Impaired Axoplasmic Transport Research Articles

Extent of impaired axoplasmic transport and neurofilament compaction in traumatically injured axon at various strains and strain rates

ABSTRACTPrimary objective: Secondary axotomy is more prevalent than the primary axotomy and involves subtle intraaxonal changes in response to the injury leading to cytoskeletal disruptions including neurofilament (NF) misalignment and compaction, which is associated with the genesis of impaired axoplasmic transport (IAT). Recent studies have reported two differential axonal responses to injury, one associated with the cytoskeletal collapse and another with the IAT. The objective of this study was to determine the extent of IAT and early NF changes in axons that were subjected to a stretch of various degrees at different strain rates.Research design and methods: Fifty-six L5 dorsal spinal nerve roots were subjected to a predetermined strain at a specified displacement rate (0.01 and 15 mm/second) only once. The histological changes were determined by performing standard immunohistochemical procedures using beta amyloid precursor protein (β APP) and NF-68 kDa antibodies.Results and conclusions: No significant differences in the occurrence rate of either of the staining in the axons were observed when subjected to similar loading conditions, and the occurrence rate of both β APP and NF68 staining was strain and rate-dependent.

Rescue of tau-induced synaptic transmission pathology by paclitaxel

Behavioral and electrophysiological studies of Alzheimer’s disease (AD) and other tauopathies have revealed that the onset of cognitive decline correlates better with synaptic dysfunctions than with hallmark pathologies such as extracellular amyloid-β plaques, intracellular hyperphosphorylated tau or neuronal loss. Recent experiments have also demonstrated that anti-cancer microtubule (MT)-stabilizing drugs can rescue tau-induced behavioral decline and hallmark neuron pathologies. Nevertheless, the mechanisms underlying tau-induced synaptic dysfunction as well as those involved in the rescue of cognitive decline by MTs-stabilizing drugs remain unclear. Here we began to study these mechanisms using the glutaminergic sensory-motoneuron synapse derived from Aplysia ganglia, electrophysiological methods, the expression of mutant-human tau (mt-htau) either pre or postsynaptically and the antimitotic drug paclitaxel. Expression of mt-htau in the presynaptic neurons led to reduced excitatory postsynaptic potential (EPSP) amplitude generated by rested synapses within 3 days of mt-htau expression, and to deeper levels of homosynaptic depression. mt-htau-induced synaptic weakening correlated with reduced releasable presynaptic vesicle pools as revealed by the induction of asynchronous neurotransmitter release by hypertonic sucrose solution. Paclitaxel totally rescued tau-induced synaptic weakening by maintaining the availability of the presynaptic vesicle stores. Postsynaptic expression of mt-htau did not impair the above described synaptic-transmission parameters for up to 5 days. Along with earlier confocal microscope observations from our laboratory, these findings suggest that tau-induced synaptic dysfunction is the outcome of impaired axoplasmic transport and the ensuing reduction in the releasable presynaptic vesicle stores rather than the direct effects of mt-htau or paclitaxel on the synaptic release mechanisms.

Temporal assessment of traumatic axonal injury in the rat corpus callosum and optic chiasm

Impaired axoplasmic transport (IAT) and neurofilament compaction (NFC), two common axonal pathology processes involved in traumatic axonal injury (TAI), have been well characterized. TAI is found clinically and in animal models in brainstem white matter (WM) tracts and in the corpus callosum (CC), optic chiasm (Och), and internal capsule. Previous published quantitative studies of the time course of TAI expression induced by the Marmarou impact acceleration model have been limited to the brainstem. Accordingly, this study assessed the extent of IAT and NFC in the CC and Och at 8h, 28h, 3days and 7days after traumatic brain injury (TBI) induction by the Marmarou impact acceleration model. IAT peak density was observed at 8h in the CC and 28h in the Och post-TBI. NFC peak density was observed at 28h in both structures. The density of IAT and NFC decreased with increasing survival time in both structures. The NFC density time profile followed a similar trend in both the Och and CC, whereas the IAT density time profile was variable between the Och and CC. Furthermore, a strong linear relationship was observed between IAT and NFC in the CC but not in the Och. These findings highlight the heterogeneity of TAI as evidenced by variable IAT and NFC injury time profiles in each anatomical structure. This variability indicates the requirement of multiple markers for a comprehensive TAI evaluation and multiple targeted treatments for TAI polypathology within its therapeutic window time frame.

