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Background and Objectives A persistent increase in calcitonin gene-related peptide (CGRP) immunoreactivity in motoneurons may serve as an indicator for regeneration after peripheral nerve injury [Borke et al., J Neurocytol 1993;22:141–153]. Study Design/Materials and Methods We examined the effects of low power laser treatment (633 nm) on axotomy-induced changes in α-CGRP mRNA and long-term neuronal survival in facial motoneurons. A quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) assay for α-CGRP mRNA was used to detect changes in the response to axotomy and laser irradiation. Cell counts of neurons in injured and non-injured facial motor nuclei of laser-treated and non-treated rats were done to estimate neuronal survival. Results A 10-fold increase (P

Low power laser irradiation has been reported to cause biological effects due to the photochemical and/or photophysical action of the radiation. This study determined quantitatively if transcu-taneous low power laser irradiation can affect the regeneration of the rat facial nerve. The facial nerve was crushed unilaterally in anesthetized rats and transcutaneously irradiated daily with a laser beam directed at the area of the crush injury. Laser treatment began on the day of the crush injury and was continued daily for 7, 8, or 9 days. Preliminary experiments determined the most effective wavelength, laser power, length of irradiation, and treatment schedule. The wavelengths examined were 361, 457, 514, 633, 720, and 1064. The laser powers and lengths of irradiation examined ranged from 8.5 to 40 mW and 13 to 120 min. Irradiation treatment was done daily, on alternating days and on the first 4 days postcrush. The most effective laser parameters for the low power treatment included daily irradiation with a helium-neon (HeNe) or argon pumped tunable dye laser a wavelength of 633 nm, with a power of 8.5 mW for 90 minutes (45.9 J, 162.4 J/cm2). The number of horseradish peroxidase (HRP) labeled neurons in the facial motor nucleus was used as an assay of the degree of regeneration. In rats in which the facial nerve was crushed but not irradiated, the average number of HRP labeled neurons in the facial nucleus was 22 on day 7 postcrush, 54 on day 8, 116 on day 9, and 1,149 on day 10. After HeNe or argon pumped tunable dye laser irradiation, the average number of HRP-labeled neurons increased to 34 on day 7 postcrush, 148 on day 8, and 1,725 on day 9. There was a statistically significant difference between the control and irradiated rats on day 9 post-crush (P < 0.01). These data indicate that transcutaneous low power irradiation with the lasers and parameters involved in this study increased the rate of regeneration of rat facial nerve following crush injury. © 1993 Wiley-Liss, Inc.

Background and Objective Familial amyotrophic lateral sclerosis (FALS) is a neurodegenerative disease characterized by progressive loss of motor neurons and death. Mitochondrial dysfunction and oxidative stress play an important role in motor neuron loss in ALS. Light therapy (LT) has biomodulatory effects on mitochondria. Riboflavin improves energy efficiency in mitochondria and reduces oxidative injury. The purpose of this study was to examine the synergistic effect of LT and riboflavin on the survival of motor neurons in a mouse model of FALS. Study Design/Materials and Methods G93A SOD1 transgenic mice were divided into four groups: Control, Riboflavin, Light, and Riboflavin+Light (combination). Mice were treated from 51 days of age until death. A single set of LT parameters was used: 810 nm diode laser, 140-mW output power, 1.4 cm2 spot area, 120 seconds treatment duration, and 12 J/cm2 energy density. Behavioral tests and weight monitoring were done weekly. At end stage of the disease, mice were euthanized, survival data was collected and immunohistochemistry and motor neuron counts were performed. Results There was no difference in survival between groups. Motor function was not significantly improved with the exception of the rotarod test which showed significant improvement in the Light group in the early stage of the disease. Immunohistochemical expression of the astrocyte marker, glial fibrilary acidic protein, was significantly reduced in the cervical and lumbar enlargements of the spinal cord as a result of LT. There was no difference in the number of motor neurons in the anterior horn of the lumbar enlargement between groups. Conclusions The lack of significant improvement in survival and motor performance indicates study interventions were ineffective in altering disease progression in the G93A SOD1 mice. Our findings have potential implications for the conceptual use of light to treat other neurodegenerative diseases that have been linked to mitochondrial dysfunction. Lasers Surg. Med. 41:52–59, 2009. © 2008 Wiley-Liss, Inc.

