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PubMed2005

Gonadal steroid attenuation of developing hamster facial motoneuron loss by axotomy: equal efficacy of testosterone, dihydrotestosterone, and 17-beta estradiol.

In the hamster facial nerve injury paradigm, we have established that androgens enhance both functional recovery from facial nerve paralysis and the rate of regeneration in the adult, through intrinsic effects on the nerve cell body response to injury and via an androgen receptor (AR)-mediated mechanism. Whether these therapeutic effects of gonadal steroids encompass neuroprotection from axotomy-induced cell death is the focus of the present study. Virtually 100% of adult hamster facial motoneurons (FMNs) survive axotomy at the stylomastoid foramen (SMF), whereas, before postnatal day 15 (P15), developing FMNs undergo substantial axotomy-induced cell death. The first part of the present study focuses on determining when ARs are first expressed in developing hamster FMNs. Using AR immunocytochemistry, it was found that males express ARs by P2 and females by P4, which is the earliest demonstration of AR expression in mammalian motoneurons reported thus far in the literature. The second half examines the neuroprotective effects of testosterone propionate, 17-beta estradiol, and dihydrotestosterone on FMNs of P7 hamsters after facial nerve transection at the SMF. The results demonstrate that androgens and estrogens are equally able to rescue approximately 20% of FMNs from axotomy-induced cell death, with the effects permanent. This study is the first to investigate the effects of both androgens and estrogens on axotomy-induced cell death in one system and, with our previously published work, to validate the hamster FMN injury paradigm as a model of choice in the investigation of both neurotherapeutic and neuroprotective actions of gonadal steroids.

Age Factors↗
PubMed2000

Role of p38 mitogen-activated protein kinase in axotomy-induced apoptosis of rat retinal ganglion cells.

p38 is a member of the mitogen-activated protein (MAP) kinase superfamily and mediates intracellular signal transduction. Recent studies suggest that p38 is involved in apoptotic signaling in several cell types, including neurons. In the mammalian retina, approximately 50% of the retinal ganglion cells (RGCs) die by apoptosis during development. Additionally, transection of the optic nerve close to the eye bulb causes apoptotic cell death of RGCs in adulthood. We investigated the role of p38 in axotomy-induced apoptosis of RGCs. One day after axotomy, activated (phosphorylated) p38 was visualized by immunocytochemistry in the nuclei of RGCs, but not in control retinas. Phosphorylated p38 was first detected on immunoblots 12 hr after axotomy, reached a maximum at 1 d, and then decreased. To investigate possible roles of p38 in RGC death, a p38 MAP kinase inhibitor, SB203580, was administered intravitreally at the time of axotomy and repeated at 5 and 10 d. Assayed 14 d after axotomy, SB203580 increased the number of surviving RGCs in a dose-dependent manner (the minimum effective concentration was 1.6 micrometer). Furthermore, MK801, a selective inhibitor of NMDA receptors, not only showed protective effects against RGC apoptosis but also attenuated p38 MAP kinase activation in a dose-dependent manner. Our findings imply that p38 is in the signaling pathway to RGC apoptosis mediated by glutamate neurotoxicity through NMDA receptors after damage to the optic nerve. p38 inhibitors could be potentially useful for the treatment of optic nerve trauma and neurodegenerative diseases that affect RGCs, such as glaucoma.

Animals↗
PubMed1998

Upregulation of growth associated protein 43 expression and neuronal co-expression with neuropeptide Y following inferior alveolar nerve axotomy in the rat.

