Evidence for survival of the central arbors of trigeminal primary afferents after peripheral neonatal axotomy: experiments with galanin immunocytochemistry and Di-I labelling.

Studies employing axoplasmic transport techniques have suggested that the central arbors of vibrissae-related primary afferents are rapidly and permanently lost from the trigeminal (V) brainstem complex after transection of the intraorbital nerve (ION). The present study reexamined this issue using immunocytochemistry for galanin (GAL) and anterograde labelling with Di-I to evaluate V brainstem organization in rats that sustained damage to the ION or individual vibrissae follicles in infancy or adulthood. After adult nerve damage, GAL-positive fibers are increased in layers I and II of V subnucleus caudalis (SpC). This was apparent by 3 days after the lesion. In rats that sustained nerve damage at birth (P0), GAL immunoreactivity (IR) appeared throughout the V brainstem complex and had a patchy distribution similar to that of vibrissae-related V primary afferents in normal rats. Increased GAL-IR in rostral portions of the V brainstem complex was observed in rats that sustained ION damage as late as P14. Additional experiments in which nerve damage was followed by destruction of the V ganglion demonstrated that this GAL-IR was contained in primary afferents. Damage to single vibrissa follicles or to a row of follicles produced a single patch or row of GAL-IR terminals in the somatotopically appropriate portion of the ipsilateral V brainstem complex. Di-I labelling in neonatally nerve-damaged rats demonstrated that primary afferent axons filled the central territory normally innervated by this nerve and that their terminal distribution was patchy. These results suggest that the V ganglion cells that survive neonatal axotomy may retain somatotopically organized projections to the V brainstem complex for at least a limited postnatal period.

Immunochemical heterogeneity in the tecto-LP pathway of the rat.

The projection from the rat's superior colliculus (SC) to the lateral posterior nucleus of the thalamus (LP) has previously been described as arising from a morphologically homogeneous population of neurons in the stratum opticum (SO). The present study combined immunocytochemistry with retrograde tracing and lesion techniques to determine whether or not the SC-->LP projection arose from neurons that were also neurochemically homogeneous. The combination of retrograde tracing and immunocytochemistry with an antibody directed against calbindin-D 28K (CBD) showed that 64.4% of the neurons that project from SC to LP contain this calcium-binding protein. Retrograde tracing and immunocytochemistry for adenosine deaminase (ADA) showed that a smaller number of tecto-LP cells (15.7%) were immunoreactive (IR) for this enzyme. Moreover, nearly all (93.0%) of the ADA-IR tecto-LP cells also contained CBD-IR. Adenosine deaminase-IR axons in LP were restricted to the dorsomedial portion of the nucleus and their density was substantially reduced after ablation of the ipsilateral superficial SC laminae. The lateral posterior nucleus contained numerous CBD-IR cells and fibers throughout its extent and it was thus difficult to determine the extent to which the extra-perikaryal CBD-IR in this nucleus was dependent upon the tecto-LP pathway. Nevertheless, destruction of the ipsilateral SC did reduce the density of CBD-IR in LP. These results suggest that the SC-->LP projection in rat arises from at least four neurochemically distinct cell groups: 1) those that contain CBD, 2) those that contain both CBD and ADA, 3) a very small population that contains only ADA, and 4) a group that is not recognized by either of these markers. Our results further suggest that ADA containing fibers may have a more restricted terminal distribution in LP than axons that contain only CBD.

Lesion-induced changes in the central terminal distribution of galanin-immunoreactive axons in the dorsal column nuclei.

Rats that sustained forelimb removal on either embryonic day (E) 16, on the day of birth (P-0), or transection of the brachial plexus in adulthood had brainstem sections stained for galanin, calcitonin gene-related peptide (CGRP), or substance P (SP) at various intervals after these lesions were made. In normal adult rats, only a few galanin-immunoreactive fibers are present in the cuneate nucleus and most are located in its caudal portion. CGRP-positive axons are also sparse in the cuneate and are distributed mainly in the periphery of the nucleus. SP-positive axons are seen throughout the cuneate nucleus. In rats that sustained forelimb removals at birth or transection of the brachial plexus in adulthood, dense galanin immunoreactivity was present throughout the cuneate nucleus at all rostrocaudal levels on the side of the brainstem ipsilateral to the lesion. The changes after lesions that were made in the adult animals were apparent within 1 week, the earliest time analyzed. Increases in galanin immunoreactivity in the cuneate of animals that sustained forelimb removals on P-0 were first visible on P-2. Neither forelimb removal at birth nor brachial plexus lesions in adulthood had any qualitative effect upon the distribution or density of CGRP- or SP-immunoreactivity in the cuneate nucleus. Removal of a forelimb on E-16 did not increase the density of galanin-immunoreactive fibers in the cuneate nucleus. Such lesions also failed to produce any appreciable change in the density of either CGRP- or SP-positive fibers in the cuneate nucleus. The present data raise the possibility that large caliber, non-peptidergic primary afferent axons which innervate the cuneate nucleus may express galanin after damage at birth or in adulthood.