Background and Objectives Light therapy has biomodulatory effects on central and peripheral nervous tissue. Spinal cord injury (SCI) is a severe central nervous system trauma with no effective restorative therapies. The effectiveness of light therapy on SCI caused by different types of trauma was determined. Study Design/Materials and Methods Two SCI models were used: a contusion model and a dorsal hemisection model. Light (810 nm) was applied transcutaneously at the lesion site immediately after injury and daily for 14 consecutive days. A laser diode with an output power of 150 mW was used for the treatment. The daily dosage at the surface of the skin overlying the lesion site was 1,589 J/cm2 (0.3 cm2 spot area, 2,997 seconds). Mini-ruby was used to label corticospinal tract axons, which were counted and measured from the lesion site distally. Functional recovery was assessed by footprint test for the hemisection model and open-field test for the contusion model. Rats were euthanized 3 weeks after injury. Results The average length of axonal re-growth in the rats in the light treatment (LT) groups with the hemisection (6.89±0.96 mm) and contusion (7.04±0.76 mm) injuries was significantly longer than the comparable untreated control groups (3.66±0.26 mm, hemisection; 2.89±0.84 mm, contusion). The total axon number in the LT groups was significantly higher compared to the untreated groups for both injury models (P

Ts65Dn, a mouse model of Down syndrome (DS), demonstrates abnormal hippocampal synaptic plasticity and behavioral abnormalities related to spatial learning and memory. The molecular mechanisms leading to these impairments have not been identified. In this study, we focused on the G-protein-activated inwardly rectifying potassium channel 2 (GIRK2) gene that is highly expressed in the hippocampus region. We studied the expression pattern of GIRK subunits in Ts65Dn and found that GIRK2 was overexpressed in all analyzed Ts65Dn brain regions. Interestingly, elevated levels of GIRK2 protein in the Ts65Dn hippocampus and frontal cortex correlated with elevated levels of GIRK1 protein. This suggests that heteromeric GIRK1-GIRK2 channels are overexpressed in Ts65Dn hippocampus and frontal cortex, which could impair excitatory input and modulate spike frequency and synaptic kinetics in the affected regions. All GIRK2 splicing isoforms examined were expressed at higher levels in the Ts65Dn in comparison to the diploid hippocampus. The pattern of GIRK2 expression in the Ts65Dn mouse brain revealed by in situ hybridization and immunohistochemistry was similar to that previously reported in the rodent brain. However, in the Ts65Dn mouse a strong immunofluorescent staining of GIRK2 was detected in the lacunosum molecular layer of the CA3 area of the hippocampus. In addition, tyrosine hydroxylase containing dopaminergic neurons that coexpress GIRK2 were more numerous in the substantia nigra compacta and ventral tegmental area in the Ts65Dn compared to diploid controls. In summary, the regional localization and the increased brain levels coupled with known function of the GIRK channel may suggest an important contribution of GIRK2 containing channels to Ts65Dn and thus to DS neurophysiological phenotypes.