Growth associated protein 43 (GAP 43) is an acidic membrane-bound phosphoprotein produced at high levels in developing and regenerating neurons. It is a substrate for protein kinase C and suggested to be involved in calcium-regulated release of axonal vesicular-contained neurotransmitters. Expression of GAP 43 has been demonstrated in the uninjured cat dental pulp which receives its sensory nerve supply from the trigeminal ganglion. The aim of this study was a detailed mapping of the spatial and time-dependent expression of GAP 43 and co-expression of neuropeptide Y (NPY) in dental peripheral target tissues and trigeminal neurons subsequent to inferior alveolar nerve (IAN) axotomy in rats, as background for later low-level laser studies. Unilateral sectioning of IAN, resulting in an almost complete loss of sensory nerve fibers in the ipsilateral dental pulp of the first molar, was performed. The avidin biotin complex (ABC) method was used to evaluate peripheral changes in GAP 43 expression at 4, 7 and 10 days. Ganglionic changes in GAP 43 and co-localization of neuronal NPY expression was examined at 4, 10 and 21 days using either the ABC method or double immunofluorescence labelling techniques and confocal microscopy. Axotomy resulted in an early upregulation and change in the peripheral distribution of GAP 43 in nerve profiles already 4 days post IAN axotomy suggesting a Schwann cell origin. Ten days post axotomy a pronounced upregulation of GAP 43 immunoreactivity could be demonstrated in neurons located in the mandibular region of the trigeminal ganglion, compared to the contralateral uninjured side. The peripheral and ganglionic upregulation of GAP 43 continued to persist at 21 days. A concomitant time-delayed shift and co-expression of NPY was demonstrated throughout in the GAP 43-upregulated ganglion cells 10 days post axotomy. Furthermore, confocal microscopy indicated that the intraneuronal distribution of NPY and upregulated GAP 43 expression showed a similar conformity and distribution in both perinuclear regions and cell periphery.

Animals↗
PubMed1997

Effect of permanent axotomy on number and volume of dorsal root ganglion cell bodies.

The effects of axotomy on the size and number of rat dorsal root ganglion cells was studied using stereological methods. Twenty adult Wistar rats were axotomized by transection of the right fifth lumbar spinal nerve approximately 7 mm distal to the fifth lumbar dorsal root ganglion (DRG-L5). The corresponding ganglia from the nonaxotomized side served as controls. The DRG-L5 were removed for study 4, 8, 15, and 45 days after axotomy. The number of neurons in each DRG-L5 was determined from estimates of the numerical density, NV, made with disectors and estimates of the volume of the ganglion using the Cavalieri principle. The mean cell body volume was determined with the vertical planar rotator method. There was a progressive loss of nerve cells during the postoperative period. There was a loss of 6% (not significant) after 4 days, 19% (not significant) after 8 days, 22% (2P < 0.05) after 15 days, and 35% (2P < 0.005) after 45 days. The relative reduction in cell number 45 days after axotomy was larger for B-cells (43%) than for A-cells (15%). The mean nerve cell body volume for the entire DRG-L5 cell population was reduced by 33% (2P < 0.005) 4 days after axotomy and remained so throughout the experimental period. The distribution of the individual cell volumes in the ganglia appeared to be uniformly shifted to lower values. It is concluded that permanent axotomy of the fifth lumbar spinal nerve results in a substantial loss of dorsal root ganglion cells and is well-suited as a model for studying the potential protective effects of neurotrophic factors using modern stereological techniques.

Animals↗
PubMed1999

Activation of protein kinase A contributes to the expression but not the induction of long-term hyperexcitability caused by axotomy of Aplysia sensory neurons.

Nociceptive sensory neurons (SNs) in Aplysia provide useful models to study both memory and adaptive responses to nerve injury. Induction of long-term memory in many species, including Aplysia, is thought to depend on activation of cAMP-dependent protein kinase (PKA). Because Aplysia SNs display similar alterations in models of memory and after nerve injury, a plausible hypothesis is that axotomy triggers memory-like modifications by activating PKA in damaged axons. The present study disproves this hypothesis. SN axotomy was produced by (1) dissociation of somata from the ganglion [which is shown to induce long-term hyperexcitability (LTH)], (2) transection of neurites of dissociated SNs growing in vitro, or (3) peripheral nerve crush. Application of the competitive PKA inhibitor Rp-8-CPT-cAMPS at the time of axotomy failed to alter the induction of LTH by each form of axotomy, although the inhibitor antagonized hyperexcitability produced by 5-HT application. Strong activation of PKA in the nerve by coapplication of a membrane-permeant analog of cAMP and a phosphodiesterase inhibitor was not sufficient to induce LTH of either the SN somata or axons. Furthermore, nerve crush failed to activate axonal PKA or stimulate its retrograde transport. Therefore, PKA activation plays little if any role in the induction of LTH by axotomy. However, the expression of LTH was reduced by intracellular injection of the highly specific PKA inhibitor PKI several days after nerve crush. This suggests that long-lasting activation of PKA in or near the soma contributes to the maintenance of long-term modifications produced by nerve injury.

