Changes in Two Point Discrimination and the law of mobility in Diabetes Mellitus patients

Background

Though retrograde neuronal death and vascular insufficiency have been well established in plegics following intracerebral hemorrhage, the effects of plegia on arterial nervorums of peripheral nerves have not been reported. In this study, the histopathological effects of the intracerebral hemorrhage on the dorsal root ganglions and sciatic nerves via affecting the arterial nervorums were investigated.

Methods

This study was conducted on 13 male hybrid rabbits. Three animals were taken as control group and did not undergo surgery. The remaining 10 subjects were anesthetized and were injected with 0.50 ml of autologous blood into their right sensory-motor region. All rabbits were followed-up for two months and then sacrificed. Endothelial cell numbers and volume values were estimated a three dimensionally created standardized arterial nervorums model of lumbar 3. Neuron numbers of dorsal root ganglions, and axon numbers in the lumbar 3 nerve root and volume values of arterial nervorums were examined histopathologically. The results were analyzed by using a Mann-Whitney-U test.

Results

Left hemiplegia developed in 8 animals. On the hemiplegic side, degenerative vascular changes and volume reduction in the arterial nervorums of the sciatic nerves, neuronal injury in the dorsal root ganglions, and axonal injury in the lumbar 3 were detected. Statistical analyses showed a significant correlation between the normal or nonplegic sides and plegic sides in terms of the neurodegeneration in the dorsal root ganglions (p < 0.005), axonal degeneration in the lumbar 3 nerve roots (p < 0.005), endothelial cell degeneration in the arterial nervorums (p < 0.001), and volume reduction in the arterial nervorums (p < 0.001).

Conclusion

Intracerebral hemorrhage resulted in neurodegeneration in the dorsal root ganglion and axonolysis in the sciatic nerves, endothelial injury, and volume reduction of the arterial nervorums in the sciatic nerves. The interruption of the neural network connection in the walls of the arterial nervorums in the sciatic nerves may be responsible for circulation disorders of the arterial nervorums, and arterial nervorums degeneration could result in sciatic nerves injury.     

End-Structure of Afferent Axons Injured in the Peripheral and Central Nervous System

The end-structure of afferent axons chronically severed in the rat sciatic nerve or dorsal column (DC) was visualized by centrifugal transport of horseradish peroxidase (HRP) or wheatgerm agglutinin conjugated to HRP (WGA:HRP) injected into the L₄ or L₅ dorsal root ganglion. Nerve regeneration was prevented and neuroma formation encouraged by tightly ligating the cut nerve end. For the first few weeks postoperative, the time during which afferents trapped in a nerve-end neuroma generate their most intense ectopic impulse barrage, the developing neuroma was dominated by swollen terminal end-bulbs. There was some axonal dying-back, retrograde fiber growth, and terminal sprouting, but little preterminal branching. The rich tangle of fine preterminal branches usually thought of in relation to nerve-end neuromas did not elaborate until several months postoperative, a time when the neuroma is relatively quiescent electrically. Afferents cut in the DC, which never develop dramatic ectopic electrical activity, showed morphological peculiarities similar to nerve-end neuromas during the early postoperative period, including retrograde fiber growth and minimal sprouting. They did not, however, go on to form luxuriant branches. These data provide preliminary clues as to the structure of the ectopic impulse-generating mechanism thought to underlie paresthesias and pain associated with peripheral nerve injury.     

Deletion of p75NTR impairs regeneration of peripheral nerves in mice

AimsAfter peripheral nerve injury, p75NTR was upregulated in Schwann cells of the Wallerian degenerative nerves and in motor neurons but down-regulated in the injured sensory neurons. As p75NTR in neurons mediates signals of both neurotrophins and inhibitory factors, it is regarded as a therapeutic target for the treatment of neurodegeneration. However, its physiological function in the nerve regeneration is not fully understood. In the present study, we aimed to examine the role of p75NTR in the regeneration of peripheral nerves.Main methodsIn p75NTR knockout mice (exon III deletion), the sciatic nerves and facial nerves on one side were crushed and regenerating neurons in the facial nuclei and in the dorsal root ganglia were labelled by Fast Blue. The regenerating fibres in the sciatic nerve were also labelled by an anterograde tracer and by immunohistochemistry.Key findingsThe results showed that the axonal growth of injured axons in the sciatic nerve of p75NTR mutant mice was significantly retarded. The number of regenerated neurons in the dorsal root ganglia and in the facial nuclei in p75NTR mutant mice was significantly reduced. Immunohistochemical staining of regenerating axons also showed the reduction in nerve regeneration in p75NTR mutant mice.SignificanceOur data suggest that p75NTR plays an important role in the regeneration of injured peripheral nerves.     

