay sensitivity was the same as in the standard method, which uses anti-species IgG conjugates. The protein A modification of oLAb allows direct reading of animal oLAb titres from human calibrators. With this method, results of animal experiments can be interpreted on the basis of the situation in humans. Preliminary results obtained show that immunisation experiments with oxidised LDL give serum titres in animals, which are in the same order of magnitude as human sera with high oLAb concentrations. The results of this study, in accordance with findings of other authors, give further indications that atherosclerotic processes are influenced by the specific immune system.

Animals↗
PubMed2002

Furfural, 5-hydroxymethyl furfural, and acetoin act as external electron acceptors during anaerobic fermentation of xylose in recombinant Saccharomyces cerevisiae.

The electron acceptors acetoin, acetaldehyde, furfural, and 5-hydroxymethylfurfural (HMF) were added to anaerobic batch fermentation of xylose by recombinant, xylose utilising Saccharomyces cerevisiae TMB 3001. The intracellular fluxes during xylose fermentation before and after acetoin addition were calculated with metabolic flux analysis. Acetoin halted xylitol excretion and decreased the flux through the oxidative pentose phosphate pathway. The yield of ethanol increased from 0.62 mol ethanol/mol xylose to 1.35 mol ethanol/mol xylose, and the cell more than doubled its specific ATP production after acetoin addition compared to fermentation of xylose only. This did, however, not result in biomass growth. The xylitol excretion was also decreased by furfural and acetaldehyde but was unchanged by HMF. Thus, furfural present in lignocellulosic hydrolysate can be beneficial for ethanolic fermentation of xylose. Enzymatic analyses showed that the reduction of acetoin and furfural required NADH, whereas the reduction of HMF required NADPH. The enzymatic activity responsible for furfural reduction was considerably higher than for HMF reduction and also in situ furfural conversion was higher than HMF conversion.

Acetoin↗
PubMed1980

Brain stem projections of sensory and motor components of the vagus complex in the cat: I. The cervical vagus and nodose ganglion.

The motor and sensory connections of the cervical vagus nerve and of its inferior ganglion (nodose ganglion) have been traced in the medulla oblongata of 32 adult cats with the neuroanatomical methods of horseradish peroxidase (HRP) histochemistry and amino acid autoradiography (ARG). In 14 of these subjects, an aqueous solution of HRP was applied unilaterally to the central end of the severed cervical vagus nerve. In 13 other cases, HRP was injected directly into the nodose ganglion. Three of these 13 subjects had undergone infranodose vagotomy 6 weeks prior to the HRP injection. A mixture of tritiated amino acid was injected into the nodose ganglion in five additional cats. The retrograde transport of HRP yielded reaction product in nerve fibers and perikarya of parasympathetic and somatic motoneurons in the medulla oblongata. Furthermore, a tetramethyl benzidine (TMB) method for visualizing HRP enabled the demonstration of anterograde and transganglionic transport, so that central sensory connections of the nodose ganglion and of the vagus nerve could also be traced. The central distribution of silver grain following injections of tritiated amino acids in the nodose ganglion corresponded closely with the distribution of sensory projections demonstrated with HRP, thus confirming the validity of HRP histochemistry as a method for tracing these projections. The histochemical and autoradiographic experiments showed that the vagus nerve enters the medulla from its lateral aspect in multiple fascicles and that it contains three major components--axons of preganglionic parasympathetic neurones, axons of skeletal motoneurons, and central processes of the sensory neurons in the nodose ganglion. Retrogradely labeled neurons were seen in the dorsal motor nucleus of X(dmnX), the nucleus ambiguus (nA), the nucleus retroambigualis (nRA), the nucleus dorsomedialis (ndm) and the spinal nucleus of the accessory nerve (nspA). The axons arising from motoneurons in the nA did not traverse the medulla directly laterally; rather, all of these axons were initially directed dorsomedially toward the dmnX, where they formed a hairpin loop and then accompanied the axons of dmnX neurons to their points of exit. Afferent fibers in the vagus nerve reached most of the subnuclei of the nTS bilaterally, with the more intense labeling being found on the ipsilateral side. Labeling of sensory vagal projections was also found in the area postrema of both sides and around neurons of the dmnX. These direct sensory projections terminating within the dmnX may provide an anatomical substrate for vagally mediated monosynpatic reflexes. Following deefferentiation by infranodose vagotomy 6 weeks prior to HRP injections into the nodose ganglion, a number of neurons in the dmnX were still intensely labeled with the HRP reaction product. The axons of these HRP-labeled perikarya may constitute the bulbar component of the accessory nerve.

