For the neurobiologist, this year's annual American Society for Cell Biology meeting offered a small gem in the form of the minisymposium "The Cell Biology of Neurodegenerative Diseases" chaired by Dennis Selkoe (Brigham and Women's Hospital) and Sam Sisodia (Johns Hopkins University). Three important points were made in their introduction. First, the identification of the genetic basis for many neurodegenerative disorders has opened new and technically useful approaches to elucidating molecular mechanisms. Second, a common pathological feature, the production of fibrillar aggregates, appears to underlie various types of neurodegenerations. Third (and to my mind, the most crucial) is the need to establish the relevance of any experimental model to the disease process in situ. It is this last point that needs to be especially emphasized by anybody using a cell culture system to investigate neurodegeneration.

The first talk, by Anat Shirvan (Yitzhak Rabin Medical Center, Israel), reported studies on the role of collapsin, an axon retracting factor, as a potentially deleterious agent in oxidative stress-induced neuropathologies. Dopamine treatment of chick sympathetic neurons was used to kill neurons by an oxidative stress. Shirvan suggested that this model is relevant to similar neuropathologies in Alzheimer's disease (AD), stroke, and Creutzfeldt-Jacob disease in addition to Parkinson's disease. Differential display of messages in neurons treated with dopamine revealed that both collapsin-responsive mediator (CRM) and collapsin 1 were coordinately induced.

CRM is a developmentally regulated protein that facilitates the repulsive cues of collapsin 1 to prohibit unwanted synaptic connectivity. Antibodies to collapsin provide protection against the oxidative stress. But what is the relevance to neurodegeneration of the re-expression of this developmentally regulated protein? Shirvan presented the hypothesis that neurons undergoing apoptotic cell death may secrete collapsin, which results in degeneration of neighboring neurons. However, only those neurons expressing receptors for collapsin would be vulnerable to this neurodegeneration. Thus, collapsin, a secreted factor that is integral to appropriate synaptogenesis during brain development, may upon re-expression in the adult brain lead to neurodegeneration. This hypothesis leads to many important questions. The most pressing issue may be determining if a subpopulation of adult neurons express the collapsin receptor. If so, it will be crucial to determine whether this pattern of expression reflects the selective neuron vulnerability observed in diseases such as Alzheimer's.

The next two talks focused on two distinct neuropathologies that involve the selective vulnerability of long neurons, which rely upon efficient axonal transport. William Saxton (Indiana University) used null mutations of the kinesin gene (Khc) in Drosophila to investigate the consequences of disrupted fast axonal transport. He argued such a model is particularly relevant to peripheral neuropathies that show evidence of such disrupted transport, including axonal swellings, atrophy of distal axonal bodies, and progressive loss of specific neurons that have long axons, which would be most vulnerable.

Indeed, the Khc mutants show a posterior-first loss of neuron function and reduced innervation of the neuromuscular junction. Only fast, anterograde transport was affected. Electron microscopy provided evidence for swollen axons containing various intracellular organelles caught in a "traffic jam." The result of such jams may be the eventual degeneration of the neuron. Interestingly, several mutations along the Khc gene gave essentially the same phenotype. These mutants may provide an animal model for the early events in similar peripheral neuropathies in humans. Are there gene mutations for kinesin, or other cellular motor proteins involved in axonal transport, in the human population? This model should inspire a candidate gene search for such mutations.

A role for disrupted axonal tubulin transport in amyotrophic lateral sclerosis (ALS) was discussed by T.L. Williamson (University of California at San Diego). ALS is the result of degeneration of large motor neurons of the spinal cord. A small fraction of ALS cases are caused by a mutation in the superoxide dismutase 1 (SOD1) gene. Williamson et al. used a transgenic mouse harboring the human mutated SOD1 gene to examine axonal transport, using the rationale that the motor neurons that degenerate in ALS are selectively vulnerable because of their axonal length.

They found evidence of slowed transport of tubulin (but not actin or neurofilament) and accumulation of "exploded organelles" in the axons of the spinal cord neurons in the transgenic mice. This disruption occurred prior to the loss of axons and motor neuron function. The authors argue that the reduced axon transport leads to the loss of neuromuscular innervation and muscle atrophy that occurs in ALS. The curious absence of reduced neurofilament transport (which is normally observed in ALS) in these mice suggests that such a disruption may be secondary to the effects on tubulin transport.

A cellular model for the neuropathology of prion diseases was presented by D.A. Harris (Washington University School of Medicine). Prions are involved in several human neuropathologies, including Creutzfeldt-Jacob disease, kuru, Gerstmann-Straussler-Scheinker syndrome, and fatal familial insomnia. Prions also play a role in the animal pathologies scrapie and bovine spongiform encephalopathy. The molecular pathology of these diseases occurs as a consequence of the post-translational conversion of PrPc, a normally expressed protein of unknown function, to PrPsc, a protease-resistant, aggregatable protein.