Characterization of proteins transported at different rates by axoplasmic flow in the dorsal root afferents of rats

Proteins synthesized by soma located in L₄ dorsal root ganglia and supplied to the axonal branches extending centrally in the dorsal root and peripherally towards the sciatic nerve were analyzed for radioactivity following injections of [³H] leucine into the L₄ dorsal root ganglia. All proteins located in the dorsal root and sciatic nerve were analyzed by SDS acrylamide gel electrophoresis at various times post injection. The differences in radioactivity between the dorsal root and sciatic nerve proteins were mainly quantitative and not qualitative, with many proteins of various molecular weight ranges being transported into both segments. Generally, it appears that in both axonal branches the high molecular weight proteins are transported at the highest rate, medium weights slower and low molecular weight proteins slowest. More proteins of high and low molecular weights are transported into the dorsal root whereas more of those of medium molecular weight are transported towards the sciatic nerve.     

Soluble Nogo Receptor Down-regulates Expression of Neuronal Nogo-A to Enhance Axonal Regeneration

Nogo-A, a member of the reticulon family, is present in neurons and oligodendrocytes. Nogo-A in central nervous system (CNS) myelin prevents axonal regeneration through interaction with Nogo receptor 1, but the function of Nogo-A in neurons is less known. We found that after axonal injury, Nogo-A is increased in dorsal root ganglion (DRG) neurons unable to regenerate following a dorsal root injury or a sciatic nerve ligation-cut injury and that exposure in vitro to CNS myelin dramatically enhanced neuronal Nogo-A mRNA and protein through activation of RhoA while inhibiting neurite growth. Knocking down neuronal Nogo-A by small interfering RNA results in a marked increase of neurite outgrowth. We constructed a nonreplicating herpes simplex virus vector (QHNgSR) to express a truncated soluble fragment of Nogo receptor 1 (NgSR). NgSR released from QHNgSR prevented myelin inhibition of neurite extension by hippocampal and DRG neurons in vitro. NgSR prevents RhoA activation by myelin and decreases neuronal Nogo-A. Subcutaneous inoculation of QHNgSR to transduce DRG neurons resulted in improved regeneration of myelinated fibers in both the dorsal root and the spinal dorsal root entry zone, with concomitant improvement in sensory behavior. The results indicate that neuronal Nogo-A is an important intermediate in neurite growth dynamics and its expression is regulated by signals related to axonal injury and regeneration, that CNS myelin appears to activate signaling events that mimic axonal injury, and that NgSR released from QHNgSR may be used to improve recovery after injury.     

Electro-acupuncture Reduces Neuronal Apoptosis Linked to Bax and Bcl-2 Expression in the Spinal Cords of Cats Subjected to Partial Dorsal Root Ganglionectomy

While electro-acupuncture (EA) has been well known to contribute towards neuroplasticity occurring in both the central and the peripheral nervous system after injury, the underlying mechanism remains largely unknown. This study evaluated the effects and the possible mechanism of EA on neuronal apoptosis in the spinal cords of cats subjected to the removal of L₁–L₅ and L₇–S₂ dorsal root ganglion, sparing the L₆ dorsal root ganglion. EA treatment decreased the number of TUNEL-positive apoptotic cells in lamina II of the L₃ and L₆ cord segments at 7 and 14 days post operation (dpo). This EA-mediated neuroprotection is associated with a decrease in the number of Bax immunoreactive neurons and an increase in the number of Bcl-2 immunoreactive neurons. Furthermore, Western blot and RT-PCR analysis revealed a significant downregulation of Bax protein and its mRNA, but an upregulation of Bcl-2 in the dorsal horn of L₃ and L₆ cords at both 7 and 14 dpo. The present findings suggest that EA could inhibit neuronal apoptosis in dorsal root deafferentated cat spinal cords, possibly by Bax downregulation and Bcl-2 upregulation. Wei Zhao and Qi Zhao contributed equally to this work.     