Animals↗
PubMed1990

Ultrastructure of an identified molluscan neuron in organ culture and cell culture following axotomy.

We examined the ultrastructure of neuron 5 from the buccal ganglion of the mollusc Helisoma trivolvis after axotomy and organ culture, and after isolation of the same neuron in culture. Buccal ganglia containing axotomized neurons 5 were cultured either in host snails or in Leibovitz medium conditioned with ganglia. In addition, some neurons 5 were isolated from buccal ganglia by micro-dissection and plated into culture. Neuron 5 and its processes were identified in both whole mounts and plastic sections of buccal ganglia after intracellular injection with Lucifer Yellow or horseradish peroxidase. Five days after axotomy of neuron 5, thick sections of buccal ganglia stained with toluidine blue revealed that densely staining basophilic bodies (Nissl bodies) within the cytoplasm had dispersed, i.e., they had undergone chromatolysis. Coincident with chromatolysis was an overall increase in diffuse basophilic staining within the cytoplasm of neuron 5 when maintained in organ culture. The dispersion of Nissl bodies viewed by light microscopy correlated with a more freely arranged rough endoplasmic reticulum and associated polysomes within neuron 5 as seen by electron microscopy. Isolated neurons 5 did not possess densely staining Nissl bodies when examined after 2 days in vitro, thus indicating that chromatolysis occurred earlier in isolated neurons. Furthermore, no increase in diffuse cytoplasmic basophilia was observed within isolated neurons 5 cultured in vitro. However, isolated neurons 5 exhibited a marked increase in the number of lipid-like bodies (0.5-1.5 micron in diameter) that were particularly evident in scanning electron micrographs. Scanning and transmission electron micrographs revealed that the isolated neurons were free of associated glia, but non-neuronal cells (hemocytes) would attach themselves to the somata and neurites. Glia surrounding neuron 5 within buccal ganglia exhibited a marked hypertrophy following axotomy and organ culture. Hypertrophy of glia was absent, however, if ganglia were axotomized and left within the animal or axotomized ganglia were implanted into host animals and examined 5 days later by electron microscopy. These observations indicate that, following axotomy, a molluscan neuron may exhibit different morphological features depending on its microenvironment. In addition, the hypertrophy of glia surrounding neurons in Helisoma was not associated with axotomy per se, but with organ culture.

Animals↗
PubMed1994

Axotomy induces preprotachykinin gene expression in a subpopulation of dorsal root ganglion neurons.

The distribution of dorsal root ganglion (DRG) cell sizes that show changes in preprotachykinin (PPT) gene expression and substance P (SP) levels following axotomy was examined using RNA blot analysis, in situ hybridization histochemistry, and immunocytochemistry. PPT mRNA was induced in medium-sized (1,000-2,000 microns 2) and large-sized (> 2,000 microns 2) cells in the DRG after axotomy. There was a 165% increase in the number of labeled cells after sciatic transection and a 260% increase after spinal nerve transection which results in axotomy of all the cells in the ganglion. The further increase after spinal nerve transection suggests that the induction occurred in axotomized neurons. PPT mRNA label was also present in a reduced number of small (< 1,000 microns 2) cells after axotomy. SP immunoreactivity was also induced in medium- and large-sized cells and reduced in small-sized cells. Our findings suggest that the expression of the PPT gene and SP is differentially regulated in different subpopulations of DRG neurons after axotomy and is consistent with the hypothesis that tachykinins may be important in both sensory transmission and regeneration.

Animals↗
PubMed1991

Axotomy accelerates slow component b of axonal transport.