Galanin immunoreactivity reveals a vibrissae-related primary afferent pattern in perinatal rats after neonatal infraorbital nerve transection.

Immunocytochemistry was used to demonstrate the distribution of galanin immunoreactivity in the trigeminal (V) brainstem complex of normal rats and rats that sustained neonatal transection of the infraorbital nerve (ION, the V branch that supplies the mystacial vibrissae). In normal adult and perinatal (postnatal day [P-] 7) rats, there is very little galanin immunoreactivity in the rostral part of the V brainstem complex. However, there is dense immunoreactivity for this peptide in layers I and II of V subnucleus caudalis (SpC). There was a marked upregulation of galanin immunoreactivity at all levels of the V brainstem complex of P-7 rats that sustained ION transection on P-0. In V subnucleus interpolaris and the magnocellular part of SpC, this immunoreactivity occurred in clusters which had a pattern resembling that of the mystacial vibrissae. In animals examined at 30 days or more after ION transection, there were a few more galanin-immunoreactive fibers on the deafferented side of the brainstem than on the intact side, but there was no evidence of a vibrissae-related pattern. Our interpretation of these results is that galanin immunocytochemistry reveals a vibrissae-related pattern in the central arbors of axotomized primary afferents for at least 1 week after they have been disconnected from the periphery.

Development and plasticity of the serotoninergic projection to the hamster's superior colliculus.

Immunocytochemical techniques were employed to investigate the normal adult organization, development, and effects of both neonatal and adult eye removal upon the organization of the serotoninergic projection to the hamster's superior colliculus. Immunocytochemistry, both alone and in combination with retrograde tracing with true blue and fluorogold, was used to determine the organization of the serotoninergic projection to the superior colliculus in normal adult hamsters. Immunoreactive fibers were present in all laminae of the superior colliculus, but they were most dense in the lower part of the stratum griseum superficiale, the stratum opticum, the stratum griseum profundum, and the stratum album profundum. When retrograde tracing from the colliculus was combined with immunocytochemistry for serotonin, cells containing both labels were found in the lateral portions of the nucleus raphe dorsalis and also in periaqueductal gray, the median and pontine raphe nuclei, and in the region of the medial lemniscus. Such cells were visible both ipsilateral and contralateral to the injection site. Serotonin immunoreactive fibers were visible in the superior colliculus by embryonic day 14 (2 days prior to birth). On the day of birth, a small number of immunoreactive fibers were present just below the pial surface and others were generally oriented either parallel or orthogonal to the collicular laminae. At this age, there were also many immunoreactive fibers that crossed from one side of the brainstem to the other in the commissure of the superior colliculus. Some serotonin-positive axons appeared to terminate as growth cones in fetal and newborn hamsters. Over the next 2 weeks, the serotoninergic innervation of the SC increased in density and assumed the laminar distribution observed in adult animals. By this age, only a very few immunoreactive fibers were present in the commissure of the superior colliculus. Removal of one eye on either on the day of birth or in adulthood resulted in reorganization of the serotoninergic innervation of the partially deafferented colliculus. There was a marked increase in the density of serotonin-positive fibers in the upper stratum griseum superficiale on the side ipsilateral to the remaining eye. This change was apparent within 2 weeks after enucleation in either neonatal or adult animals. Quantitative analysis demonstrated that the magnitude of this effect was greater after adult enucleation than after neonatal eye removal. Both neonatal and adult enucleation also resulted in an increase in the density of the serotoninergic projection to the dorsal and ventral lateral geniculate nuclei ipsilateral to the remaining eye.