Axonal Conduction & Degeneration In Multiple Sclerosis

A noninactivating, persistent Sodium current has been demonstrated previously in Dorsal Root Ganglia Neurons and in rat Optic Nerve.

We report here that Na⁺ channel blockade with TetrodoToxin (TTX) in isolated Dorsal and Ventral Roots elicits membrane HyperPolarization, suggesting the presence of a persistent Na⁺ current in Peripheral Axons.

We used a modified Sucrose-gap chamber to monitor resting and Action Potentials and observed a HyperPolarizing shift in the Nerve potential of rat Dorsal and Ventral roots by TTX.

The block of transient inward Na⁺ currents was confirmed by the abolition of Compound Action Potentials (CAPs).

Moreover, Depolarization of Nerve Roots by elevating ExtraCellular K⁺ concentrations to 40 mM eliminated CAPs but did not significantly alter TTX-induced HyperPolarizations, indicating that the persistent Na⁺ currents in Nerve Roots are not Voltage-dependent.

TetrodoToxin-sensitive persistent inward Na⁺ currents are present in both Dorsal and Ventral root Axons at rest and may contribute to Axonal excitability.

Axotomy of Nerve Fibers leads to the subsequent degeneration of their distal part, a process termed Wallerian Degeneration (WD).

While WD in the Peripheral Nervous System is usually followed by regeneration of the lesioned Axons, Central Nervous System (CNS) Neurons are generally unable to regrow.

In this study, we investigated the process of WD in the Dorsal Columns of the rat Spinal Cord rostral to a mid-Thoracic lesion.

We confirm earlier studies describing a very delayed Microglial and an early and sustained Astroglial reaction finally leading to scar formation. Interestingly, we found a differential time course in the loss of Myelin proteins depending on their location.

Proteins situated on the PeriAxonal Myelin membrane such as Myelin Associated Glycoprotein disappeared early, within a few days after lesion, concomitantly with CytoSkeletal Axonal proteins.

Whereas, compact Myelin and outer Myelin membrane proteins such as MBP and Nogo-A remained for long intervals in the degenerating tracts.

Two distinct mechanisms are probably responsible for this difference: processes of protein destruction emanating from and initially probably located in the Axon, act on a time scale of 1-3 days.

In contrast, the bulk of Myelin destruction is due to Phagocytosis known to be slow, prolonged, and inefficient in the CNS. These results may also have implications for future intervention strategies aiming at enhancing CNS regeneration.

Copyright 2003 Wiley-Liss, Inc.

Multiple Sclerosis (MS) is an AutoImmune Central Nervous System (CNS) DeMyelinating Disease that causes relapsing and chronic Neurologic impairment. Recent observations have altered certain traditional concepts regarding MS pathogenesis.

A greater diversity of cell types and molecules involved in MS is now evident. While ReMyelination can occur during the early inflammatory phase when damage may be reversible.

It is impaired in the later stages, which involve Axonal death. These observations have important therapeutic implications.

The pivotal role of Axons in the PathoPhysiology and PathoGenesis of Multiple Sclerosis (MS) is increasingly becoming the focus of our attention. Axonal injury, considered at one time to be a late phenomenon, is now recognized as an early occurrence in the inflammatory lesions of MS.

There is converging evidence from HistoPathologic, as well as Magnetic Resonance Imaging and Magnetic Resonance Spectroscopy studies, that Axons play a crucial and dynamic role during the evolution of MS pathology and the development of clinical disability.

It has been repeatedly demonstrated that Neurologic functional impairment correlates best with Axonal, rather than Myelin, injury. The PathoPhysiology of Axonal injury remains speculative.

Although generally considered to be sequelae of DeMyelination, it is possible that Axonal injury in MS is indeed a primary event. The discovery that Axonal injury can be reversible has provided an impetus to institute early therapy.

The finding that irreversible Axonal transection occurs in early lesions has underscored now, more than ever before.

The need to curtail inflammation and the need to institute early treatment with disease-modifying agents. The Axon will undoubtedly remain the focus of our attention regarding research on MS now and in the future.

Myelination of Axons by Oligodendrocytes enables rapid Impulse propagation in the Central Nervous System. But long-term interactions between Axons and their Myelin sheaths are poorly understood.

Here we show that Cnp1, which encodes 2',3'-Cyclic Nucleotide Phosphodiesterase in Oligodendrocytes, is essential for Axonal survival but not for Myelin assembly.

In the absence of Glial Cyclic Nucleotide Phosphodiesterase, mice developed Axonal swellings and NeuroDegeneration throughout the Brain, leading to Hydrocephalus and premature death.

But, in contrast to previously studied Myelin mutants, the UltraStructure, periodicity and physical stability of Myelin were not altered in these mice.