This study takes advantage of fibroblast growth factor 2 (FGF2) knock-out mice to determine the contribution of FGF2 to the regeneration of oligodendrocytes in the adult CNS. The role of FGF2 during spontaneous remyelination was examined using two complementary mouse models of experimental demyelination. The murine hepatitis virus strain A59 (MHV-A59) model produces focal areas of spinal cord demyelination with inflammation. The cuprizone neurotoxicant model causes extensive corpus callosum demyelination without a lymphocytic cell response. In both models, FGF2 expression is upregulated in areas of demyelination in wild-type mice. Surprisingly, in both models, oligodendrocyte repopulation of demyelinated white matter was significantly increased in FGF2 -/- mice compared with wild-type mice and even surpassed the oligodendrocyte density of nonlesioned mice. This dramatic result indicated that the absence of FGF2 promoted oligodendrocyte regeneration, possibly by enhancing oligodendrocyte progenitor proliferation and/or differentiation. FGF2 -/- and +/+ mice had similar oligodendrocyte progenitor densities in normal adult CNS, as well as similar progenitor proliferation and accumulation during demyelination. To directly analyze progenitor differentiation, glial cultures from spinal cords of wild-type mice undergoing remyelination after MHV-A59 demyelination were treated for 3 d with either exogenous FGF2 or an FGF2 neutralizing antibody. Elevating FGF2 favored progenitor proliferation, whereas attenuating endogenous FGF2 activity promoted the differentiation of progenitors into oligodendrocytes. These in vitro results are consistent with enhanced progenitor differentiation in FGF2 -/- mice. These studies demonstrate that the FGF2 genotype regulates oligodendrocyte regeneration and that FGF2 appears to inhibit oligodendrocyte lineage differentiation during remyelination.

Ensheathing cells exclusively enfold olfactory axons. The ability of olfactory axons to reinnervate the adult mammalian olfactory bulb throughout the lifetime of an organism is believed to result from the presence of this unique glial cell in the olfactory system. This theory has been substantiated by research demonstrating the ability of transplanted ensheathing cells to promote axonal regrowth in areas of the central nervous system that are normally nonpermissive. A simple method for purifying ensheathing cells resulting in a large yield of cells is therefore invaluable for transplantation studies. We have developed such a method based on the differing rates of attachment of the various harvested cell types. The greatest percentage of cells (70.4%) that attached during the first step of the separation was determined to be fibroblasts. The remainder of the cells were classified as astrocytes (20.8%) and ensheathing cells (6.8%). The percentage of attached astrocytes (67.6%) was greatly increased during the second purification step while the percentage of fibroblasts decreased greatly (27.9%) and the percentage of ensheathing cells (5.3%) slightly decreased. In the final cultures, 93.2 % of the attached cells were ensheathing cells, while astrocytes (5.9%) and fibroblasts (1.4%) were only minor components. This simple, inexpensive method of purifying ensheathing cells will facilitate their use in central nervous system regeneration research.

This study examined the fate of axon terminals of one of the major sources of hypoglossal afferents, the spinal V nucleus, after XIIth nerve resection in adult Sprague-Dawley rats. In order to anterogradely label trigemino-hypoglossal projections, small quantities of horse radish peroxidase were pressure-injected into the ipsilateral dorsal (mandibular) portion of the spinal V nucleus two days before the animals were killed. Survival periods ranged from 5 to 33 days after nerve injury (dpo). Axonal injury produced relative changes in the association of labelled axon terminals to structures in the hypoglossal nucleus on the injured side. The proportion of horse radish peroxidase-labelled spinal V nucleus terminals with spherical vesicles (S-terminals) that were unapposed to hypoglossal somata or dendrites increased rapidly and reached maximal levels by 11 dpo. By contrast, the isolation of labelled terminals with pleomorphic/flattened vesicles (P/F-terminals) from postsynaptic structures began later, advanced at a slower rate and did not attain maximal levels until 20 dpo. S-terminals not apposed to neuronal cell parts increased at a rate of 2.2 times greater than unapposed P/F-terminals. In addition, at peak levels, the proportion of labelled S-terminals that were detached from somata and dendrites was significantly greater than unapposed, labelled P/F-terminals. Axotomy did not alter the caliber of the labelled axon terminals. However, by 29 days after axotomy, the average diameter of dendrites remaining in contact with SPVN terminals was 1/3 the diameter of dendrites of uninjured neurons apposed to labelled axon terminals. These findings provide the morphological correlate for physiological and pharmacological evidence that the effectiveness of excitatory and inhibitory synapses are down-regulated in a coordinated manner after hypoglossal nerve injury.