Impaired axoplasmic transport is the dominant injury induced by an impact acceleration injury device: An analysis of traumatic axonal injury in pyramidal tract and corpus callosum of rats

Traumatic axonal injury (TAI) involves neurofilament compaction (NFC) and impaired axoplasmic transport (IAT) in distinct populations of axons. Previous quantification studies of TAI focused on limited areas of pyramidal tract (Py) but not its entire length. Quantification of TAI in corpus callosum (CC) and its comparison to that in Py is also lacking. This study assessed and compared the extent of TAI in the entire Py and CC of rats following TBI. TBI was induced by a modified Marmarou impact acceleration device in 31 adult male Sprague Dawley rats by dropping a 450gram impactor from either 1.25m or 2.25m. Twenty-four hours after TBI, TAI was assessed by beta amyloid precursor protein (β-APP-IAT) and RMO14 (NFC) immunocytochemistry. TAI density (β-APP and RMO14 axonal swellings, retraction balls and axonal profiles) was counted from panoramic images of CC and Py. Significantly high TAI was observed in 2.25m impacted rats. β-APP immunoreactive axons were significantly higher in number than RMO14 immunoreactive axons in both the structures. TAI density in Py was significantly higher than in CC. Based on our parallel biomechanical studies, it is inferred that TAI in CC may be related to compressive strains and that in Py may be related to tensile strains. Overall, IAT appears to be the dominant injury type induced by this model and injury in Py predominates that in CC.

The effects of cyclosporin-A on axonal conduction deficits following traumatic brain injury in adult rats

Immunophilin ligands, including cyclosporin-A (CsA), have been shown to be neuroprotective in experimental models of traumatic brain injury (TBI) and to attenuate the severity of traumatic axonal injury. Prior studies have documented CsA treatment to reduce essential components of posttraumatic axonal pathology, including impaired axoplasmic transport, spectrin proteolysis, and axonal swelling. However, the effects of CsA administration on axonal function, following TBI, have not been evaluated. The present study assessed the effects of CsA treatment on compound action potentials (CAPs) evoked in corpus callosum of adult rats following midline fluid percussion injury. Rats received a 20 mg/kg bolus of CsA, or cremaphor vehicle, at either 15 min or 1 h postinjury, and at 24 h postinjury CAP recording was conducted in coronal brain slices. To elucidate how injury and CsA treatments affect specific populations of axons, CAP waveforms generated largely by myelinated axons (N1) were analyzed separately from the CAP signal, which predominantly reflects activity in unmyelinated axons (N2). CsA administration at 15 min postinjury resulted in significant protection of CAP area, and this effect was more pronounced in N1, than in the N2, CAP component. This treatment also significantly protected against TBI-induced reductions in high-frequency responding of the N1 CAP signal. In contrast, CsA treatment at 1 h did not significantly protect CAPs but was associated with atypical waveforms in N1 CAPs, including decreased CAP duration and reduced refractoriness. The present findings also support growing evidence that myelinated and unmyelinated axons respond differentially to injury and neuroprotective compounds.

Structural and Functional Changes in Nerve Roots Due to Tension at Various Strains and Strain Rates: An In-Vivo Study

This study investigates the functional and structural responses of spinal nerve roots in vivo to various strains and strain rates. Seventy-two L5 dorsal nerve roots from male Sprague-Dawley rats were each subjected to a predetermined strain (<10%, 10-20%, and >20%; n = 8) and rate (0.01 mm/sec, 1 mm/sec, or 15 mm/sec; n = 24). Neurophysiologic recordings were performed before and after stretch to determine changes in conduction velocity (CV), amplitude, and area of the compound action potential (CAP). Morphological injury as evident by primary and secondary axotomy as well as impaired axoplasmic transport (IAT) was determined using the palmgren silver impregnation technique and betaAPP immunostaining, respectively. The results from neurophysiologic recordings indicate that as strain and rate increased, there was a decrease in CV, amplitude, and area of the CAP. Further, high strains led to a complete conduction block that appeared to be rate dependent. Strains of 16%, 10%, and 9%, at 0.01 mm/sec, 1 mm/sec, and 15 mm/sec, respectively, led to 50% probability of complete conduction block in the nerve roots. Results from histological assessment indicate an increase in periaxonal spacing (secondary axotomy) and torn fibers (primary axotomy), as well as impaired IAT, with increasing strain and rate. Overall, the results from the current study indicate that (1) functional nerve root injuries as evident by changes in the CV, amplitude, and area of the CAP are strain- and rate-dependent; (2) high strains at low rates cause complete conduction block in the roots, while a similar block was observed at lower strains at the high rate; (3) the extent of IAT and primary and secondary axotomy occurred concomitant with functional injury and were strain- and rate-dependent.