Afferent Pathways↗
PubMed1982

Brainstem projections of sensory and motor components of the vagus nerve in the rat.

The sensory and motor connections of the cervical vagus nerves and of its inferior ganglion (nodose ganglion) have been traced in the medulla and upper cervical spinal cord of 16 male Wistar rats by using horseradish peroxidase (HRP) neurohistochemistry. The use of tetramethyl benzidine (TMB) as the substrate for HRP permitted the visualization of transganglionic and retrograde transport in sensory nerve terminals and perikarya, respectively. The vagus nerve in the rat enters the medulla in numerous fascicles with points of entry covering the entire lateral aspect of the medulla extending from level +4 to -6 mm rostrocaudal to the obex. Fascicles of vagal sensory fibers enter the dorsolateral aspect of the medulla and travel to the tractus solitarius (TS) which was labeled for over 8.8 mm in the medulla. The caudal extent of the TS receiving vagal projections was found in lamina V of the cervical spinal cord (C1 to C2). Sensory terminal fields could be visualized bilaterally in the nucleus of the tractus solitarius (nTS), area postrema (ap) and dorsal motor nucleus of the vagus nerve (dmnX). The ipsilateral projection to the nTS and the dmnX was heavier than that found on the contralateral side. The area postrema was intensely labeled on both sides. Motor fibers from HRP-labeled perikarya in the dmnX travel ventromedially in a distinct fascicle and subsequently subdivide into a number of small fiber bundles that traverse the medullary reticular formation in the form of a fine network of HRP-labeled fibers. As these fibers from the dmnX approach the ventrolateral aspect of the medulla they are joined by axons from the nucleus ambiguus (nA), nucleus retroambigualis (nRA) and the retrofacial nucleus (nRF). These latter fibers form hairpin loops in the middle of the reticular formation to accompany the axons from the dmnX exiting from the medulla in a ventrolateral location. HRP-labeled perikarya, in contrast to transganglionically transported HRP in sensory terminals in the nTS, were visualized on one side only, thus indicating that motor control via the vagus nerve is exerted only by motor neurons located ipsilaterally. Sensory information on the other hand, diverges to many nuclear subgroups located on both sides of the medulla.

Accessory Nerve↗
PubMed1983

The development and postnatal organization of primary afferent projections to the rat thoracic spinal cord.

Primary afferent projections to the thoracic spinal cord in fetal and postnatal rats were labelled by applying horseradish peroxidase (HRP) to the central stumps of cut peripheral nerves. Diaminobenzidine (DAB) and tetramethyl benzidine (TMB) histochemical processing procedures were used to reveal the HRP reaction product. In postnatal rats, individual muscle nerves were labelled to reveal the organization of muscle afferent projections to the motor nuclei. The terminals of muscle afferents were distributed widely across the dendritic arbors of motoneurons supplying the same muscles. No spatial segregation of the terminations of different populations of muscle afferents was discernable. Afferents supplying different regions of the skin were labelled by applying HRP to the dorsal and ventral primary rami of the spinal nerves. Afferents in the dorsal rami projected to lateral portions of both the ipsilateral and contralateral dorsal horns while afferents in the ventral rami projected to the medial portions of both dorsal horns. The projections of the dorsal rami were shifted caudally relative to those of the ventral rami. This relationship reflects the fact that the regions of skin innervated by the dorsal rami are displaced caudally relative to those innervated by the corresponding ventral rami. In fetuses, dorsal rami were labelled alone or in combination with ventral rami. These experiments disclosed the time course of development of the projections to different laminae of the spinal gray matter and revealed that afferents in the two primary rami project to appropriate regions in the ipsilateral and contralateral dorsal horns from the very outset.