Because the integrity of an axon depends on the supply of proteins synthesized in the cell body, we examined the effect of axotomy on the transport of structural proteins in rat motor axons, and the effect of altered transport on the rate of outgrowth after a subsequent testing axotomy. To examine the axonal transport of structural proteins, we labeled newly synthesized proteins with 35S-methionine 7 days after a "conditioning" lesion of the sciatic nerve, and removed the nerve 7-21 days later for SDS-PAGE. Tubulin, actin, calmodulin, and the 68-kD light neurofilament protein (NF-L) were identified by fluorography and removed for liquid scintillation counting. The fastest moving structural proteins were carried by slow component b (SCb) of axonal transport, which advanced 20% faster in conditioned axons: 4.2 versus 3.5 mm/day (p less than 0.01). NF-L was not accelerated, indicating that the motor for subcomponent a (SCa) of slow axonal transport was unaffected by axotomy. To measure outgrowth distances, the testing lesions was made 7 days after the conditioning lesion, and growth cones were located by the fast transport method 3 or 9 days later. The regression analysis of outgrowth distance on time showed that sprouts elongated 25% faster in conditioned axons: 4.0 versus 3.2 mm/day (p less than 0.001). These accelerated sprouts were formed too far from the spinal cord to contain SCb proteins that were synthesized after axotomy. Because the rate of outgrowth correlated closely with the rate of SCb in outgrowing sprouts (McQuarrie and Jacob, J. Comp. Neurol. 305:139-147, 1991), we conclude that SCb is accelerated throughout the length of the axon by 7 days after axotomy.

Actins↗
PubMed1994

Regulation of immediate-early gene expression in rat retinal ganglion cells after axotomy and during regeneration through a peripheral nerve graft.

To determine mechanisms of structural plasticity in adult CNS neurons, we investigated the expression of immediate early genes (IEGs) in the rat retina. Gene products of different IEG families (JUN and FOS proteins) and cAMP-responsive element binding protein (CREBP) were examined by immunohistochemistry under three different paradigms. Normal rats which were not axotomized were compared with axotomized animals, were retinal ganglion cells (RGCs) were axotomized by intraorbital optic nerve cut and retrogradely labeled with fluorogold (FG). Under these circumstances, RGCs show only transient sprouting, followed by continuous retrograde RGC degeneration. In the third group, after the optic nerve lesion, adult rats additionally received a sciatic nerve graft to the transected optic nerve stump. This allows some RGCs to regenerate an axon into the grafted nerve. In both groups, the time course of RGC survival and JUN, CREB, and FOS protein expression was monitored. In normal animals, JUN-Immunoreactivity (JUN-Ir) was not detectable in the retinal ganglion cell layer. JUN-Ir was induced in about 70% of all FG-positive RGCs 5 days after axotomy. The expression of JUN-Ir stated to decline 8 days after axotomy. Only a few JUN-Ir-positive RGCs were found after 2 weeks. In transplanted animals, however, the numbers of JUN-Ir-positive RGCs were significantly higher 2 and 3 weeks after transplantation compared to animals that exclusively received axotomy. Furthermore, in grafted rats, about 70% of the regenerating RGCs expressed JUN-Ir 2 weeks after grafting as compared to only 38% JUN-positive RGCs among the surviving but not regenerating RGCs. In normal animals CREBP-Ir was constitutively expressed in nearly all cells of the retinal ganglion cell layer. The decline in number of CREBP-Ir-positive cells paralleled the axotomy-induced RGC death. FOS-Ir-positive cells were not found in the ganglion cell layer at any time. These results demonstrate a selective and transient JUN expression of RGCs after axotomy which is sustained during axonal regeneration. This suggests that sciatic nerve grafts are able to regulate the expression of JUN proteins in axotomized RGCs of adult rats.

Activating Transcription Factor 2↗
PubMed1995

Importance of target innervation in recovery from axotomy-induced loss of androgen receptor in rat perineal motoneurons.

In adult male rats, axotomy of the spinal nucleus of the bulbocavernosus (SNB) motoneurons transiently down-regulates androgen receptor (AR) immunoreactivity. The present study investigates the importance of target reinnervation in the recovery of AR expression in axotomized SNB motoneurons after short (up to 5 days) and long (1 to 6 weeks) periods of recovery. In the long-term recovery experiment, animals were divided into two groups. In one, the two stumps of the cut pudendal nerve, which carries the axons of the SNB motoneurons, were sutured together immediately after axotomy. In the second group, the proximal stump was ligated immediately after axotomy to prevent target reinnervation. Axotomy of the SNB motoneurons caused a significant down-regulation in AR immunoreactivity within 3 days. At 6 weeks, AR immunoreactivity was still depressed in ligated animals but had recovered to control levels in resutured animals. The recovery in the resutured group was coincident with the first signs of reinnervation of the target perineal muscles, although reinnervation seemed to lag behind AR immunoreactivity. SNB soma size was significantly reduced 2 weeks after axotomy and returned to control levels after 6 weeks of recovery only in the resutured animals. These findings suggest that the target perineal muscles play a role in the regulation of AR expression and androgen sensitivity in the SNB motoneurons, perhaps mediated by muscle-derived trophic factors.

