Transganglionic Horseradish Peroxidase Research Articles

Evidence for prenatal competition among the central arbors of trigeminal primary afferent neurons

Previous studies have shown that damage to vibrissa follicles in newborn rats and mice does not alter the brainstem representations of the remaining vibrissa as demonstrated by staining for mitochondrial enzymes such as cytochrome oxidase (CO) succinic dehydrogenase. This study asked whether this lack of effect might be due to the fact that the trigeminal primary afferents in rodents are already quite well developed at birth. We assessed this possibility by using CO staining the evaluate patterns in the brainstems of pre- and postnatal rats. A vibrissa-related pattern began to emerge in trigeminal nucleus principalis and subnucleus interpolaris (Spl) by embryonic day (E-) 19 and appeared fully developed by the day of birth (P-0). We also made partial lesions of the vibrissa pad on E-15-20 and on P-0, killed pups on P-5-7, and measured the size of the CO-stained patches in Spl on both sides of the brainstem. The correspondence between CO patches and clusters of primary afferent terminal arbors was verified in some animals by combining transganglionic horseradish peroxidase tracing and CO staining. Vibrissa pad damage on E-15-18 resulted in significant (20.1-36.9%) increases in the average area of the remaining CO patches in Spl ipsilateral to the lesion. Vibrissa pad damage on E-19, E-20, and P-0 produced small (6.2-8.9%), but insignificant, increases in patch size in Spl ipsilateral to the lesion. We used anatomical and electrophysiological methods to determine whether our lesions altered the trigeminal innervation of surviving vibrissa follicles. We recorded single trigeminal ganglion cells from 12 rats that sustained vibrissa pad lesion on E-17. As in normal rats, all of the 49 vibrissa-sensitive ganglion cells isolated in the lesioned animals were responsive to deflection of one and only one vibrissa. We also dissected 11 deep vibrissal nerves from intact follicles in adult rats that sustained fetal vibrissa pad damage on E-17, and counted numbers of myelinated axons in 1 microns plastic sections. These data were compared with counts from corresponding follicles on the intact side of the face. The average number of myelinated axons innervating follicles in the damaged vibrissa pads was 196.8 +/- 27.9, and that for the corresponding contralateral nerves was 194.6 +/- 25.7. These data suggest that competitive interactions among the central arbors of trigeminal primary afferents in fetal life may influence the development of central vibrissa representations and, further, that lesion-induced central changes need not be correlated with alterations in the peripheral innervation of undamaged follicles.

Brainstem projections of different branches of the vestibular nerve: An experimental study by transganglionic transport of horseradish peroxidase in the cat

The brainstem projections of the ampullar nerves from the vertical semicircular canals, the anterior (AAN) and the posterior ampullar nerve (PAN), were studied in adult cats using the transganglionic horseradish peroxidase (HRP) method. Each nerve was exposed in three experiments. Two animals in each group had labeling which allowed detailed mapping. From the AAN, terminal-type labeling was found in two separate groups, one laterally and one medially, both in the lateral (LV) and in the superior (SV) vestibular nucleus. In addition, such labeling was found in all parts of the medial vestibular nucleus (MV). Labeled structures were found also in the descending vestibular nucleus, (DV) more densely over its lateral part, except for cell group f, where no labeling was found. From the PAN, terminal-type labeling was found medially and laterally in the LV and in the medial part of the SV. In the MV, such labeling was evenly distributed rostrally but concentrated laterally in caudal parts. In the DV, terminal-type labeling was present rostrally, whereas no labeling was seen caudally. In the interstitial nucleus of the vestibular nerve, terminal-type labeling was observed from the AAN but not from the PAN. No labeled fibers from either of the two ampullar nerves were seen outside the vestibular root and nuclei, except for small-caliber fibers from the SV heading towards the brachium conjunctivum. The findings clearly indicate a specific termination for each of the two ampullar nerves.