Afferent Pathways↗
PubMed1984

Projections from the periaqueductal gray matter to the B3 cellular area (nucleus raphe magnus and nucleus reticularis paragigantocellularis) as revealed by the retrograde transport of horseradish peroxidase in the rat.

Afferent projections from the periaqueductal gray matter (PAG) to the B3 region (nucleus raphe magnus, NRM; and nucleus reticularis paragigantocellularis, NPG) were examined by means of the horseradish peroxidase (HRP) method revealed by using the tetramethylbenzidine (TMB) procedure. Following iontophoretic injection of HRP into the B3 cellular area, numerous labeled neurons were found between the third oculomotor complex and the rostral part of the tegmental nucleus dorsalis of Gudden. The most densely labeled regions were the lateral parts of the dorsal raphe nucleus (wings of the DRN) and Hamilton's nucleus dorsalis of the PAG. Very few neurons were found within the medial part of the DRN. In contrast, results with control injections performed outside the B3 area gave a different distribution of labeled neurons. The functional significance of these connections, particularly those emanating from the wings of the DRN which represent new information, is briefly discussed with regard to mechanisms operative in the control of pain. Although the major PAG-B3 direct projections could underlie a role for the B3 area in PAG-induced analgesia, the comparison of our anatomical and behavioral results points out some problems. Indeed, there is no strict correlation between the location of stimulation sites which induce analgesia in the freely moving rats and the distribution within the PAG of B3 projections. These observations lead us to question the role of direct PAG-B3 connections in the antinociception induced by PAG stimulation and alternative hypotheses are proposed.

Analgesia↗
PubMed1984

Efferent connections of the caudate nucleus, including cortical projections of the striatum and other basal ganglia: an autoradiographic and horseradish peroxidase investigation in the cat.

Tritiated tracer was injected into the head of the caudate nucleus in cats. Following such injections, labeling is present within extensive regions of both the globus pallidus and entopeduncular nucleus, where it presents a mottled or meshlike appearance. These projections are topographically organized in that there is simple correspondence between the mediolateral, dorsoventral, and rostrocaudal origin of the caudate projection and its input to the globus pallidus and entopeduncular nucleus. Transported tracer is also present within the substantia nigra, where it is most abundant within the pars reticularis. However, distinct labeling also overlies cells of the pars compacta, and lesser amounts of labeling are present within the pars lateralis and within the retrorubral area. Following injections of horseradish peroxidase into the caudate nucleus, and subsequent tissue processing by the tetramethylbenzidine (TMB) method of Mesulam ('78), labeled anterograde fibers are present in abundance within the globus pallidus, entopeduncular nucleus, and all subdivisions of the substantia nigra, thus confirming the autoradiographic findings. Also, it is especially obvious in this HRP material that, contrary to previous degeneration studies, both the rostromedial and caudolateral parts of the pars lateralis of the substantia nigra contain numerous anterogradely labeled fibers. Retrogradely labeled neurons are also present within the substantia nigra of these same tissue sections, where they are most abundant within the pars compacta, but lesser numbers of labeled neurons are also present within the pars reticularis, pars lateralis, retrorubral area, and ventral tegmental area on the ipsilateral side, and all of these same subdivisions of the substantia nigra on the contralateral side. Also, within the subthalamic nucleus in these experiments, there are anterogradely labeled fibers, as well as retrogradely labeled neurons, which are interpreted to represent a reciprocal connection between the subthalamic nucleus and the striatum. In a separate series of experiments, horseradish peroxidase was injected into the motor cortex-specifically into the anterior sigmoidal gyrus. Following such injections, labeled neurons representing afferents to the motor cortex are found in all subcortical nuclei commonly known as the "basal ganglia," including the caudate nucleus, putamen, globus pallidus, entopeduncular nucleus, substantia innominata, nucleus of the diagonal band of Broca, medial septal nucleus, claustrum, and basolateral amygdaloid nucleus.

Amygdala↗
PubMed1984

Cortical projections to the paramedian tegmental and basilar pons in the monkey.