Analysis of Variance↗
PubMed1995

Agrin gene expression in ciliary ganglion neurons following preganglionic denervation and postganglionic axotomy.

Agrin is an extracellular matrix protein that has been implicated as a synaptogenic agent in the peripheral and central nervous systems. Both the level of expression and pattern of alternative splicing of agrin mRNA are developmentally regulated. As a step toward identifying signals important in regulating agrin gene expression in neurons, we examined the effects of postganglionic axotomy or preganglionic denervation on agrin mRNA levels and alternative splicing in ciliary ganglia of posthatch chicks. In comparison to unoperated age-matched controls, in situ hybridization with a pan-specific agrin cRNA probe demonstrated a significant decrease in neuronal agrin mRNA expression as a result of axotomy. Reverse transcription-polymerase chain reaction analysis demonstrated that axotomy also resulted in changes in the pattern of alternative splicing of agrin mRNA. Underlying these changes are decreases in the molar amounts of transcripts encoding the neuron-specific isoforms agrin8 and agrin19, homologous to rat agrin proteins that have high AChR aggregating activity. Similar, but less dramatic changes in agrin expression following axotomy were also observed in unoperated neurons on the contralateral side. In contrast, the only significant change in agrin gene expression following ganglionic denervation was a small decline in the relative abundance of agrin 8 mRNA in operated versus unoperated age-matched control ganglia. Major changes in agrin gene expression following axotomy but not denervation are consistant with the notion that agrin synthesized by ganglionic neurons exerts its effects in the periphery rather than at synapses formed between ciliary ganglion neurons and their preganglionic input. These data suggest that the pattern of alternative splicing and the absolute amount of agrin mRNA in ciliary ganglion neurons may be regulated by target tissue interactions.

Agrin↗
PubMed1997

Expression of superoxide dismutase following axotomy.

Oxidative injury has been implicated in the pathophysiology of neuronal injury and neurodegenerative disease. Antioxidant proteins provide an endogenous defense against such oxidative injury and may yield important clues to mechanisms of cytoprotection and neuronal recovery. Axotomy is the simplest model of neuronal injury and lesioning the sciatic nerve allows concurrent study of both motor (spinal cord) and sensory (dorsal root ganglia, DRG) neurons affected by the same injury. This study was designed to examine the expression of superoxide dismutase (SOD), an essential antioxidant protein, in motor and sensory neurons following complete axotomy of peripheral nerve. Immunocytochemical, quantitative immunoblot, and enzymatic activity assay techniques are used. By 12 days after axotomy, immunocytochemical expression of Mn-SOD is markedly increased in affected DRG and spinal cord. A similar increase in Cu/Zn-SOD is not seen in DRG or spinal cord. This immunocytochemical staining is associated with a significant increase in specific activity and Mn-SOD protein content as measured on quantitative immunoblots. This report suggests, for the first time, that Mn-SOD and not Cu/Zn-SOD increases in sensory neurons of the DRG and motor neurons of the spinal cord following distal axotomy of the sciatic nerve. Quantitative measurements of Mn-SOD following axotomy reveals that the increase in immunocytochemical reactivity is associated with an approximately 30% increase in specific activity when comparing lesioned and contralateral spinal cord samples. These data suggest that Mn-SOD may have a more significant role in the pathophysiology of neuronal injury than Cu/Zn-SOD.

Animals↗
PubMed1976

Morphometric analysis of rat superior cervical ganglion after axotomy and nerve growth factor treatment.

The changes in neuronal number, cell body size and nuclear size have been followed for 12 weeks after postganglionic axotomy of the rat superior cervical ganglion. Axotomy was carried out at 6 days post partum and treatment with nerve growth factor (NGF) was from 6-21 days. During normal development there is a 30% decrease in the number of neurons in the superior cervical ganglion; axotomy increases the loss of cells resulting in a 90% decrease by 28 days post partum. The normal decrease is prevented and the enhanced loss of cells after axotomy is decreased by administration of NGF. Thus the increased number of cells observed after NGF administration appears to be due to the survival of cells that otherwise would have degenerated. NGF causes a rapid enlargement of both the cell bodies and the nucleus in the normal and axotomized ganglia. This increase in size rapidly reverses after cessation of treatment. These changes in cell size may account for the previously observed changes in cell profile number with NGF. There is a large increase in the number of non-neuronal cells during normal development and axotomy prevents this increase. NGF treatment results in a 6 fold increase in the number of non-neuronal cells and it is suggested that these are required to support the massive fibre outgrowth that occurs in NGF treated ganglia. It is concluded that these results are consistent with a physiological role for NGF as the trophic substance supporting adrenergic neurons making the appropriate contact with their target cell.