Mystacial vibrissae representation within the trigeminal sensory nuclei of the cat

Somatotopic arrangements of axon terminals of primary afferent fibers innervating follicles of the mystacial vibrissae were examined in the cat by the transganglionic horseradish peroxidase (HRP) method. Forty to 60 hours after injecting HRP into a single or a group of vibrissal follicles, transported HRP was visualized by the tetramethylbenzidine technique. HRP-labeled axon terminals were distributed in the ventral subnucleus of the principal sensory trigeminal nucleus (ventral Vp), in the oral and interpolar spinal trigeminal nuclei (Vo and Vi), and in the caudal spinal trigeminal nucleus (Vc) (layer I, deep part of layer II, layers III-V) with its spinal extension into the dorsal horn of the first cervical cord segment (rostral C1). In cross sections through the caudal parts of the ventral Vp, Vi, and layer IV of the Vc and rostral C1, a single mystacial vibrissa was represented in a one-to-one fashion by a patch of dense terminal arbors of primary afferent fibers. The more dorsally a horizontal row of the mystacial vibrissae was located, the more ventrally was it represented in the ventral Vp, the more ventrolaterally in the Vi, and the more ventrally in layer IV of the Vc and the rostral C1. In addition, the more anteriorly a vibrissa was located in a horizontal row of the mystacial vibrissae, the more medially was it represented in the ventral Vp, the more ventromedially in the Vi, and the more laterally in layer IV of the Vc and rostral C1; the most posteriorly located vibrissae in the horizontal rows of the mystacial vibrissae were represented along the lateral border of the ventral Vp and Vi, and most medially in layer IV of the Vc and rostral C1. Thus, the representation pattern in the ventral Vp was rotated clockwise at about 45 degrees angle in the Vi, and projected as a mirror image in layer IV of the Vc and rostral C1. It was also indicated that the anterior-posterior arrangement of the mystacial vibrissae was represented in a rostral-caudal organization within layer IV of the Vc and rostral C1. It was also indicated that the anterior-posterior arrangement of the mystacial vibrissae was represented in a rostral-caudal organization within layer IV of the Vc and rostral C1. Patchy patterns probably replicating the distribution of the vibrissae on the face of the cat were also revealed by the cytochrome oxidase histochemical staining in cross sections through the caudal parts of the ventral Vp, Vi, and layer IV of the Vc and rostral C1.

Trigeminal nerve branches containing primary afferent fibers to the dorsal division of the principal sensory trigeminal nucleus: A transganglionic horseradish peroxidase study in the cat

Trigemial nerve branches containing primary afferent fibers to the dorsal division of the principal sensory trigeminal nucleus (Vp) were examined in the cat by the transganglionic horseradish peroxidase method. The results indicate that the pterygopalatine, superior alveolar, infraorbital, buccal, lingual, inferior alveolar and mental nerves contain primary afferent fibers terminating in the dorsal division of the Vp probably conveying sensory information chiefly from the intraoral structures.

The central projections of primary afferent neurons of greater splanchnic and intercostal nerves in the rat. A horseradish peroxidase study.

The central projections of primary afferent fibers of the greater splanchnic nerve of the rat were investigated using the transganglionic horseradish peroxidase transport technique. In addition, the corresponding spinal ganglion cells and the preganglionic sympathetic neurons were demonstrated. For comparing visceral and somatic afferents, intercostal nerve afferents were labelled by the same technique. Splanchnic afferent dorsal root ganglion cells were found at segments T3 to T13 ipsilaterally, with the greatest density at T8 to T12. Labelled cells represented about 10%-15% of all neurons in the ganglia at maximal projection levels. They were randomly distributed within individual ganglia. The great majority were medium to small sized and round to slightly oval in shape. In the spinal cord, labelled visceral afferent axons were found maximally at T8 to T11, but could be detected in decreasing density up to T1 and down to L1. They were distributed over Lissauer's tract and the dorsal funiculus to a medial and lateral collateral pathway (MCP and LCP, respectively). The MCP, somewhat more prominent than the LCP, was destined primarily to clustered presumptive terminal fields in medial lamina I and outermost lamina IIa. Only a few axons continued further to laminae V and X. Splanchnic afferent axons, most likely derived from the MCP, formed a longitudinal bundle ventral to the central canal. The LCP consisted of more or less well-defined axon bundles emanating from the lateral Lissauer's tract and curving round the lateral edge of the dorsal horn and through the dorsolateral funiculus. Presumptive terminal sites of LCP axons are the lateral laminae I and IIa, the nucleus of the dorsolateral funiculus and the dorsal part of lamina V. A few LCP axons were seen in the vicinity of lateral dendrites of preganglionic sympathetic axons. Visceroafferent terminals were absent from laminae IIb-IV and VII. The possible consequences of the MCP/LCP duality for the central connections of splanchnic afferents are discussed. Some splanchnic afferents ascended to the gracile and cuneate nuclei, and rarely to the spinal trigeminal nucleus. These results fit into the general concept of visceroafferent terminal organization that has emerged during the last few years. Differences to other reports in the detailed arrangement of fibers and terminals are discussed. Somatoafferent cell bodies represented the vast majority of neurons in the respective spinal ganglia. Cell sizes encompassed the whole range from very small to very large without a clear predominance of one particular size class.(ABSTRACT TRUNCATED AT 400 WORDS)

