Pathological changes of Alzheimer disease are characterized by cerebral cortical atrophy as a result of degeneraton and oss of neurons. Typical histologcal lesions include numerous senile plaques composed of deposits of amyloid 3-protein and neurofibrillary tangles consisting predominantly of ubiquitin and highly phosphorylated tau proteins. Previousy, tau protein kinase I (TPK I) was purified and its cDNA was cloned. To emine the biological role of this enzyme in neurons, we have studied the induction of its kinase activity in primary cultures of embryonic rat hippocampal neurons. Treatment of cultures with amyloid 1-protein significantly increased TPK I activity and induced the appearance of tau proteins cognized by the Alz-50 monoclonal antibody. In addition, though amyloid 1-protein was neurotoxic, either cycloheximide or actinomycin D prevented neuronal death.Death was also prevented by TPK I antisense oligonudeotides but not by sense oligonucleotides. These observations suggest that rat hippocampal neurons undergo programmed ceil death in respons to amyloid 1-protein and that TPK I is a key enzyme In this process.The histopathological lesions of Alzheimer disease (AD) are well known yet poorly understood. At autopsy, AD brains typically show diffuse cerebral cortical atrophy with varying distributions that often include the hippocampus, reflecting widespread granulovacular degeneration and loss ofneurons. Microscopically, amyloid deposits are found in large numbers of senile plaques scattered throughout the extracellular matrix as well as in the walls of the cerebral blood vessels. Many neurons also contain intracytoplasmic neurofibrillary tangles (NFI) (1). Amyloid (-protein (AP), a polypeptide derived by proteolytic cleavage ofamyloid precursor protein, is a major component of senile plaques (2). Ubiquitin (3,4) and highly phosphorylated tau proteins, members of a family of microtubule-associated phosphoproteins, predominantly compose NFT. The tau proteins form paired helical filaments (PHF), which are found in NFT and degenerative neurites of senile plaques (5-7). The molecular mechanisms by which these lesions arise are unknown. They are thought to result from defects in proteolytic processing of the corresponding precursor polypeptides, but whether they are the primary causes of neurodegeneration or merely remnants of this process remains obscure. and neuronal death (1, 2, 11, 12). The specific details of this process are lacking, however, and whether neurons die by passive necrosis or by programmed cell death requiring protein synthesis is a matter of speculation. It is a reasonable assumption that the ability of NFT-containing neurons to function normally is severely damaged. Accordingly, the appearance of NFT may be an important indicator of the extent of brain damage. Despite intensive efforts, however, the sequence of events leading to the formation of NFT is unknown in AD, and for ethical reasons in vitro model systems are needed for analysis of these events at the molecular level.In sea...

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Immunoglobulin G fractions from patients with Lambert-Eaton myasthenic syndrome (LEMS), an autoimmune disease of neuromuscular transmission, immunoprecipitate '2SI-labeled a-conotoxin GVIA-labeled calcium channels solubilized from rat brain. A 58-kDa antigen was detected by probing Western blots of partially purified calcium channels with LEMS plasma and IgG and was shown to be the relevant antigen in w-conotoxin receptor immunoprecipitation. Monoclonal antibody 1D12, produced by immunizing mice with synaptic membranes, has properties similar to these autoimmune IgGs in both immunoprecipitation and Western blotting assays. 1D12 antigen was purified by immunoaffinity chromatography and shown to bind LEMS IgG. The antigen was identified by screening a rat brain cDNA library with 1D12 and was found to have strong homology to the synaptic vesicle membrane protein synaptotagmin. Our results indicate therefore that these antibodies immunoprecipitate w-conotoxin receptors by binding to synaptotagmin that is associated with calcium channels. We suggest that the interaction between synaptotagmin and the voltage-gated calcium channel plays a role in docking synaptic vesicles at the plasma membrane prior to rapid neurotransmitter release and that autoantibody binding to a synaptotagmin-calcium-channel complex may be involved in the etiology of LEMS.

Synaptotagmin (p65) is an abundant synaptic vesicle protein of neurons and contains regions similar to the regulatory domain of protein kinase C. These domains are thought to be involved in calcium-dependent interaction with membrane phospholipids during exocytosis. To assess the functional role of synaptotagmin, synaptotagmin-deficient clonal variants of PC12 cells were isolated. All of the variant cells released catecholamine and adenosine triphosphate in response to elevated intracellular concentrations of calcium, which suggests that synaptotagmin is not essential for secretion of catecholamine and adenosine triphosphate from PC12 cells.