Rasmussen's encephalitis is a progressive childhood disease of unknown cause characterized by severe epilepsy, hemiplegia, dementia, and inflammation of the brain. During efforts to raise antibodies to recombinant glutamate receptors (GluRs), behaviors typical of seizures and histopathologic features mimicking Rasmussen's encephalitis were found in two rabbits immunized with GluR3 protein. A correlation was found between the presence of Rasmussen's encephalitis and serum antibodies to GluR3 detected by protein immunoblot analysis and by immunoreactivity to transfected cells expressing GluR3. Repeated plasma exchanges in one seriously ill child transiently reduced serum titers of GluR3 antibodies, decreased seizure frequency, and improved neurologic function. Thus, GluR3 is an autoantigen in Rasmussen's encephalitis, and an autoimmune process may underlie this disease.

The cloning of cDNAs that encode functional glutamate receptors makes it possible to produce antibodies that can be used as high-affinity probes for the localization and characterization of these receptors in the mammalian brain. We have made antibodies to different regions of the first cloned member of this family, GluR1, using bacterially overproduced antigen. On Western blots, these antisera detect glycoprotein(s) of 105 kDa present in crude membranes of the hippocampus and cerebellum. The 105-kDa band is associated with postsynaptic densities, and it is observed in cultured cells upon transfection with the GluR1 cDNA. Although glutamate receptors are thought to be the most prevalent excitatory ligand-gated ion channel in the mammalian brain, immunohistochemistry reveals that the receptors recognized by these antisera are localized predominantly in neurons of the cerebellum and some structures of the limbic system, including the hippocampus, the central nucleus of the amygdala, and portions of the septum. This pattern of expression is, in general, consistent with the distribution of GluR1 mRNA as determined by in situ hybridization histochemistry. Our results suggest that glutamate excitatory circuits recognized by these antisera are predominantly found in regions of the limbic system that are reciprocally interconnected.

The expression of neuronal nicotinic ACh receptors (nAChRs) and the subunits that compose these receptors by PC12 cells exposed to NGF has been studied. The analysis of total RNA reveals that the neuronal nAChR subunits alpha 3, alpha S, beta 2, beta 3, and beta 4, but not alpha 2 and alpha 4, are expressed in our PC12 cells. Within 48 hr of adding NGF to cultures, the RNA corresponding to alpha 3, alpha 5, beta 3, and beta 4 is decreased, but the beta 2 RNA increases for up to 6 d after NGF treatment. To determine the influence of NGF treatment on subunit protein expression, subunit-specific antisera were prepared. Immunocytochemistry detected antigen for alpha 3, alpha 5, beta 2, beta 3, and beta 4 (but not alpha 2 and alpha 4) in both NGF-treated and nontreated PC12 cells. The expression of nAChR subunit proteins, as measured by direct binding of antibodies to PC12 cells, does not change subsequent to 6 d of treatment with NGF. Whole-cell recording of PC12 cells shows that both the individual cell current density and response to the agonist cytisine were not altered after 5-7 d in NGF. However, the number of cells exhibiting detectable ACh-induced currents doubled. These results indicate that NGF increases the number of PC12 cells expressing ACh-sensitive nAChR currents but the activation is not the result of altering the amounts of individual nAChR subunit proteins. These data, taken together with the decrease in most nAChR subunit RNAs (except beta 2), suggest that NGF regulation of nAChRs may be through a posttranscriptional mechanism.

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