This study examined changes in choline acetyltransferase and calcitonin gene-related peptide immunoreactivity in hypoglossal motoneurons of rats at 1, 3, 7, 20 and 50 days after three types of nerve injury: crush, transection and resection. Peripheral reinnervation was assayed by retrograde labelling of the motoneurons after injections of the exogenous protein, horseradish peroxidase, into the tongue. Maximal reduction in choline acetyltransferase immunostaining occurred at seven days after nerve damage and the amount of the decrease was related to the nature of the injury. The recovery of choline acetyltransferase to normal levels was related to the timing of reinnervation after nerve crush, but not after transection or resection injuries. In contrast to these findings, a rapid increase in calcitonin gene-related peptide immunoreactivity preceded the decrease in choline acetyltransferase levels. A striking increase in calcitonin gene-related peptide immunoreactivity was observed at one day postoperative and was maximal at three days postoperatively for all injuries. Later changes in calcitonin gene-related peptide levels were dependent on the type of injury. Increased calcitonin gene-related peptide staining persisted to 20 days after nerve crush. After nerve transection or resection, calcitonin gene-related peptide immunoreactivity decreased to basal levels at seven days postoperatively. This declination was followed by a second rise in calcitonin gene-related peptide immunolabeling at 20 days for nerve transection or at 50 days after resection. Nearly complete reinnervation was established by 20 days after nerve crush. At 50 days after transection, less than half the number of normally-labelled neurons contained horseradish peroxidase. At this time only 1% of those whose axons had been resected were labelled. These observations suggest that different mechanisms regulate the responses of choline acetyltransferase and calcitonin gene-related peptide to nerve injury. The present results indicate that choline acetyltransferase levels in motoneurons can not be used to predict either the likelihood of or the timing of reinnervation after nerve transection or resection. However, our results strengthen the premise that an increase of calcitonin gene-related peptide immunoreactivity serves as a reliable index for predicting nerve regeneration/reinnervation after cranial nerve injury.

The pathogenesis of superficial siderosis of the central nervous system (CNS) may be examined by the repeated intracisternal injection of washed autologous red blood cells (RBC). In rabbits, the injections cause the accumulation of iron in the cytoplasm of microglial cells and astrocytes of cerebellar and cerebral cortices. Immunocytochemistry for ferritin reveals enhanced reaction product mainly in microglia but hemosiderin occurs only after extending the injections to 6 months. In an effort to determine the biochemical correlates of these morphological changes, iron, ferritin, ferritin subunits and the ferritin repressor protein (FRP) were quantitated. There was no increase of total iron or ferritin in the exposed cortical areas. However, the injections of RBC caused dramatic shifts of the relative contributions by heavy (H-) and light (L-) ferritin subunits. The initial response was a prompt increase of the H/L ratio to over 4.0 from the normal ratio near 1.0. Extended injections caused the ratio to drop to below unity, and the predominance of L-ferritin at 6 months coincided with the appearance of granular hemosiderin. This investigation also confirmed the presence of FRP in rabbit brain cytosols but the induction of experimental superficial siderosis did not change its levels or in vitro affinity for the iron-responsive element in ferritin messenger ribonucleic acid. It is proposed that the incrustation by hemosiderin which characterizes superficial siderosis of the CNS in humans occurs when prolonged exposure to hemoglobin produces persistent shifts of the H/L-ratios by accumulation of L-ferritin.

Autologous washed red blood cells were injected weekly over a period of three to six months into the cisterna magna of adult New Zealand white rabbits. After three months, the surface of the brain stem, cerebellum, and piriform cortex showed a distinct brown color, and staining of the gross specimens for iron produced an intense blue color which extended for a distance of 1-2 mm into the brain parenchyma. Enhanced iron stains of vibratome sections revealed the accumulation of reaction product in microglia and Bergmann glia of the cerebellar cortex, and in microglia and astrocytes of the piriform cortex. Ferritin immunocytochemistry revealed reaction product in cerebellar microglia and Bergmann glia which strongly resembled that obtained by the enhanced iron stain. In the piriform cortex, only microglia were reactive with anti-ferritin. Electron microscopy confirmed the accumulation of electron-dense ferritin granules only in the cytoplasm of microglia. Bergmann glia in the cerebellum and astrocytic processes in the piriform cortex were replete with intermediate filaments and contained an excess of glycogen. After six months, small granules of hemosiderin began to appear in cerebellar and piriform cortices. The observations support that the sequence of conversion of hemoglobin to ferritin and hemosiderin occurs in brain as in other organs.