The efferent connections of the cerebral cortex to paramedial tegmental and basilar pons were studied in the monkey by using the retrograde and orthograde capabilities of the horseradish peroxidase (HRP) technique. Six capuchin monkeys (Cebus apella) received transcannular pontine HRP gel implants to retrogradely label the cells of origin of corticopontine projections. Four additional capuchin monkeys, one rhesus (Macaca mulatta), and one cynomolgus (Macaca fascicularis) monkey, received HRP gel implants in premotor (area 6), frontal eye field (FEF, area 8), superior (area 5), and inferior (area 7) parietal lobules to orthogradely label the course and termination of corticopontine projections, and thus to confirm the retrograde studies. The brains were processed according to the tetramethylbenzidine (TMB) protocol of Mesulam ('78) and studied with darkfield microscopy. Premotor (area 6) frontal cortex and FEF (area 8) were found to be the main sources of cortical inputs to the ipsilateral paramedian basilar pons, whereas FEF, dorsal prefrontal convexity, and dorsal medial prefrontal (granular frontal association) cortex were the main sources of bilateral projections to the paramedian pontine tegmentum. The medial portion of the nucleus reticularis tegmenti pontis (NRTP), considered to be a tegmental extension of the basilar pontine gray, also received its principal cortical input from the frontal lobe. Parietal cortex, on the other hand, was observed to project to lateral NRTP and lateral basilar pons. Although the possibility exists of convergence of frontal and parietal eye field efferents in the NRTP, the frontal eye field and prefrontal cortex appear to be the principal source of cortical projections to the paramedian pontine tegmentum, which contains the physiologically defined PPRF (paramedian pontine reticular formation), an important preoculomotor center. The results are discussed primarily with regard to their significance for potential cortical influence on the oculomotor system.

Animals↗
PubMed1986

Afferents to the abducens nucleus in the monkey and cat.

The abducens nucleus is a central coordinating element in the generation of conjugate horizontal eye movements. As such, it should receive and combine information relevant to visual fixation, saccadic eye movements, and smooth eye movements evoked by vestibular and visual stimuli. To reveal possible sources of these signals, we retrogradely labeled the afferents to the abducens nucleus by electrophoretically injecting horseradish peroxidase into an abducens nucleus in four monkeys and two cats. The histologic material was processed by the tetramethyl benzidine (TMB) method of Mesulam. In both species the largest source of afferents to the abducens nucleus was bilateral projections from the ventrolateral vestibular nucleus and the rostral pole of the medial vestibular nucleus. Scattered neurons were also labeled in the middle and caudal levels of the medial vestibular nucleus. Large numbers of neurons were labeled in the ventral margin of the nucleus prepositus hypoglossi in the cat and in the common margin of the nucleus prepositus and the medial vestibular nucleus in the monkey, a region we call the marginal zone. Substantial numbers of retrogradely labeled neurons were found in the dorsomedial pontine reticular formation both caudal and rostral to the abducens nuclei. In the monkey, large numbers of labeled neurons were present in the contralateral medial rectus subdivision of the oculomotor complex, while smaller numbers occurred in the ipsilateral medial rectus subdivision and elsewhere in the oculomotor complex. In the cat, large numbers of retrogradely labeled cells were present in a small periaqueductal gray nucleus immediately dorsal to the caudal pole of the oculomotor complex, and a few labeled neurons were also dispersed through the caudal part of the oculomotor complex. Occasional labeled neurons were present in the contralateral superior colliculus in both species. The size and distribution of the labeled neurons within the intermediate gray differed dramatically in the two species. In the cat, the retrogradely labeled neurons were very large and occurred predominantly in the central region of the colliculus, while in the monkey, they were small to intermediate in size and were distributed more uniformly within the middle gray. Among the afferent populations present in the monkey, but not in the cat, was a group of scattered neurons in the ipsilateral rostral interstitial nucleus of the medial longitudinal fasciculus and a denser, bilateral population in the interstitial nucleus of Cajal.(ABSTRACT TRUNCATED AT 400 WORDS)

Abducens Nerve↗
PubMed1986

The overlap of spinothalamic and dorsal column nuclei projections in the ventrobasal complex of the rat thalamus: a double anterograde labeling study using light microscopy analysis.