Genetically, the chief function of Glia in supporting Axonal integrity can thus be completely uncoupled from its function in maintaining compact Myelin.

Oligodendrocyte dysfunction, such as that in Multiple Sclerosis lesions, may suffice to cause secondary Axonal loss.

All previous analyzes of Axonal responses to Traumatic Axonal Injury (TAI) have described the UltraStructure of changes in the CytoSkeleton and Axolemma within 6 h of injury.

In the present study we tested the hypothesis that there are, in addition, UltraStructural pathological changes up to 1 week after injury. TAI was induced in the adult guinea pig Optic Nerve of nine animals.

Three animals were killed at either 4 h, 24 h, or 7 days (d) after injury. Quantitative analysis of the number or proportion of Axons within 0.5-micro m-wide bins.

Showed an increase in the number of Axons with a diameter of less than 0.5 micro m at 4 h, 24 h, and 7 d, the presence of lucent Axons at 24 h and 7 d and that the highest number of injured Axons occurred about half way along the length of the Nerve.

A spectrum of pathological changes occurred in injured Fibers-pathology of Mitochondria:

1. Dissociation of Myelin lamellae but little damage to the Axon
2. Loss of linear register of the Axonal CytoSkeleton
3. Differential responses between MicroTubules (MT) and NeuroFilaments (NF) in different sizes of Axon
4. Two different sites of compaction of NF
   • Loss of both NF (with an increase in their spacing)
5. And MT (with a reduction in their spacing)
6. Replacement of the Axoplasm by a flocculent precipitate
7. Increased length of the Nodal gap

These provide the first UltraStructural evidence for Wallerian Degeneration of Nerve Fibers in an animal model of TAI.

Axonal degeneration has been identified as the major determinant of irreversible Neurological disability in patients with Multiple Sclerosis (MS).

Axonal injury begins at disease onset and correlates with the degree of inflammation within lesions, indicating that inflammatory DeMyelination influences Axon Pathology during Relapsing/Remitting MS (RR-MS).

This Axonal Loss remains clinically silent for many years, and irreversible Neurological disability develops when a threshold of Axonal Loss is reached and compensatory CNS resources are exhausted.

Experimental support for this view - the Axonal Hypothesis - is provided by data from various animal models with primary Myelin or Axonal pathology, and from pathological or Magnetic Resonance studies on MS patients.

In mice with Experimental Autoimmune Encephalomyelitis (EAE), 15-30% of Spinal Cord Axons can be lost before permanent Ambulatory Impairment occurs.

During Secondary/Progressive MS (SP-MS), chronically DeMyelinated Axons may degenerate due to lack of Myelin-derived Trophic support.

In addition, we hypothesize that reduced Trophic support from damaged targets or degeneration of Efferent Fibers may trigger preprogrammed NeuroDegenerative mechanisms.

The concept of MS as an Inflammatory NeuroDegenerative Disease has important clinical implications regarding therapeutic approaches, monitoring of patients, and the development of NeuroProtective treatment strategies.

Axonal injury in Multiple Sclerosis has attracted considerable interest during the past few years. It has been demonstrated in association with inflammation within active lesions, but it is also present in Normal-Appearing White Matter.

Because Axonal Loss appears to be responsible for persistent Neurological deficits in patients with Multiple Sclerosis, treatment strategies to prevent damage to Neurites and restore function are of paramount importance in controlling the disease process.

Some of the currently available ImmunoModulatory therapies may also reduce Axonal damage, as demonstrated using improved imaging technologies, but the precise mechanisms that could protect Axons during the inflammatory attack are yet to be identified.

Factors that are involved in functional impairment of Axonal Conduction and those elements that are responsible for direct structural damage to the Axon are both potential targets for therapeutic interventions.

Spinal Cord regeneration in adult mammals is limited by Neurite outgrowth inhibitors and insufficient availability of outgrowth-promoting agents.

Formation of degenerative swellings at the proximal ends of severed Axons (Terminal Clubs), which starts early after injury, also may hinder recovery and their rupture may contribute to secondary Spinal Cord damage.

We investigated whether NeuroTrophins would reduce these degenerative processes. Adult rats received a transection of the Dorsal Column Sensory and CorticoSpinal Motor Tracts at T9 and anterograde tracing of the Axons from the Sciatic Nerve and Motor Cortex, respectively.

The highest number of terminal clubs was found at 1 day and approximately half remained present until at least 28 days.

A single injection immediately after injury of a mixture of Nerve Growth Factor, Brain-Derived Neurotrophic Factor and NeuroTrophin-3 into the lesion site, reduced the number of Terminal Clubs in the Sensory System by approximately half at 1 and 7 days (but not 14) after the lesion.