Age Factors↗
PubMed1983

Effect of axotomy on the cyclic GMP increase induced by preganglionic stimulation and high extracellular K+ concentration in superior cervical sympathetic ganglion of the rat.

Cyclic GMP generation, induced by preganglionic nerve stimulation or by high extracellular potassium ion concentration (70 mM) in the medium, was studied during aerobic incubation of the excised superior cervical sympathetic ganglion of the rat with and without axotomy, and the results were compared with that of preganglionic denervation. Both axotomy and denervation of the ganglion for a week caused complete loss of increase of cyclic GMP content in the ganglion in response to the preganglionic nerve stimulation. However, the increase of the ganglionic cyclic GMP content evoked by raising the extracellular potassium ion concentration was maintained at a level about two-thirds of the control after axotomy for 1-4 days, while it was abolished within a day after denervation. Ganglionic choline acetyltransferase activity was maintained for several days after axotomy, but it was decreased rapidly by denervation. Acetylcholinesterase, dopamine-beta-hydroxylase and muscarinic cholinergic receptor sites had been lost almost completely a week after axotomy. These results suggest that the increase of ganglionic cyclic GMP content caused by depolarization with high extracellular K+ is associated with the preganglionic nerve terminals rather than with postganglionic receptor.

Acetylcholinesterase↗
PubMed1984

Cholinesterase activity in the rat superior cervical ganglion: effect of denervation and axotomy.

Cholinesterase activity and its various forms in the superior cervical ganglion of the rat were studied after denervation, axotomy and double section. The response of this activity to various inhibitors was also studied. The specific activity of cholinesterase activity was 323 +/- 25 nmol/mg protein/min in homogenates of normal ganglia and the total enzyme activity was 78 +/- 5 nmol/ganglion/min. Axotomy produced approximately a 40% decrease in total activity in ganglia at 3 days compared to contralateral ganglia. Activity in the contralateral ganglia was also decreased by about 30% compared to normal ganglia. Decentralization alone for 3 days did not affect the specific or total cholinesterase activity. However, decentralization followed by axotomy 3 days later resulted in a greater loss in activity compared to axotomy and a significant change in the forms of cholinesterase. After chromatography on Sepharose 6B-100 columns, 3 forms (a, c, d) of cholinesterase activity could be separated from homogenates of single ganglia. The 3 forms had estimated molecular weights of 1,934,000, 129,000 and 59,000 daltons, respectively. A fourth form (b) presented as a shoulder and had an estimated molecular weight of 404,000 daltons. Decentralization or axotomy decreased forms a and d. A reduction in the proportion of form c was also observed in the axotomized ganglion. Decentralization followed by axomoty resulted in large decreases in forms a, b and c and complete loss of form d. Assay of the separated forms in the presence of the specific inhibitor of acetylcholinesterase, BW284C-51, indicated that the majority of the enzyme recovered after chromatography is acetylcholinesterase.

Acetylcholinesterase↗
PubMed1989

Nerve growth factor (NGF) reverses axotomy-induced decreases in choline acetyltransferase, NGF receptor and size of medial septum cholinergic neurons.