Projections from the spinal cord and the dorsal column nuclei (DCN) to the ventrobasal complex of the thalamus (VB) were studied in the rat by using double anterograde labeling strategy. This strategy was based on the injection of 3H-leucine into the DCN and of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into the spinal cord and their subsequent transport. Adjacent 30-micron-thick sections were then processed differentially for autoradiography or for HRP by using tetramethyl benzidine (TMB) as a chromogen. Similar areas of the ventrobasal complex were labeled, in adjacent sections, after a large injection of 3H-leucine into the DCN and when wheat germ agglutinin-HRP had been injected in any part of the spinal cord. If, however, a small injection of the radioactive tracer was centered in the gracile nucleus and compared with an injection of WGA-HRP placed in the lumbar enlargement of the cord, the rostral and dorsal portions of the lateral VB were labeled from both sources. On the other hand, if tritiated leucine was injected into the cuneate nucleus, and WGA-HRP placed in the cervical enlargement, then the caudal and ventral portions of the lateral VB demonstrated overlap of both labels. The present results show that, in the rat, areas of termination of both the spinothalamic tract and the lemniscal pathway originating from the DCN overlap in the lateral VB. This overlap is somatotopically organized, thus indicating that the same area of the VB receives somatic inputs from one particular part of the body through both pathways. These results are discussed in comparison to those of comparable studies performed in the cat and in the monkey and with reference to the electrophysiological data that have demonstrated that, in the rat VB, neurons responding to noxious stimulation are intermingled with neurons exclusively responding to non-noxious stimulation.

Afferent Pathways↗
PubMed1987

Amygdalofugal and amygdalopetal connections with modality-specific visual cortical areas in macaques (Macaca fuscata, M. mulatta, and M. fascicularis).

The origins and terminations of the amygdaloid connections with the modality-specific visual cortical areas TEa (anterior TE area), TEp (posterior TE area), TEO, V4, V2, MST (medial superior temporal visual area), MT (middle temporal visual area), and V1 were studied in macaques. These were compared with the amygdaloid connections of a vision-related polysensory area TG by making cortical injections of horseradish peroxidase (HRP) and incubating the sections with tetramethylbenzidine (TMB) as the chromogen. Both areas TEa and TEp receive a major projection from the lateral basal nucleus and a minor one from the accessory basal nucleus of the amygdaloid complex, whereas these areas send a major projection to the lateral nucleus and a minor one to the lateral basal nucleus. Areas TEO, V4, V2, MST, MT, and V1 receive projections only from the lateral basal nucleus; none of them project to any amygdaloid nucleus. Thus, the amygdalofugal projections are more widespread and more complex than the amygdalopetal projections. These findings indicate that the connections between the amygdaloid nuclei and the visual areas are generally nonreciprocal and underlie the importance of a feedback mechanism from the amygdala to the visual cortical areas in visual information processing. There appears to be a caudorostral (occipitotemporal) gradient in the distribution and density of the amygdaloid projections, which become progressively more widespread and heavier among the progressively more rostral visual areas (from area V1 to area TEa). The amygdaloid connections with area TG are distinctly different from the connections with the visual areas. Area TG is reciprocally connected mainly with the periamygdaloid cortex, and with the lateral, accessory basal, and medial basal nuclei of the amygdala as well.

Amygdala↗
PubMed1988

Sympathetic innervation of the rat cornea as demonstrated by the retrograde and anterograde transport of horseradish peroxidase-wheat germ agglutinin.