Posttraumatic administration of pituitary adenylate cyclase activating polypeptide in central fluid percussion injury in rats

Several in vitro and in vivo experiments have demonstrated the neuroprotective effects of pituitary adenylate cyclase activating polypeptide (PACAP) in focal cerebral ischemia, Parkinson's disease and traumatic brain injury (TBI). The aim of the present study was to analyze the effect of PACAP administration on diffuse axonal injury (DAI), an important contributor to morbidity and mortality associated with TBI, in a central fluid percussion (CFP) model of TBI. Rats were subjected to moderate (2 Atm) CFP injury. Thirty min after injury, 100 microg PACAP was administered intracerebroventricularly. DAI was assessed by immunohistochemical detection of beta-amyloid precursor protein, indicating impaired axoplasmic transport, and RMO-14 antibody, representing foci of cytoskeletal alterations (neurofilament compaction), both considered classical markers of axonal damage. Analysis of damaged, immunoreactive axonal profiles revealed significant axonal protection in the PACAP-treated versus vehicle-treated animals in the corticospinal tract, as far as traumatically induced disturbance of axoplasmic transport and cytoskeletal alteration were considered. Similarly to our former observations in an impact acceleration model of diffuse TBI, the present study demonstrated that PACAP also inhibits DAI in the CFP injury model. The finding indicates that PACAP and derivates can be considered potential candidates for further experimental studies, or purportedly for clinical trials in the therapy of TBI.

Minor Traumatic Brain Injury (mTBI) in Ice Hockey and Other Contact Sports

Minor Traumatic Brain Injury (mTBI) is caused by the inertial effect of a mechanical impact to the head with sudden rotational acceleration forces. MTBI produces, in the less severe cases, only transient disturbances of ionic homeostasis with temporary disturbances of brain function. Depending on the severity of the trauma, animal and human studies have demonstrated focal intraaxonal alterations in neurofilamentous/cytoskeletal network and impairment of axoplasmic transport, which may lead to progressive axonal swelling, detachment or even cell death over a period of hours or days, the so-called process of delayed axotomy. Disturbances of ionic homeostasis, acute metabolic changes and cerebral blood flow alterations compromise the ability of neurons to function and render brain cells vulnerable. These processes may predispose brain cells to a vulnerable state for an undetermined period; therefore, it is recommended that any confused player with or without amnesia should be taken off the ice and not be permitted to play again for at least 72 h.

P3-286 UB+1 causes mitochondria jam in neurites and neuronal degeneration through impairment of axoplasmic transport

P3-286 UB+1 causes mitochondria jam in neurites and neuronal degeneration through impairment of axoplasmic transport

Diffuse Axonal Injury in Head Trauma

Diffuse axonal injury (DAI) is one of the most common and important pathologic features of traumatic brain injury (TBI). The susceptibility of axons to mechanical injury appears to be due to both their viscoelastic properties and their high organization in white matter tracts. Although axons are supple under normal conditions, they become brittle when exposed to rapid deformations associated with brain trauma. Accordingly, rapid stretch of axons can damage the axonal cytoskeleton resulting in a loss of elasticity and impairment of axoplasmic transport. Subsequent swelling of the axon occurs in discrete bulb formations or in elongated varicosities that accumulate transported proteins. Calcium entry into damaged axons is thought to initiate further damage by the activation of proteases. Ultimately, swollen axons may become disconnected and contribute to additional neuropathologic changes in brain tissue. DAI may largely account for the clinical manifestations of brain trauma. However, DAI is extremely difficult to detect noninvasively and is poorly defined as clinical syndrome. Future advancements in the diagnosis and treatment of DAI will be dependent on our collective understanding of injury biomechanics, temporal axonal pathophysiology, and its role in patient outcome.