Injections of horseradish peroxidase (HRP) or wheat-germ agglutinin-horseradish peroxidase (WGA-HRP) into the nucleus reticularis parvocellularis (RPc) produced anterograde labeling of axon terminals within the hypoglossal nucleus. Based on morphological parameters of vesicle population, membrane specializations, and postsynaptic articulations, two types of axon terminals derived from neurons in RPc end on hypoglossal neurons. More than half of the terminals contained spherical vesicles (S-type), established asymmetrical membrane specializations and contacted proximal and medium-sized dendrites. The remaining labeled terminals had flattened vesicles (F-type), symmetrical membrane densities and apposed medium and small dendrites. The morphological differences expressed in the two types of terminals may reflect physiological and/or pharmacological differences in the action of RPc neurons on motoneurons in the hypoglossal nucleus.

Comparisons of anterograde labeling of axon terminals originating from short and long projection neurons were made in the hypoglossal nucleus. Injections of dilute and concentrated horseradish peroxidase (HRP) or wheat germ-agglutinin-horseradish peroxidase (WGA-HRP) were made via a glass micropipette into the nucleus reticularis parvocellularis (RPc = short projection neurons) and the Spinal V trigeminal complex (Sp. V=long projection neurons). Axon terminals in the hypoglossal nucleus, a common projection site of the two efferent systems, were evaluated ultrastructurally using diaminobenzidine (DAB) as the chromogen for the cobalt-glucose oxidase (CO-GOD) method of HRP labeling. Labeled axon terminals from these two sources demonstrated different distribution patterns of the reaction product. For the short pathway, high concentrations of the tracers resulted in diffuse, agranular labeling in the majority of axon terminals. Dilute concentrations of the tracers were associated with membrane-bound, granular type of labeling. All anterograde labeling of terminals of long projection neurons (Sp. V) was membrane-bound and granular irrespective of the tracer concentration. The length of the pathway and the concentration of the enzyme tracers are factors that affect the pattern of anterograde label in axon terminals of hypoglossal afferents.

Axon terminals projecting to the hypoglossal nucleus have been identified and characterized by electron microscopy following injections of horseradish peroxidase (HRP) into pars interpolaris of the spinal trigeminal nucleus (SPVN) in adult rats. Over 70% of the anterogradely labeled terminals contained spherical vesicles (S-terminals) and their synaptic densities were chiefly asymmetrical (Gray Type I). The rest (28%) of the labeled terminals had flattened vesicles (F-terminals) and predominantly established symmetrical (Gray Type II) synaptic contacts. The diameters of labeled terminals were 0.5-2.5 micron. Two-thirds of the S-terminals had diameters less than 1.25 micron, whereas, F-terminals were distributed equally in the higher (greater than 1.25) and lower (less than 1.25) diameter ranges. Most axon terminals ended on dendrites of hypoglossal neurons; some, chiefly F-terminals, formed axosomatic endings. Dendrites had diameters of 0.5-5 micron. The majority of S- and F-terminals ended on dendrites with diameters of less than 2.5 micron. However, more F-terminals (17%) than S-terminals (11%) were presynaptic to dendrites greater than 2.5 micron in diameter. Experiments in which anterograde HRP labeling of trigemino-hypoglossal projections was combined with retrograde WGA-HRP labeling of motoneurons projecting to the tongue, demonstrated that SPVN axons end on dendrites of these motoneurons. Whether some of the trigeminal fibers also terminate on intrinsic hypoglossal interneurons remains to be determined.