Direct activation of Transient Receptor Potential Vanilloid 1(TRPV1) by Diacylglycerol (DAG)

Voltage-gated sodium channels play important roles in modulating dorsal root ganglion (DRG) neuron hyperexcitability and hyperalgesia after peripheral nerve injury or inflammation. We report that chronic compression of DRG (CCD) produces profound effect on tetrodotoxin-resistant (TTX-R) and tetrodotoxin-sensitive (TTX-S) sodium currents, which are different from that by chronic constriction injury (CCI) of the sciatic nerve in small DRG neurons. Whole cell patch-clamp recordings were obtained in vitro from L₄ and/or L₅ dissociated, small DRG neurons following in vivo DRG compression or nerve injury. The small DRG neurons were classified into slow and fast subtype neurons based on expression of the slow-inactivating TTX-R and fast-inactivating TTX-S Na⁺ currents. CCD treatment significantly reduced TTX-R and TTX-S current densities in the slow and fast neurons, but CCI selectively reduced the TTX-R and TTX-S current densities in the slow neurons. Changes in half-maximal potential (V_1/2) and curve slope (k) of steady-state inactivation of Na⁺ currents were different in the slow and fast neurons after CCD and CCI treatment. The window current of TTX-R and TTX-S currents in fast neurons were enlarged by CCD and CCI, while only that of TTX-S currents in slow neurons was increased by CCI. The decay rate of TTX-S and both TTX-R and TTX-S currents in fast neurons were reduced by CCD and CCI, respectively. These findings provide a possible sodium channel mechanism underlying CCD-induced DRG neuron hyperexcitability and hyperalgesia and demonstrate a differential effect in the Na⁺ currents of small DRG neurons after somata compression and peripheral nerve injury. This study also points to a complexity of hyperexcitability mechanisms contributing to CCD and CCI hyperexcitability in small DRG neurons.     

Axoplasmic importins enable retrograde injury signaling in lesioned nerve

Axoplasmic proteins containing nuclear localization signals (NLS) signal retrogradely by an unknown mechanism in injured nerve. Here we demonstrate that the importin/karyopherin alpha and beta families underlie this process. We show that importins are found in axons at significant distances from the cell body and that importin beta protein is increased after nerve lesion by local translation of axonal mRNA. This leads to formation of a high-affinity NLS binding complex that traffics retrogradely with the motor protein dynein. Trituration of synthetic NLS peptide at the injury site of axotomized dorsal root ganglion (DRG) neurons delays their regenerative outgrowth, and NLS introduction to sciatic nerve concomitantly with a crush injury suppresses the conditioning lesion induced transition from arborizing to elongating growth in L4/L5 DRG neurons. These data suggest a model whereby lesion-induced upregulation of axonal importin beta may enable retrograde transport of signals that modulate the regeneration of injured neurons.     

A conditioning lesion promotes in vivo nerve regeneration in the contralateral sciatic nerve of rats

A conditioning lesion in the sciatic nerve increases in vivo axonal regeneration in the nerve after a second transection. We studied whether this increased regeneration also occurs in the contralateral nerve. The left sciatic nerve was transected and sutured in Wistar rats; the nerve was exposed but not transected in controls. After 5 days, the right sciatic nerves of all rats were transected and sutured. Neuronal regeneration was measured at 0, 1, 3, 5, and 7 days with the pinch test and histological staining. IL-1beta and TGF-beta1 expression was also measured. The initial delay in the experimental group was significantly shorter, but the regeneration rates were the same. The expression of IL-1beta and TGF-beta1 in the right dorsal root ganglia was significantly higher in the experimental group. Nerve injury enhances cytokine expression in the contralateral dorsal root ganglion and promotes contralateral nerve regeneration in vivo by shortening the initial delay.     