The Immunophilin Ligand FK506 Attenuates the Axonal Damage Associated with Rapid Rewarming Following Posttraumatic Hypothermia

Our laboratory has shown that traumatically induced axonal injury (TAI) is significantly reduced by posttraumatic hypothermia followed by slow rewarming. Further, TAI can be exacerbated by rapid rewarming, and the damaging consequences of rapid rewarming can be reversed by cyclosporin A, which is believed to protect via blunting mitochondrial permeability transition (MPT). In this communication, we continue investigating the damaging consequences of rapid posthypothermic rewarming and the protective role of immunophilin ligands using another member of the immunophilin family, FK506, which does not affect MPT but rather inhibits calcineurin. Rats were subjected to impact-acceleration brain injury followed by the induction of hypothermia with subsequent rapid or slow posthypothermic rewarming. During rewarming, animals received either FK506 or its vehicle. Three hours postinjury, animals were prepared for the visualization of TAI via antibodies targeting impaired axoplasmic transport (APP) and/or overt neurofilament alteration (RMO-14). Rapid rewarming exacerbated TAI, which was attenuated by FK506. This protection was statistically significant for the APP-immunoreactive fibers but not for the RMO-14-positive fibers. Combined labeling, using one chromagen to visualize both axonal changes, suggested that these two immunoreactive profiles revealed two distinct pathologies not occurring along the same axon. Collectively, these studies confirmed previous observations identifying the adverse consequences of rapid rewarming while also showing the complexity of the pathobiology of TAI. Additionally, the demonstration that FK506 is protective suggests that calcineurin may be a major target for neuroprotection.

Axonal Damage in Traumatic Brain Injury

Axonal damage is one of the most common and important pathologic features of traumatic brain injury. Severe diffuse axonal injury, resulting from inertial forces applied to the head, is associated with prolonged unconsciousness and poor outcome. The susceptibility of axons to mechanical injury appears to be due to both their viscoelastic properties and their highly organized structure in white matter tracts. Although axons are supple under normal conditions, they become brittle when exposed to rapid deformations associated with brain trauma. Accordingly, rapid stretch of axons can damage the axonal cytoskeleton, resulting in a loss of elasticity and impairment of axoplasmic transport. Subsequent swelling of the axon occurs in discrete bulb formations or in elongated varicosities that accumulate organelles. Calcium entry into damaged axons is thought to initiate further damage by the activation of proteases and the induction of mitochondrial swelling and dysfunction. Ultimately, swollen axons may become disconnected and contribute to additional neuropathologic changes in brain tissue. However, promising new therapies that reduce proteolytic activity or maintain mitochondrial integrity may attenuate progressive damage of injured axons following experimental brain trauma. Future advancements in the prevention and treatment of traumatic axonal injury will depend on our collective understanding of the relationship between the biomechanics and pathophysiology of various phases of axonal trauma.

Initiating mechanisms involved in the pathobiology of traumatically induced axonal injury and interventions targeted at blunting their progression.

To gain better insight into the initiating factors involved in traumatically induced axonal injury cats and rats were subjected to various forms of traumatic brain injury. Following injury at intervals ranging from 10 min. to 3 hours, the animals were sacrificed and prepared in accordance with multiple immunocytochemical strategies capable of detecting focal changes in the axolemma, the subaxolemmal spectrin network, the underlying cytoskeleton as well as any related abnormalities in axoplasmic transport. Through these approaches it was recognized that the most severe forms of injury resulted in focal abnormalities of axonal permeability which were observed together with calpain-mediated spectrin proteolysis in the subaxolemmal network. These events were associated with compaction of the underlying neurofilaments and some microtubular loss which occurred without any direct evidence of overt axoplasmic proteolysis with the exception of the most severely injured fibers. In addition to these severely injured axonal profiles, other injured axons did not manifest overt changes in axolemmal permeability or early calpain-mediated spectrin proteolysis but demonstrated dramatic neurofilament and microtubular misalignment and impaired axoplasmic transport. Lastly, other small caliber axons showed another form of intraaxonal change manifested in the local pooling of organelles in the nodal and paranodal regions, with the suggestion that some of these changes may be reversible. In relation to these axonal responses the efficacy of various therapeutic investigations were assessed. The use of calcium chelators showed a trend for protection in those axons manifesting altered axolemmal permeability. However, the use of early and delayed hypothermia demonstrated dramatic protection resulting in significant reduction in the number of damaged axonal profiles. These studies illustrate the diversity and complexity of those axonal responses evoked by traumatic brain injury, suggesting that multiple forms of therapy may be needed to blunt these multifaceted forms of progression.