The origin of afferent connections of the hypoglossal nucleus in rats was investigated using horseradish peroxidase (HRP) as a retrograde tracer. Pressure injections (0.15–0.17 μl) of 15% HRP were introduced into the rostral, middle and caudal portions of the nucleus. Projections to the hypoglossal nucleus originated from 3 regions of the brainstem: the reticular formation, the spinal V complex and the nucleus of the solitary tract. Bilateral projections with ipsilateral predominance came from the lateral reticular formation: the dorsal aspect of the nucleus reticularis parvocellularis and its caudal continuation, the nucleus reticularis dorsalis. Fewer projections emerged from two nuclei of the medial reticular formation. The dorsal part of the nucleus reticularis ventralis at the spinomedullary junction contributed bilateral with mainly contralateral input to hypoglossal neurons. A few labeled neurons were situated bilaterally in the nucleus reticularis gigantocellularis of the rostral medulla. The input from the spinal V complex originated from the dorsal aspect along most of its length but particularly from the pars interpolaris and oralis subdivisions. Labeled neurons were located primarily in the posterior portion of the nucleus of the solitary tract. Projections from the spinal V complex and the solitary nucleus exhibited ipsilateral predominance. These results suggest that somatic and visceral centers of the rat brainstem play an important role in the control of the activity of hypoglossal motoneurons.

Axon terminals were identified and characterized by electron microscopy after injections of horseradish peroxidase (HRP) into the spinal V nucleus (SPVN) or the medullary reticular formation adjacent to the XIIth nucleus. The synaptic organization and topology of these two different populations of hypoglossal afferents (T-XII and R-XII respectively) were determined by quantitative comparisons. Significant differences were obtained in the ratios of morphological types of terminals, sizes of axonal endings and their location on postsynaptic structures. Axon terminals containing spherical vesicles (S-terminals) and those with flattened/pleomorphic vesicles (F-terminals) were anterogradely labeled with HRP from both injection sites. However, the S/F ratio for R-XII terminals was 1.2:1 compared to 2.6:1 for T-XII afferents. Asymmetrical membrane densities (Gray Type I) were the predominant form of junctional specialization for S-terminal synapses. Asymmetrical densities with subjunctional dense bodies/bars (S-Taxi) were associated with a higher proportion of T-XII synapses than R-XII synapses. Almost all of the F-terminals from both sources had symmetrical densities (Gray Type II). The average diameter of R-XII terminals was greater than that of T-XII terminals. R-XII-F terminals were the largest terminals. The majority of axon terminals from both sources formed axodendritic synapses. However, R-XII terminals had a higher incidence (10% vs. 3%) of axosomatic contacts. The proportion of R-XII-F-terminals decreased from the central toward the distal dendrites, whereas the opposite was found for T-XII-F and T-XII-S-terminals. In contrast to these findings, R-XII-S-terminals were more uniformly distributed on dendrites of all sizes.(ABSTRACT TRUNCATED AT 250 WORDS)

The intrasomatic reactions to different types of peripheral nerve injury during postnatal maturation were investigated by light and electron microscopy. The hypoglossal nerve was crushed in 7 day postnatal (dpn) rats and crushed, ligated or transected in 10 and 21 day rats. Survival intervals ranged from 3 to 40 days postoperative (dpo). Normal and sham operated rats of corresponding ages served as controls. The initial intrasomatic reactions in young (7-10 dpn) rats were identical after each type of nerve injury. These reactions involved the nucleus and the perinuclear cytoplasm: severe nuclear eccentricity and elaborate infoldings of the nuclear membrane were seen. The processes of cytoplasm indenting the nuclear membrane were intensely basophilic and contained numerous polyribosomes and cisterns of rough endoplasmic reticulum (RER). The formation of organized RER was not disrupted after axonal injury. Disorganization, fragmentation and degranulation of the cisterns were not apparent until 13-20 dpo. Comparable nerve injuries to older (21 dpn) rats produced structural alterations of the same organelles. However, the initial intrasomatic response involved the organized RER and the extent of the changes was directly related to the severity of nerve injury. Nuclear changes occurred later and only after nerve ligation and transection. Therefore, two major differences characterized the intrasomatic reactions to axonal injury in young and older motoneurons. The timetable of involvement of two organelles, the nucleus and the organized RER, was reversed in the sequence of intrasomatic reactions after axonal damage during successive periods of postnatal development. The magnitude of intrasomatic reactions to different types of nerve injury was age-dependent.