Harris presented a cell biological approach to investigating this conversion, using Chinese hamster ovary (CHO) cells transfected with mutated forms of the PrP gene. The kinetics of PrPsc generation indicate that the conversion occurs in distinct biochemical steps. One step appears to be an initial resistance to cleavage of the GPI anchor to the cell membrane, which may occur in the endoplasmic reticulum (ER) soon after its synthesis. The other biochemical steps may involve two events that occur on the cell surface: insolubility to detergent treatment, and resistance to normal proteolytic cleavage. It was suggested that the first step in the conversion was the co-translational alteration of PrPc conformation in the ER and may involve an inappropriate association of the nascent PrPc with its chaperone. Thus, these studies may have pinpointed the pathological conversion of PrP to a disruption of co-translational processing. However, determining how the PrPsc kills neurons remains an open question. Why are only certain cells vulnerable to these mutations in various neuropathologies?

The final two talks discussed the molecular pathology of AD: Dennis Selkoe's talk from the perspective of amyloid precursor protein (APP), and Sam Sisodia's talk from that of the presenilins (PS). Selkoe argued that production of the Ab peptide from APP is the initial central event in the pathology of AD, particularly, when APP is processed to generate the longer form of Ab (1-42). Thus, it is important to elucidate the cellular mechanisms responsible for this production.

Selkoe's laboratory has investigated the processing of APP from the cell surface into clathrin-coated endosomes to lysosomes. They initially found that the shorter form of Ab (1-40) was generated from APP in the endosomes by g-secretase activity. However, in cells transfected with PS1, the long (1-42) form of Ab was produced. Although PS1 is present predominantly in the ER, both PS1 and APP were fractionated in the Golgi apparatus, along with Ab (both forms) in PS1-transfected cells. Furthermore, Selkoe presented preliminary evidence that APP co-precipitates with PS1, which argues for a direct interaction between the two proteins in the ER/Golgi pathway. Can this interaction account for the role of PS mutations (which make up the majority of familial cases) in AD pathology? Transgenic mice containing both mutated APP and PS1 exhibit an exacerbated pathology including earlier deposition of amyloid deposits. It will be important to determine if the in vitro observations in the transfected cells are replicated in the appropriate neurons in the brains of these animals.

The final talk was a presentation by Sam Sisodia on the biology of the presenilins (PS1 and PS2). Although PS mRNA appears at high levels in neurons, very little protein is present. The protein is rarely evident in its intact form but is predominantly present as two cleavage products (29 and 18 kD). This cleavage appears to be the normal processing of the protein because they always occur in a one to one stoichiometry and co-reside within cells. Overexpression of either PS leads to the inhibition of expression of the other.

Sisodia discussed the homology between PS and SEL12 (of C. elegans), which is related to notch. Indeed, PS knockout mice exhibit developmental problems of somitogenesis that would be expected from disrupted notch function. What is the relevance of the PS knockouts to neuropathology? These knockouts result in Ab levels that are dramatically reduced for both short and long forms, and APP and its C-terminal fragments (the precursors to Ab) accumulate in the neurons. Contrary to the previous talk, there is no evidence for a direct interaction between APP and PS1. Instead, Sisodia suggested that PS may affect the g-secretase cleavage site on the APP molecule. Clearly, based upon these two talks, it will be crucial to understand the molecular interaction between PS and APP. However, AD is most often not associated with mutations of either of these proteins. It will eventually be necessary to determine the relevance of these mutated proteins to the general neuropathology of AD to test the initial hypothesis that production of Ab is the central event.

What causes neurons to die? The answer seems to be as varied as the number of different neuropathologies that have been described. Although the subject of this symposium was neurodegeneration, the variety of disorders was matched by the variety of molecular mechanisms that appear to underlie the diseases. It is becoming clear that the etiology of each disorder is distinct from the others, as evidenced by the different cellular pathologies exhibited by the affected neurons.

How do neurons die? Any common themes, such as fibrillar aggregation (AD and prion diseases) or disrupted axonal transport (peripheral neuropathies and ALS) shared by these neurodegenerations may represent the late stages of the neurodegenerative processes. These late stages may represent the important targets for therapeutic interventions.

Why do certain neurons die and others appear to be unaffected? Understanding the genetic basis for neuropathologies offers new experimental approaches to the problems, including transgenic mice, transfected cell lines, and knockout Drosophila, but raises an important question. How can we take advantage of those neurons that remain selectively invulnerable to these genetic diseases while they are also in the environment of neurodegeneration?