The Plasticity of the DRG Neurons Belonging to Different Subpopulations After Dorsal Rhizotomy

SUMMARY 1. The plasticity of sensory neurons following the injury to their axons is very important for prognosis of recovery of afferent fibers with different modality. It is evident that the response of dorsal root ganglion (DRG) neurons after peripheral axotomy is different depending on the deficiency in neurotrophic factors from peripheral region. The loss of cells appears earlier and is more severe in B-cells (small, dark cells with unmyelinated axons) than in A-cells (large, light cells with myelinated axons).2. We studied using immunohistochemical methods the response of DRG neurons to dorsal rhizotomy and combined injury of central and peripheral neuronal processes. A quantitative analysis of DRG neurons tagged by the selective markers isolectin B4 (IB4) and the heavy molecular component of the neurofilament triplet (NF200) antibody, selective for subpopulations of small and large/medium DRG neurons, respectively, was performed after dorsal rhizotomy, peripheral axotomy, and their combination.3. The number of NF200⁺-neurons is reduced substantially after both dorsal rhizotomy and peripheral axotomy, while the decrease of IB4⁺-neurons is observed only in combined injury, i.e., dorsal rhizotomy accompanied with sciatic nerve injury.4. Our results show that distinct subpopulations of DRG neurons respond differently to the injury of their central processes. The number of NF200⁺-neurons decreases to greater degree following dorsal rhizotomy in comparison to IB4⁺-neurons.     

Peripheral nerve lesion-induced uptake and transport of choleragenoid by capsaicin-sensitive c-fibre spinal ganglion neurons

In the present experiments the effect of systemic capsaicin treatment on the retrograde labelling of sensory ganglion cells was studied following the injection of choleratoxin B subunit-horseradish peroxidase conjugate (CTX-HRP) into intact and chronically transected peripheral nerves. In the control rats CTX-HRP injected into intact sciatic nerves labelled medium and large neurons with a mean cross-sectional area of 1,041 +/- 39 gm2. However, after injection of the conjugate into chronically transected sciatic nerves of the control rats, many small cells were also labelled, shifting the mean cross-sectional area of the labelled cells to 632 +/- 118 microm2. Capsaicin pretreatment per se induced a moderate but significant decrease in the mean cross-sectional area of the labelled neurons (879 +/- 79 microm2). More importantly, systemic pretreatment with capsaicin prevented the peripheral nerve lesion-induced labelling of small cells. Thus, the mean cross-sectional areas of labelled neurons relating to the intact and transected sciatic nerves, respectively, did not differ significantly. These findings provide direct evidence for a phenotypic switch of capsaicin-sensitive nociceptive neurons after peripheral nerve injury, and suggest that lesion-induced morphological changes in the spinal cord may be related to specific alterations in the chemistry of C-fibre afferent neurons rather than to a sprouting response of A-fibre afferents.     

Impaired regenerative response of primary sensory neurons in ZPK/DLK gene-trap mice

Rapid and persistent activation of c-JUN is necessary for axonal regeneration after nerve injury, although upstream molecular events leading to c-JUN activation remain largely unknown. ZPK/DLK/MAP3K12 activates the c-Jun N-terminal kinase pathway at an apical level. We investigated axonal regeneration of the dorsal root ganglion (DRG) neurons of homozygous ZPK/DLK gene-trap mice. In vitro neurite extension assays using DRG explants from 14 day-old mice revealed that neurite growth rates of the ZPK/DLK gene-trap DRG explants were reduced compared to those of the wild-type DRG explants. Three ZPK/DLK gene-trap mice which survived into adulthood were subjected to sciatic nerve axotomy. At 24 h after axotomy, phosphorylated c-JUN-positive DRG neurons were significantly less frequent in ZPK/DLK gene-trap mice than in wild-type mice. These results indicate that ZPK/DLK is involved in regenerative responses of mammalian DRG neurons to axonal injury through activation of c-JUN.     