Progressive changes in the postnatal incidence, distribution and duration of glycogen in neurons of the pons, medulla and spinal cord were studied by light and electron microscopy using cytochemical and quantitative methods. Albino rats of 11 ages ranging from new-born to adult were used for this investigation. Methacrylate sections, stained with periodic acid Schiff-dimedone (PAS) were surveyed to identify nerve cell groups containing the polysaccharide, glycogen. The PAS reaction was positive in neuronal cell groups of the hypoglossal nucleus, the mesencephalic nucleus of V, nucleus ambiguus, the abducens nucleus, the facial motor nucleus and anterior horn cells of the spinal cord. The intensity and duration of the PAS reaction appeared greatest in the hypoglossal nucleus. Neurons of the mesencephalic nucleus of V demonstrated a reaction of moderate intensity and duration. The remaining nerve cell groups exhibited a weak, diffuse reaction of brief duration. Postnatal differences in the incidence and patterns of disposition of glycogen were quantified using ultrathin sections of the hypoglossal nucleus, the site richest in glycogen. The presence of glycogen was verified by the periodic acid-thiosemicarbizide-silver proteinate (PA-TSC-SP) ultracytochemical stain. The incidence of glycogen in neuronal perikarya of hypoglossal nuclei was related to age. All neurons contained some glycogen during the first postnatal week. By 24 days postnatal (dpn), the majority of hypoglossal neurons lacked glycogen and all neurons of adult rats were glycogen-free. Likewise, the distribution of glycogen particles varied with age. At birth, two perikaryal zones served as the primary sites for glycogen localization: the juxtanuclear and marginal regions. Postnatally, changes in the concentration and patterns of glycogen deposits look place principally during the first postnatal week: large accumulations of free glycogen particles were replaced by a small number of particles, randomly dispersed in the perikaryon. To date, glycogen storage in the developing rat brain has been localized almost exclusively in radial glial cells. The present findings provide the initial ultrastructural demonstration of the temporary storage in and subsequent depletion of glycogen from neurons in the brainstem and spinal cord of early postnatal rats.

A detailed morphometric evaluation of the somata and organelles of regenerating hypoglossal motoneurons from the rat was conducted. The volume of the hypoglossal nucleus and various parameters used to appraise neuronal size were estimated from 50 microns sections. The subcellular composition of randomly selected neurons was quantified from 1 micron and ultrathin sections. The volume of neuronal nuclei, nucleoli, mitochondria and lysosomes as well as the surface area of intracellular membranes were determined. Seven to 30 days following axotomy the volume of the hypoglossal nucleus was significantly diminished, undoubtedly reflecting dendritic retraction (P less than 0.05). Concomitantly, all estimates of neuronal size indicated significant neuronal enlargement (P less than 0.05). Ultrastructural alterations were most prominent 7 days following nerve transection: nucleolar volume was significantly increased, rough endoplasmic reticulum surface area was reduced, and non-Golgi smooth membrane surface area increased (P less than 0.05). In general, other organelles resisted the influence of axotomy and all ultrastructural parameters returned to control levels 21 to 30 days following the nerve transection. Functional recovery was detected in all animals 21 and 30 days following axotomy. The measured responses of axotomized hypoglossal motoneurons are similar to those reported for retinal ganglion cells of the goldfish (WhitnallGrafstein, 1982, 1983), suggesting common metabolic events among these distinct neuronal populations following axonal transection.

Glial fibrillary acidic protein (GFAP) immunoreactivity within rat hypoglossal (XIIth) nuclei was examined 1–50 days following either unilateral nerve transection or modest electrical stimulation using indirect immunofluorescence and PAP immunohistochemistry. Both nerve transection and stimulation provoked an increase in the immunodetected GFAP within the XIIth nucleus.