Primary vestibular projections in the Hagfish, Eptatretus burgeri

The VIIIth cranial nerve projections in the hagfish, which has only one circular canal in the ear, were studied by transganglionic HRP transport. This nerve has two branches, the nervus utricularis (N. utr.) and the nervus saccularis (N. sac.), each with its own ganglion, the ganglion utriculare (G. utr.) and the ganglion sacculare (G. sac.), respectively. Although the G. sac. has uniformly small cells, the G. utr. consists of two separate cell masses, a ventral mass of large cells and a dorsal mass of small cells. The small cells were labeled in both ganglia after horseradish peroxidase (HRP) injection into the endolymphatic space. The greater part of the terminal areas of these two branches overlapped in the ventral nucleus of the area acoustico-lateralis, but the terminals of the N. sac. extended slightly further in a caudal direction. No projections to the primordial cerebellum and no retrogradely labeled cells in the brain were found. The large cells in the ventral part of the G. utr. seem to be general cutaneous neurons, and the dorsal part of the area acousticolateralis seems to receive lateral line input.

Ultrastructure of transganglionic HRP transport in cat trigeminal system

The ultrastructure of transganglionic transport of horseradish peroxidase (HRP) from the inferior alveolar (IA) nerve to the brainstem is being studied in the cat. The IA nerve was soaked in an HRP solution and following a two-day survival the animal was perfused transcardially with a paraformaldehyde-glutaraldehyde solution. The tissue was immediately dissected and postfixed for 1–3 h in perfusate. Sections of 75 μm thickness were cut with a Vibratome and reacted utilizing tetramethyl benzidine (TMB) as the chromagen. Optimum results for electron microscopy were obtained by osmication in a pH 6.0, 1% osmium tetroxide solution for 45 min at 45 °C, followed by rapid dehydration and embedment in Epon. The resulting HRP-TMB reaction product was characterized and identified ultrastructurally in ganglion cells, peripheral and central axons and in brainstem terminals. The HRP-TMB reaction product varied in density but had consistent crystalline-like laminations of a repeating unit and characterized by a membrane 4–5 nm in diameter. Some of the HRP-TMB reaction product found in terminals and axons was below the limit of resolution of the light microscope.

Central distribution of afferent fibers in the intermediate nerve: A transganglionic HRP study in the cat

Afferent fibers in the two components of the intermediate nerve of the cat, i.e. the chorda tympani (CTN) and greater petrosal (GPN) nerves, were traced into the lower brainstem transganglionically after applying horseradish peroxidase (HRP) to the central cut end of the CTN or GPN. CTN fibers ended in the trigeminal (principal sensory and spinal trigeminal nuclei) and solitary (medial, interstitial and dorsal solitary nuclei) nuclei, whereas GPN fibers ended only in the solitary nucleus.

Central projections of the normal and ‘regenerate’ infraorbital nerve in adult rats subjected to neonatal unilateral infraorbital lesions: a transganglionic horseradish peroxidase study

Application of horseradish peroxidase (HRP) to the proximal stump of the transected infraorbital (IO) nerve in the orbit was employed in normal adult rats to demonstrate the ganglionic representation of and brainstem innervation by the trigeminal (V) branch which supplies primarily the mystacial vibrissae. In the V ganglion, somal clusters resembling the barrels or barreloids which comprise the central representations of the whiskers were not visible. In the trigeminal brainstem nuclear complex (TBNC), however, barrel-like aggregates of HRP-labeled terminals could be clearly seen throughout subnucleus interpolaris, and to a lesser extent, in the principal sensory nucleus, subnucleus caudalis and the first cervical segment. In adult rats subjected to neonatal IO nerve section and vibrissae follicle cauterization, transganglionic HRP transport from the ‘regenerate’ IO nerve indicated an almost exclusive projection to the marginal layer of the medullary and rostral cervical dorsal horn.

Functional properties of neurons in cat trigeminal subnucleus caudalis (medullary dorsal horn). I. Responses to oral-facial noxious and nonnoxious stimuli and projections to thalamus and subnucleus oralis.

Functional properties of neurons in cat trigeminal subnucleus caudalis (medullary dorsal horn). I. Responses to oral-facial noxious and nonnoxious stimuli and projections to thalamus and subnucleus oralis.

Four aspects of trigeminal nucleus and a paradox.

Four aspects of trigeminal nucleus and a paradox.