The sympathetic innervation of the rat cornea was studied by using the method of intraaxonal transport of horseradish peroxidase-wheat germ agglutinin conjugate (HRP-WGA). In the first set of experiments, the relative number of superior cervical ganglion neurons that innervate the rat central cornea was estimated by the method of retrograde HRP-WGA transport. Following tracer application to the scarificed central corneal surface, 49-198 labeled neurons were observed in the ophthalmic region of the ipsilateral trigeminal ganglion and zero to four cells in the rostral pole of the ipsilateral superior cervical ganglion. In the second set of experiments, the three-dimensional distribution and termination sites of the corneal sympathetic nerve fibers were investigated by the technique of HRP-WGA anterograde transport from the superior cervical ganglion. HRP-WGA-labeled axons in corneal whole mounts were identified by the presence within their axoplasm of linear arrays of HRP-TMB reaction product, and their distribution was plotted faithfully onto line drawings made with a drawing tube attachment. Large numbers of HRP-labeled fibers were found in all animals within the corneoscleral limbus where the majority were associated with blood vessels. Fewer fibers (zero to 14 per animal) entered the cornea proper. The latter fibers entered the peripheral cornea in the deep to middle layers of the stroma and ascended into progressively more superficial layers as they coursed centrally. The majority of fibers branched infrequently in the peripheral cornea and increased in branching complexity near the central cornea. HRP-labeled axonal varicosities suggestive of terminal and preterminal expansions were located preferentially in the subepithelial layer of the corneal stroma and in the basal epithelium. Approximately 75% of the axonal varicosities were located in the central half of the cornea. The results of the current investigation reveal that the rat cornea is innervated sparsely by sympathetic nerve fibers derived from the superior cervical ganglion. These data provide additional support to current theories that corneal sympathetic nerve fibers may influence select aspects of corneal physiology, including ion transport and hydration, mitogenesis and wound healing, and sensitivity.

Adrenergic Fibers↗
PubMed1988

Medial gastrocnemius motor nucleus in the rat: age-related changes in the number and size of motoneurons.

The age-related alterations in the number and size of alpha- and gamma-motoneurons were studied in the medial gastrocnemius (MG) motor nuclei in rats at four ages: young (5 months), middle aged (10-13 months), old (26 months), and very old (31 months). Small volumes (0.1-0.5 microliter) of 40% horseradish peroxidase (HRP) solution were injected into the cut MG nerve bilaterally by using glass micropipettes and a pressure injection system. The number, position, and soma size (average soma diameter) of MG motoneurons were determined by using photographic maps of each TMB-stained section. The total number of myelinated axons was counted in seven MG nerves from the same animals. The average soma diameters in each MG nucleus were distributed bimodally; cells with average diameter greater than 21.0-24.0 micron were presumed to be alpha-motoneurons and those with smaller diameters were presumed to be gamma. The mean number of presumed alpha-motoneurons was significantly less in the old and very old groups as compared with the young and middle-aged. In contrast, the number of presumed gamma-motoneurons was the same across age groups. The mean average soma diameter of both alpha- and gamma-motoneurons was smaller in the old animals. The apparent decrease in the total number of labeled motoneurons in old animals was also reflected in a decrease in myelinated axon counts. We conclude that there is a significant decrease in the absolute numbers of motoneurons in rats aged 26 months and older, with most of the decrease occurring among the larger alpha-motoneurons.

Aging↗
PubMed1988

Topographical and ultrastructural investigation of the habenulo-interpeduncular pathway in the rat: a wheat germ agglutinin-horseradish peroxidase anterograde study.

The topographical and ultrastructural organization of the habenular projection to the interpeduncular nucleus (IPN) of the rat was examined employing the anterogradely transported tracer wheat germ agglutinin-horseradish peroxidase (WGA-HRP) and the chromogen tetramethylbenzidine (TMB). Unilateral placements of WGA-HRP in the habenular complex resulted in heavy terminal labelling in the rostral, central, and intermediate subnuclei bilaterally, and in the lateral subnuclei ipsilaterally. The apical subnucleus possessed only a sparse amount of label. Placements confined to the medial habenula (mH) produced similar results to those observed when the entire habenula was filled, suggesting that the afferent contribution made by the lateral habenula (lH) to the IPN is small. Unilateral placements of WGA-HRP in the dorsal portion of the mH resulted in heavy, predominantly ipsilateral labelling in the lateral subnucleus and the dorsal cap of the rostral subnucleus. In the lateral subnucleus labelled habenular terminals consistently contacted single dendritic processes shared by one or more other boutons, possibly of nonhabenular origin. Labelled habenular terminals in the rostral subnucleus normally contacted one or two dendrites. Labelled terminals in both subnuclei possessed clear, spherical vesicles and a variable number of dense-core vesicles. Unilateral placements of WGA-HRP in the ventral portion of the mH resulted in heavy labelling in the rostral half of the rostral subnucleus with a slight ipsilateral predominance, and in the central and intermediate subnuclei bilaterally. Terminal labelling was observed in crest and S synapses in the intermediate and central subnuclei respectively. Crest synapses, which consist of two parallel habenular terminals contacting an attenuated dendritic process, normally possessed label in only one of the two boutons. In the central subnucleus labelled horizontal axons formed several en passant S synapses with dendritic processes of small and medium diameter. These synaptic specializations of habenular axons contained numerous clear, spherical vesicles. This study demonstrates that a major topographically organized projection to the IPN originates from two distinct subpopulations of habenular neurons which comprise a dorsal sector and a ventral sector of the mH. Ultrastructural examination demonstrated that axons originating from neurons in the ventral and dorsal mH form characteristic contacts in the various IPN subnuclei.