Slow transport of the cytoskeleton after axonal injury

The delivery of cytoskeletal proteins to the axon occurs by slow axonal transport. We examined how the rate of slow transport was altered after axonal injury. When retinal ganglion cell (RGC) axons regenerated through peripheral nerve grafts, an increase in the rate of slow transport occurred during regrowth of the injured axons. We compared these results to axonal injury in the optic nerve where no substantial regrowth occurs and found a completely different response. Slow transport was decreased approximately tenfold in rate in the proximal segment of crushed optic nerves. This decreased rate of slow transport was not induced immediately, but occurred about 1 week after injury. To explore whether a decrease in the rate of slow transport was induced when the regeneration of peripheral nerves was physically blocked, we examined slow transport in motor neurons after the sciatic nerve was transected and ligated. In this case, no change in the rate of the comigrating tubulin and neurofilament (NF) radioactive peaks were observed. We discuss how the changes in the rate of slow transport may reflect different neuronal responses to injury and speculate about the possible molecular changes in the expression of tubulin which may contribute to the observed changes. © 1992 John Wiley & Sons, Inc.     

RSK1 promotes mammalian axon regeneration by inducing the synthesis of regeneration-related proteins

In contrast to the adult mammalian central nervous system (CNS), the neurons in the peripheral nervous system (PNS) can regenerate their axons. However, the underlying mechanism dictating the regeneration program after PNS injuries remains poorly understood. Combining chemical inhibitor screening with gain- and loss-of-function analyses, we identified p90 ribosomal S6 kinase 1 (RSK1) as a crucial regulator of axon regeneration in dorsal root ganglion (DRG) neurons after sciatic nerve injury (SNI). Mechanistically, RSK1 was found to preferentially regulate the synthesis of regeneration-related proteins using ribosomal profiling. Interestingly, RSK1 expression was up-regulated in injured DRG neurons, but not retinal ganglion cells (RGCs). Additionally, RSK1 overexpression enhanced phosphatase and tensin homolog (PTEN) deletion-induced axon regeneration in RGCs in the adult CNS. Our findings reveal a critical mechanism in inducing protein synthesis that promotes axon regeneration and further suggest RSK1 as a possible therapeutic target for neuronal injury repair.

This study shows that p90 ribosomal S6 kinase 1 (RSK1) responds differentially to nerve injury in the peripheral and central nervous systems, and identifies it as a crucial regulator of axonal regeneration; mechanistically, RSK1 preferentially induces the synthesis of regeneration-related proteins via the RSK1-eEF2K-eEF2 axis.     

Involvement of gicerin, a cell adhesion molecule, in development and regeneration of chick sciatic nerve

We have examined the role of gicerin, an immunoglobulin superfamily cell adhesion molecule, in chick sciatic nerves during development and regeneration. Gicerin was expressed in the spinal cord, dorsal root ganglion (DRG) and sciatic nerves in embryos, but declined after hatching. Neurite extensions from explant cultures of the DRG were promoted on gicerin's ligands, which were inhibited by an anti-gicerin antibody. Furthermore, gicerin expression was upregulated in the regenerating sciatic nerves, DRG and dorsal horn of the spinal cord after injury to the sciatic nerve. These results indicate that gicerin might participate in the development and regeneration of sciatic nerves.     

Slow transport of the cytoskeleton after axonal injury.

The delivery of cytoskeletal proteins to the axon occurs by slow axonal transport. We examined how the rate of slow transport was altered after axonal injury. When retinal ganglion cell (RGC) axons regenerated through peripheral nerve grafts, an increase in the rate of slow transport occurred during regrowth of the injured axons. We compared these results to axonal injury in the optic nerve where no substantial regrowth occurs and found a completely different response. Slow transport was decreased approximately tenfold in rate in the proximal segment of crushed optic nerves. This decreased rate of slow transport was not induced immediately, but occurred about 1 week after injury. To explore whether a decrease in the rate of slow transport was induced when the regeneration of peripheral nerves was physically blocked, we examined slow transport in motor neurons after the sciatic nerve was transected and ligated. In this case, no change in the rate of the comigrating tubulin and neurofilament (NF) radioactive peaks were observed. We discuss how the changes in the rate of slow transport may reflect different neuronal responses to injury and speculate about the possible molecular changes in the expression of tubulin which may contribute to the observed changes.     