Intraventricular nerve growth factor (NGF) infusion in the adult rat can prevent and also, if delayed, reverse the disappearance of most of the axotomized medial septum cholinergic neurons immunostained for choline acetyltransferase (ChAT). We have utilized the delayed NGF treatment protocol to (i) extend to 3 months the delay time between axotomy and NGF treatment, (ii) define the time course of their recovery, (iii) determine that immunostaining for the (lower affinity) NGF receptor (NGFR) parallels loss and reversal of the ChAT marker, and (iv) evaluate changes in cholinergic somal size following axotomy and subsequent NGF treatment. While NGF treatments starting only 7 days after the fimbria-fornix transection (axotomy) almost entirely restored the number of both ChAT- and NGFR-positive medial septum neurons, longer delayed (2-3 weeks) treatment brought about recovery from the baseline of 20-25% to only about 70% of the control numbers. This limited recoverability, however, persisted even after a 95 day delay period. In all cases examined maximal recoveries were achieved within 3-7 days of NGF treatment. Neuronal size analyses provided evidence for an axotomy-induced atrophy. NGF treatments, started with 1 or 2 week delays, not only reversed fully the average somal size loss but also induced an actual hypertrophy of several of those neurons. These results provide additional evidence that at least half of the apparent loss of cholinergic medial septum neurons upon axotomy is due to a loss of markers such as the transmitter-related enzyme ChAT and NGFR rather than to actual neuronal cell death. These results also show that NGF exerts a genuine trophic influence by regulating the size of its target neurons as well as their content of several proteins.

Animals↗
PubMed1979

Muscarinic receptors in the central nervous system of the rat. IV. A comparison of the effects of axotomy and deafferentation on the binding of [3H]propylbenzilylcholine mustard and associated synaptic changes in the hypoglossal and pontine nuclei.

The reaction of axotomy has been studied in the rat hypoglossal nucleus by quantitative electron microscopical counts of numbers of synapses and by changes in muscarinic receptors assessed by counting silver grains in light microscope autoradiographs of the specific (atropine-sensitive) binding of [3H]propylbenzilylcholine mustard in cryostat sections. For the first 5 days after unilateral peripheral hypoglossal nerve axotomy the muscarinic ligand binding falls to 50% of control levels and then shows no further fall for up to 30 days. Synapse numbers decrease progressively over the first 10 days after operation, by which time they reach 50% of normal. Thus receptor changes reach completion at a time when synapse loss is still continuing. Later, both muscarinic ligand binding and synapse numbers recover to an extent which depends at least in part on the effectiveness of the peripheral nerve regeneration, suggesting that both the receptor and synapse changes may be dependent upon neuromuscular contacts. The reactions of muscarinic receptors to axotomy and deafferentation have been studied in the rat basilar pontine nuclei. Cerebellectomy, which causes axotomy of the pontine neurones and also removes their postsynaptic targets (the granule cells), causes no change in pontine muscarinic receptor over the first week after operation. This differs from the rapid fall in hypoglossal muscarinic receptors induced by axotomy. At longer survivals after cerebellectomy there is a partial loss of pontine muscarinic receptors associated with atrophy of the pontine neurones. Destruction of the neocortical afferents causes a loss of at least half of the synapses in the pontine neuropil. However, the light microscopic autoradiographic study revealed no obvious changes in the dentisy or distribution of the pontine muscarinic receptors from 4 days to more than 6 months after operation.

Afferent Pathways↗
PubMed1991

A quantitative study of neuropeptide immunoreactive cell bodies of primary afferent sensory neurons following rat sciatic nerve peripheral axotomy.

Following peripheral axotomy, fluoride resistant acid phosphatase (FRAP) and most neuropeptides are depleted in the central terminals of axotomised nerves and reduced in their corresponding cell bodies (DRG) but vasoactive intestinal polypeptide (VIP) increases. The increase in VIP probably results from a change in gene expression in other ganglion cells which do not normally express VIP. A quantitative study was performed to investigate the proportion of DRG cells immunoreactive for different peptides at increasing times after sciatic nerve section. Retrograde fluorescent neuronal labelling of sciatic nerve cell bodies by injection of fast blue into the proximal stump was combined with unlabelled antibody immunohistochemistry for CGRP and VIP. The proportion of cells immunoreactive for these peptides was quantified between two and fourteen days post-axotomy. The number of VIP immunoreactive profiles increased significantly in the first 4 days post-axotomy, followed by a slight decrease before rising again. In contrast, the number of and CGRP-immunoreactive cell profiles declined to zero by 14 days post-axotomy. 4 days post-axotomy 50% of VIP positive cells were also immunoreactive for CGRP. There was neither colocalisation between VIP and FRAP nor between CGRP and FRAP. It is concluded that many peptidergic DRG cell bodies switch their expression of peptide to VIP after injury, whereas non-peptide-containing subpopulations do not.

Acid Phosphatase↗