Animals↗
PubMed1988

Sharpening of topographical projections and maturation of geniculocortical axon arbors in the hamster.

During specification of orderly neural maps, axons correctly navigate to their targets and form terminal arbors in topographically correct positions. To learn more about this mapping process, the patterns of geniculocortical topography were correlated with growth of axon arbors in the hamster visual cortex. Topography was studied by retrograde transport of WGA-HRP from area 17 to the dorsal lateral geniculate nucleus (LGd) and visualized with TMB histochemistry. In separate experiments, geniculocortical axon arbors were filled with HRP deposited extracellularly into the optic radiations and stained with cobalt-intensified DAB. On the day of birth (P0) and on P1-2, a crude topography was detected in the geniculocortical system. At these ages, geniculocortical axons coursed in the embryonic white matter of the visual cortex, parallel to the pia. During their passage, multiple short collaterals, with no terminal arbors, were extended into the subplate and deeper portions of the cortical plate. By P3-5, the topography was more precise and simple axonal arbors had now begun to be formed on some branches within the cortical plate. During the second postnatal week, branches in the white matter without terminals were eliminated and the ramifications of branches in the gray matter became more elaborate. The arbors continued to increase in complexity and resembled adult forms by P24.

Animals↗
PubMed1991

All peroneal motoneurons of the rat survive crush injury but some fail to reinnervate their original targets.

This is a quantitative study of the motoneuronal population of the rat's common peroneal nerve following severe crush injury of the sciatic nerve or its component branches. The crush was performed unilaterally under anesthesia for 60 seconds with hemostat jaws covered with tubing to form a smooth, 2 mm long, injured zone. Recovery from injury was allowed for 14 to 188 days. It was measured behaviorally using the sciatic functional index (SFI) and electrophysiologically by comparing the conduction velocity and amplitudes of evoked muscle action potentials prior to injury, and again after injury just before the nerve was labeled with horseradish peroxidase (HRP), and/or its wheat germ agglutinin conjugate (WGA-HRP), 48-72 hours before sacrifice. The motoneurons were retrogradely labeled on both sides so that the uninjured side might serve as a control. On the injured side the nerves were labeled either distal or proximal to the crush site. The tibialis anterior muscles on both sides were removed and weighted. Spinal segments L2 to L6 were cut in serial, frozen cross-sections. HRP reaction products were formed using TMB as the chromogen. The normal peroneal nerve was found to contain 634 +/- 26 motoneurons (22 cases). The number of motoneurons labeled 5-15 mm distal to the injury site (22 cases) was 535 +/- 69 or 84.4% of normal. In 12 cases in which the nerve was labeled 5 mm proximal to the injury normal population numbers (648 +/- 30) were found. These counts demonstrated that the unlabeled 15.6% in the distal labeled cases had not vanished as a result of cell death. Instead, the unlabeled group was composed mainly of small motoneurons whose axons probably had not regenerated distal to the crushed zone. Mean soma size of injured neurons increased to maximum 3-6 weeks after injury and then gradually decreased in size over the following weeks to nearly normal values. This transient increase in size was due to two factors: 1) soma swelling in response to axonal injury, and 2) absence of many small motoneurons, presumably gamma-motoneurons, which were either incapable of, or prevented from, regenerating beyond the injury zone long after larger motoneurons had reinnervated their targets. SFI scores, muscle weights, and amplitude ratios of evoked potentials recovered to control values by 70-80 days post-injury. Conduction velocities remained 20-25% below normal at the end of 80 days.(ABSTRACT TRUNCATED AT 400 WORDS)

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