Interleukin-1 beta converting enzyme (ICE) is a mammalian homolog of CED-3, a protein required for programmed cell death in the nematode Caenorhabditis elegans. The activity of ICE can be specifically inhibited by the product of crmA, a cytokine response modifier gene encoded by cowpox virus. Microinjection of the crmA gene into chicken dorsal root ganglion neurons was found to prevent cell death induced by deprivation of nerve growth factor. Thus, ICE is likely to participate in neuronal death in vertebrates.

aggregates that may be toxic to neurons (9-11). Neurotransmitters can regulate APP processing to favor the secretion of APPS (12). In human embryonic kidney (HEK) 293 cells stably expressing the human muscarinic receptor subtypes ml or m3, stimulation with the muscarinic agonist carbachol increased APP, secretion. The muscarinic receptors mediating this effect (ml and m3 but not m2 or m4) are coupled to intracellular signaling pathways via second messengers, diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (InsP3), which are generated by phosphatidylinositol 4,5-bisphosphate (PtdInsP2) hydrolysis. In other established cell lines, accelerated APP metabolism produced by direct stimulation of protein kinase C (PKC) or inhibition of phosphatase activity also increased APP, secretion (13-16). In HEK 293 cells overexpressing ml receptors, there was a reciprocal relationship between APP, and AP3 secretion after receptor activation with carbachol (17); thus, receptor activation can enhance APP, secretion and also suppress AP3 formation.Abundant levels of both APP message and protein are present in neurons. The detection of APP, in plasma and cerebrospinal fluid (18,19) suggests that proteolysis of APP into soluble derivatives occurs in the central nervous system. Because receptor activation can regulate APP processing, impaired neurotransmission could conceivably exacerbate amyloid formation in AD, particularly in cortex and hippocampus. In addition to the loss of cholinergic basal forebrain neurons (20), glutamatergic corticocortical connections and major projections of the hippocampus are especially vulnerable to damage in AD (21). These pathways atrophy during the early stages of the disease, and in postmortem brain glutamate concentrations are decreased by as much as 80% (22). Because glutamate activates PKC in nervous tissue (23, 24), we tested the hypothesis that glutamate might divert APP processing from amyloidogenic pathways and, instead, favor increased APPs secretion.Metabotropic glutamate receptors (mGluR) are coupled to the formation of multiple second messengers via activation of phospholipase enzymes (23-25). Nonselective glutamate agonists [e.g., L-glutamate, quisqualate (QA)] can interact with both mGluR and ionotropic glutamate receptors (iGluR), but the selective mGluR agonist, trans-(1S,3R)-l-amino-1,3-cyclopentane dicarboxylic acid (ACPD) initiates signal transduction without affecting the iGluR. mGluR exist as seven subtypes and are categorized into three major groups on the

The hindbrain is evolutionarily conserved among diverse vertebrate phyla. In vertebrate embryos, the hindbrain is segmentally organized as a series of overt swellings known as rhombomeres. In the larval zebrafish Brachydanio rerio, conspicuous and identifiable reticulospinal neurons are positioned in the center of rhombomeres. Segmentally homologous reticulospinal neurons that share a range of morphological, developmental, and biochemical features occupy adjacent rhombomeres. We have recently shown that reticulospinal neurons of the zebrafish survive ontogeny without considerable morphological modification and we suggested that homologous neurons may share similar functions at different stages of development (Lee and Eaton: Journal of Comparative Neurology 304:34-52, 1991). The goldfish Carassius auratus, a related cyprinid, is especially suited for neurophysiological and behavioral studies. However, it is not yet known if the various reticulospinal neurons of zebrafish are generalizable to other species such as the goldfish. Therefore, we sought to examine the extent to which reticulospinal neurons of the zebrafish are also present in the adult goldfish. Analysis of 45 brains retrogradely labeled with horseradish peroxidase (HRP) from the spinal cord showed that reticulospinal neurons are arranged as a series of seven segments within the hindbrain; a regular interval of approximately 200 microns separates adjacent segments. Although the goldfish reticulospinal system has more neurons than the zebrafish, many reticulospinal neuron types continue to be identifiable. Moreover, comparisons of dendritic arborizations and axon paths between the two species showed that the morphology between various neuron types is virtually identical. The cross-taxonomic similarities between the reticulospinal systems of these related cyprinids make it possible to pursue functional considerations of segmentally homologous neurons in the goldfish hindbrain.

Five different Alzheimer mutations of the ␤-amyloid precursor protein (APP) were expressed in neurons via recombinant herpes simplex virus (HSV) vectors, and the levels of APP metabolites were quantified. The predominant intracellular accumulation product was a Cterminal fragment of APP that co-migrated with the protein product of an HSV recombinant expressing the C-terminal 100 amino acids (C100) of APP, which is known to cause neurodegeneration. Fractionation studies revealed that the C-terminal fragment generated by expression of the Alzheimer mutations, like C100, partitioned into membrane fractions and was particularly enriched in synaptosomes. The processing abnormality caused by expression of the Alzheimer mutations occurs predominantly in neurons. Expression of these mutations or of C100 alone in neurons caused increased secretion of A␤ relative to that of neurons infected with wild type APP recombinant vectors. These data show that expression of APP mutations that cause familial Alzheimer's disease increases the intracellular accumulation of potentially amyloidogenic and neurotoxic C-terminal fragments of APP in neurons.Some familial forms of Alzheimer's disease (FAD) 1 are caused by autosomal dominant mutations (1) in the gene for the ␤-amyloid precursor protein (APP). Analyses of ␤-amyloid (A␤) in genetically engineered cell lines expressing these mutants have shown that expression of the "Swedish" mutant of APP causes increased overall secretion of A␤ (2, 3), whereas expression of the 717 mutations causes increased secretion of a "long" (42-43-amino acid) form of A␤ relative to the shorter 40-amino acid form (4). Consequently, a leading hypothesis for the etiology of AD is that increased A␤42(43) is a shared molecular correlate of FAD mutations and that it represents a gain of deleterious function that can cause FAD (5).This gain-of-function hypothesis is attractive; however, molecular mechanisms other than those mediated by extracellular A␤ could also constitute a deleterious gain of function that leads to AD neurodegeneration. In particular, intracellular events mediated by APP metabolites may cause neuronal death. For example, amyloidogenic C-terminal fragments of APP are neurotoxic in vitro and cause AD-like neuropathology in vivo (6 -8). Moreover, whereas previous studies showing increased secretion of A␤42 (43) by cells expressing FAD APP mutants in vitro have utilized cell lines, the metabolism of APP in neurons is of particular relevance to this neurodegenerative disorder. To identify metabolic fragments of APP that are generated by neurons in FAD, we used a herpes simplex virus (HSV) vector to express five different FAD mutants of APP in primary neurons in culture. All of the mutants caused abnormal intracellular increases in the level of a C-terminal APP fragment encompassing the A␤ sequence and cytoplasmic domain, as well as increased secretion of A␤. EXPERIMENTAL PROCEDURESGeneration of Recombinant HSV Vectors-Trans-polymerase chain reaction mutagenesis (9) was used to make the Swedish mu...