Diffuse Axonal Injury: An Important Form of Traumatic Brain Damage

Diffuse axonal injury (DAI) is a frequent form of traumatic brain injury in which a clinical spectrum of in creasing injury severity is paralleled by progressively increasing amounts of axonal damage in the brain. When less severe, DAI is associated with concussive syndromes; when most severe, it causes prolonged traumatic coma that is not related to mass lesions, increased intracranial pressure, or ischemia. Pathological investigations of DAI demonstrate widespread but heterogeneous microscopic damage to axons throughout the white matter of the cerebral and cerebellar hemispheres and brainstem. There is a propensity for injury to occur in the central third of the brain, and the corpus callosum and brain stem are especially prone to injury. In these locations, traumatic axonal damage can occur in several degrees of severity, ranging from transient disturbances of ionic homeostasis to swelling, impairment of axoplasmic transport with secondary (delayed) axotomy and primary axotomy (tearing). A more detailed understanding of the processes involved in axonal damage is crucial to the development of effective treatment for the clinical syndromes of DAI. NEUROSCIENTIST 4:202-215, 1998

Traumatically induced axonal damage: evidence for enduring changes in axolemmal permeability with associated cytoskeletal change.

Recent studies have demonstrated that delayed or secondary axotomy is a consistent feature of traumatic brain injury in both animals and man. Moreover, these studies have shown that the pathogenesis of this secondary axotomy involves various forms of initiating pathology, with the suggestion that, in some cases, only the axonal cytoskeleton is perturbed, while, in other cases, both the axonal cytoskeleton and related axolemma manifest traumatically induced perturbations. In the current communication, we continue in our investigation of the significance of these traumatically induced alterations in axolemmal permeability and their relation to any related intra-axonal cytoskeletal change. This was accomplished in cats which received intrathecal infusions of peroxidase, an agent normally excluded by the intact axolemma. These animals were subjected to traumatic brain injury, and sites showing altered axolemmal permeability to the peroxidase were assessed at the light and electron microscopic level. Through this approach, we recognized that a traumatic episode of moderate severity evoked changes in axolemmal permeability which surprising endured for up to 5 hrs postinjury. At such focal sites of altered permeability, the related cytoskeleton showed a statistically significantly neurofilament compaction, with the strong suggestion of concomitant neurofilament sidearm loss, microtubular dispersion, and mitochondrial abnormality. Over time, these events led to further disorganization of the axonal cytoskeleton which translated into impaired axoplasmic transport and secondary axotomy. Most likely, these alterations in axolemmal permeability result in either the direct or indirect effects upon the axonal cytoskeleton that precipitate the damaging sequences resulting in delayed axotomy.

Are the pathobiological changes evoked by traumatic brain injury immediate and irreversible?

Traumatic brain injury has long been thought to evoke immediate and irreversible damage to the brain parenchyma and its intrinsic vasculature. In this review, we call into question the correctness of this assumption by citing two traumatically related brain parenchymal abnormalities that are the result of a progressive, traumatically induced perturbation. In this context, we first consider the pathogenesis of traumatically induced axonal damage to show that it is not the immediate consequence of traumatic tissue tearing. Rather, we illustrate that it is a delayed consequence of complex axolemmal and/or cytoskeletal changes evoked by the traumatic episode which then lead to cytoskeletal collapse and impairment of axoplasmic transport, ultimately progressing to axonal swelling and disconnection. Second, we consider the traumatized brain's increased neuronal sensitivity to secondary ischemic insult by showing that even after mild traumatic brain injury, CA1 neuronal cell loss can be precipitated by the induction of sublethal ischemic insult within 24 hrs of injury. In demonstrating this increased sensitivity to secondary insult, evidence is provided that it is triggered by the neurotransmitter storm evoked by traumatic brain injury, allowing for sublethal neuro-excitation. In relation to this phenomenon, the protective effect of receptor antagonists are discussed, as well as the concept that this relatively prolonged posttraumatic brain hypersensitivity offers a potential window for therapeutic intervention. Collectively, it is felt that both examples of the brain parenchyma's response to traumatic brain injury show that the resulting pathobiology is much more complex and progressive than previously envisioned, and as such, rejects many of the previous beliefs regarding the pathobiology of traumatic brain injury.