The Characterization of AT<Subscript>1</Subscript> Expression in the Dorsal Root Ganglia After Chronic Constriction Injury

To clarify the role of Angiotensin II in the regulation of sensory signaling, we characterized the AT₁ expression in neuronal subpopulation of lower lumbar dorsal root ganglia under normal conditions and its alteration in neuropathic pain model. The characterization of AT₁ expression was done under control and after the chronic constriction injury induced by four loose ligatures of the sciatic nerve representing the model of posttraumatic painful peripheral neuropathy. Major Angiotensin II receptor type was expressed in approximately 43 % of small-sized and 62 % of large-sized neurons in control. The AT₁ overexpression after sciatic nerve ligation lasting 7 days was detected predominantly in small-sized AT₁ immunoreactive neurons (about 38 % increase). Chronic constriction injury caused a statistically marked increase in number of the small-sized peptidergic (CGRP immunoreactive) neuronal subpopulation expressing AT₁ (about 64 %). The subpopulations of AT₁-immunoreactive and nonpeptide-containing primary sensory neurons revealed by IB4 binding, tyrosine hydroxylase- and parvalbumin-immunoreactive neurons were not markedly changed. Our results indicate that: (1) the AT₁ overexpression after the chronic constriction injury is an important factor in Angiotensin II-potentiated pain perception; (2) Angiotensin II is involved in pathological mechanisms of neuropathic pain and this effect can be mediated perhaps in combination with other neuropeptides synthesized in the primary sensory neurons.     

Time-related changes in the labeling pattern of motor and sensory neurons innervating the gastrocnemius muscle,as revealed by the retrograde transport of the cholera toxin B subunit

Summary Morphological changes in the motor and sensory neurons in the lumbar spinal cord and the dorsal root ganglia were investigated at different survival times following the injection of the B subunit of cholera toxin (CTB) into the medial gastrocnemius muscle. Unconjugated CTB, visualized immunohistochemically, was found to be retrogradely transported through ventral and dorsal roots to motor neurons in the anterior horn, each lamina in the posterior horn, and ganglion cells in the dorsal root ganglia at L₃–L₆. The largest numbers of labeled motor neurons and ganglion cells were observed 72 h after the injection of CTB. Thereafter, labeled ganglion cells were significantly decreased in number, whereas the amount of labeled motor neurons showed a slight reduction. Motor neurons had extensive dendritic trees filled with CTB, reaching lamina VII and even the pia mater of the lateral funiculus. Labeling was also seen in the posterior horn, but the central and medial parts of laminae II and III had the most extensively labeled varicose fibers, the origin of which was the dorsal root ganglion cells. The results indicate that CTB is taken up by nerve terminals and can serve as a sensitive retrogradely transported marker for identifying neurons that innervate a specific muscle.     

Injury-associated induction of GAP-43 expression displays axon branch specificity in rat dorsal root ganglion neurons

Peripheral nerve injury results in the increased synthesis and axonal trasnport of the growth-associated protein GAP-43 in dorsal root ganglion (DRG) neurons, coincident with regenerative growth of the injured peripheral axon branches. To determine wheter the injury-associated signalling mechanism which leads to GAP-43 induction also operates through the central branches of DRG axons, we used immunocytochemistry to compare the expression of GAP-43 in adult rat DRG neurons 2 weeks after dorsal root crush lesions (central axotomy) or peripheral nerve crush lesions (peripheral axotomy). In uninjured ganglia, a subpopulation of smaller DRG neurons expresses moderate levels of GAP-43, whereas larger neurons generally do not. At 2 weeks following peripheral axotomy, virtually all axotomized neurons, large and small, express high levels of GAP-43. At 2 weeks following dorsal root lesions, no increase in GAP-43 expression is detected. Thus, the injury-associated up-regulation of GAP-43 expression in DRG neurons is triggered by a mechanism that is responsive to injury of only the peripheral, and not the central, axon branches. These findings support the hypothesis that GAP-43 induction in DRG neurons is caused by disconnection from peripheral target tissue, not by axon injury per se. © 1993 John Wiley & Sons, Inc.