Individual or combinations of two NeuroTrophins were as effective, suggesting that the NeuroTrophins protected similar Axonal populations. The injected NeuroTrophins did not affect degeneration of CorticoSpinal Motor Axons.

A 7-day continuous intrathecal infusion of NeuroTrophin-3 was more effective and also reduced Terminal Club formation of CorticoSpinal Axons by approximately 60%.

Spinal tissue loss was not affected by the NeuroTrophin treatments, suggesting that Terminal Clubs are not major contributors to the pathogenesis of secondary Spinal degeneration during the first two weeks.

Thus, NeuroTrophins can reduce Axonal degeneration in the Spinal Cord after Traumatic Axonal Injury.

Copyright 2002 Elsevier Science (USA).

Axonal degeneration contributes to clinical disability in the acquired DeMyelinating Disease Multiple Sclerosis.

Axonal degeneration occurs during acute attacks, associated with inflammation, and during the chronic progressive phase of the disease in which inflammation is not prominent.

To explore the importance of interactions between Oligodendrocytes and Axons in the CNS, we analyzed the Brains of rodents and humans with a null mutation in the gene encoding the major CNS Myelin Protein, ProteoLipid Protein (PLP1, previously PLP).

Histological analyses of the CNS of Plp1 null mice and of autopsy material from patients with null PLP1 mutations were performed to evaluate Axonal and Myelin integrity.

In vivo Proton Magnetic Resonance Spectroscopy (MRS) of PLP1 null patients was conducted to measure levels of N-AcetylAspartate (NAA), a marker of Axonal integrity.

Length-dependent Axonal degeneration without DeMyelination was identified in the CNS of Plp1 null mice. Proton MRS of PLP1-deficient patients showed reduced NAA levels, consistent with Axonal loss.

Analysis of patients' Brain tissue also demonstrated a length-dependent pattern of Axonal loss without significant DeMyelination. Therefore, Axonal degeneration occurs in humans as well as mice lacking the major Myelin protein PLP1.

This degeneration is length-dependent, similar to that found in the PNS of patients with the inherited DeMyelinating Neuropathy, CMT1A, but is not associated with significant DeMyelination.

Disruption of PLP1-mediated Axonal - Glial interactions thus probably causes this Axonal degeneration. A similar mechanism may be responsible for Axonal degeneration and clinical disability that occur in patients with Multiple Sclerosis.

In this paper we report a qualitative morphological analysis of Wallerian Degeneration in a marsupial.

Right Optic Nerves of opossums Didelphis marsupialis were crushed with a fine forceps and after 24, 48, 72, 96 and 168 hours the animals were anaesthetized and perfused with fixative.

The Optic Nerves were immersed in fixative and processed for routine transmission electron microscopy.

Among the early alterations typical of Axonal degeneration, we observed nerve fibers with focal degeneration of the Axoplasmic CytoSkeleton, watery degeneration and dark degeneration, the latter being prevalent at 168 hours after crush.

Our results point to a gradual disintegration of the AxoPlasmic CytoSkeleton, opposed to the previous view of an "all-or-nothing" process (Griffin et al 1995).

We also report that, due to an unknown mechanism, fibers show either a dark or watery pattern of Axonal degeneration, as observed in Axon profiles. We also observed fibers undergoing early Myelin breakdown in the absence of Axonal alterations.

Peripheral Neuropathies and Wallerian Degeneration share a number of pathological features; the most prominent of which is Axonal degeneration.

We asked whether common PathoPhysiologic mechanisms are involved in these 2 disorders by directly comparing in vitro models of Axonal degeneration after Axotomy or exposure to the NeuroToxin Vincristine.

Embryonic rat Dorsal Root Ganglia (DRG) were allowed to extend Neurites for 5 days in culture, and then were either Axotomized or exposed to 0.01 microM Vincristine. Neurites universally degenerated by 3 days after Axotomy or after 6 days of Vincristine exposure.

The NeuroProtective effects of a low Calcium environment or pharmacologic inhibition of the Cysteine protease Calpain were compared in these 2 models of Axonal degeneration.

Addition of EGTA or growth in zero-Calcium media provided significant protection against Axonal degeneration after either Axotomy or Vincristine exposure.

Treatment with the experimental Calpain Inhibitor AK295 was equally protective in both models. Chronic exposure to AK295 was not toxic to the cultures.

These data suggest that common mechanisms involving Calcium and Calpains are involved in both Axotomy-induced and Vincristine-induced Axonal degeneration.

In addition, Calpain inhibition may provide a strategy for preventing Axonal degeneration and preserving Neurologic function in a variety of PNS and CNS Disorders.