Pathobiology of traumatically induced axonal injury in animals and man

Although diffuse axonal injury is recognized as a consistent feature of traumatic brain injury, there is confusion regarding its pathogenesis. To provide insight into its pathogenesis, animal models of traumatic brain injury complemented by post mortem human analyses were used. In animals, anterograde tracers together with antibodies targeting the neurofilament subunits were used in light and electron microscopic analyses of axonal injury. In humans, antibodies to the neurofilament subunits also were used to follow diffuse axonal injury. Animals were followed from minutes to months after injury, whereas humans were studied from six hours to 59 days after injury. In neither animals nor humans did traumatic brain injury cause direct axonal tearing. Instead, the traumatic brain injury triggered focal intra-axonal change in the 68-kd neurofilament subunit, which became disordered in its alignment and resulted in impaired axoplasmic transport. This caused axonal swelling and disconnection. The sequence of axonal change was similar in animals and man; however, its temporal progression was slower in humans. Traumatically induced axonal damage is triggered first by focal intra-axonal change involving the neurofilament subunits. This neurofilament change is due to either direct mechanical failure of the axonal cytoskeleton or the initiation of a biochemical event that causes neurofilament disassembly. In general, the temporal progression of the intra-axonal changes that lead to ultimate disconnection is influenced by the severity of the traumatic injury and the species evaluated.

Effect of IDPN on the expression of POMC-derived peptides in rat motoneurones

Immunocytochemistry was used to detect β-endorphin and α-melanotropin (α-MSH) in lumbar spinal motoneurones in rats treated with β,β′-iminodiproprionitrile (IDPN), a neurotoxicant that targets motoneurones or corn oil, which has no known neurotoxicity. After IDPN treatment most of the motoneurones were immunoreactive for both peptides but after corn oil treatment immunostaining was negligible. It is suggested that increased expression of the POMC-derived peptides may be part of the regenerative repertoire of the damaged motoneurone regardless of the cause of the lesion. Alternatively the peptides may simply accumulate in the motoneurones as a result of impaired axoplasmic transport.

Traumatically Induced Axonal Injury: Pathogenesis and Pathobiological Implications

This work reviews the pathobiology of traumatically induced axonal injury. Drawing upon literature gleaned from the experimental and clinical setting, this review attempts to emphasize that, other than the most destructive insults, traumatic brain injury does not typically cause direct mechanical disruption of the axon. Rather, this review documents that with traumatic injury focal, subtle axonal change occurs, and that over time, such change leads to impaired axoplasmic transport, continued axonal swelling, and ultimate disconnection. The initial intra-axonal events that trigger the above described sequence of reactive axonal change are considered with focus on the possibility of either traumatically altered axolemmal permeability, direct cytoskeletal damage/perturbation, or more overt metabolic/functional disturbances. Not only does this review focus on the sequence of traumatically induced axonal change, but also, it considers its attendant consequences in terms of Wallerian degeneration and subsequent deafferentation. The concept that traumatically induced diffuse axonal injury leads to diffuse deafferentation is emphasized together with its pathobiological implications for morbidity and recovery. The potential for either adaptive or maladaptive neuroplasticity subsequent to such diffuse deafferentation is considered in the context of mild, moderate and severe traumatic brain injury.

Aluminum Inhibits Calpain‐Mediated Proteolysis and Induces Human Neurofilament Proteins to Form ProteaseResistant High Molecular Weight Complexes

We studied the effects of aluminum salts on the degradation of human neurofilament subunits (NF-H, NF-M, and NF-L, the high, middle, and low molecular weight subunits, respectively) and other cytoskeletal proteins using calcium-activated neutral proteinase (calpain) purified from human brain. Calpain-mediated proteolysis of NF-L, tubulin, and glial fibrillary acidic protein (GFAP), three substrates that displayed constant digestion rates in vitro, was inhibited by AlCl3 (IC50 = 200 microM) and by aluminum lactate (IC50 = 400 microM). Aluminum salts inhibited proteolysis principally by affecting the substrates directly. After exposure to 400 microM aluminum lactate and removal of unbound aluminum, human cytoskeletal proteins were degraded two- to threefold more slowly by calpain. When cytoskeleton preparations were exposed to aluminum salt concentrations of 100 microM or higher, proportions of NF-M and NF-H formed urea-insoluble complexes of high apparent molecular mass, which were also resistant to proteolysis by calpain. Complexes of tubulin and of GFAP were not observed under the same conditions. Aluminum salts irreversibly inactivated calpain but only at high aluminum concentrations (IC50 = 1.2 and 2.1 mM for aluminum lactate and AlCl3, respectively), although longer exposure to the ion reduced by twofold the levels required for protease inhibition. These interactions of aluminum with neurofilament proteins and the effects on proteolysis suggest possible mechanisms for the impaired axoplasmic transport of neurofilaments and their accumulation in neuronal perikarya after aluminum administration in vivo.