Autoantibodies against astrocyte water channel aquaporin-4 (AQP4) are highly specific for the neuroinflammatory disease neuromyelitis optica (NMO). We measured the binding of NMO autoantibodies to AQP4 in human astrocyte-derived U87MG cells expressing M1 and/or M23 AQP4, or M23 mutants that do not form orthogonal array of particles (OAPs). Binding affinity was quantified by two-color fluorescence ratio imaging of cells stained with NMO serum or a recombinant monoclonal NMO autoantibody (NMO-rAb), together with a C terminus anti-AQP4 antibody. NMO-rAb titrations showed binding with dissociation constants down to 44 ؎ 7 nM. Different NMO-rAbs and NMO patient sera showed a wide variation in NMO-IgG binding to M1 versus M23 AQP4. Differences in binding affinity rather than stoichiometry accounted for M1 versus M23 binding specificity, with consistently greater affinity of NMO-IgG binding to M23 than M1 AQP4. Binding and OAP measurements in cells expressing different M1:M23 ratios or AQP4 mutants indicated that the differential binding of NMO-IgG to M1 versus M23 was due to OAP assembly rather than to differences in the M1 versus M23 N termini. Purified Fab fragments of NMO-IgG showed similar patterns of AQP4 isoform binding, indicating that structural changes in the AQP4 epitope upon array assembly, and not bivalent cross-linking of whole IgG, result in the greater binding affinity to OAPs. Our study establishes a quantitative assay of NMO-IgG binding to AQP4 and indicates remarkable, OAP-dependent heterogeneity in NMO autoantibody binding specificity.A defining feature of the neuroinflammatory demyelinating disease neuromyelitis optica (NMO) 2 is the presence of serum autoantibodies (NMO-IgG) against aquaporin-4 (AQP4), a water channel expressed in astrocytes throughout the central nervous system (1). The presence of NMO-IgG is specific for NMO, and in some reports serum NMO-IgG titers correlate with NMO disease activity (2, 3). Studies in rodents suggest that NMO-IgG is pathogenic in NMO. Human NMO-IgG produces many features of NMO disease in rats with preexisting experimental autoimmune encephalomyelitis (4, 5) or pretreated with complete Freund's adjuvant (6) and in naïve mice when injected together with human complement (7). These animals develop characteristic NMO lesions with neuroinflammation, perivascular deposition of activated complement, demyelination, and loss of astrocyte glial fibrillary acidic protein and AQP4 immunoreactivity.The target of the NMO autoantibody, AQP4, is involved in water balance in brain (8, 9) and spinal cord (10), sensory signal transduction (11, 12) and neuroexcitatory phenomena including seizure activity (13) and cortical spreading depression (14), and astrocyte migration and glial scarring (15, 16). AQP4 is expressed in astrocytes as two major isoforms: a long (M1) isoform with translation initiation at Met-1, and a shorter (M23) isoform with translation initiation at . M23 AQP4 assembles in membranes as regular square arrays called orthogonal arrays of particles (OAPs), whic...

The DID model demonstrates predictive validity. Both opioid and dopamine signaling are involved in ethanol drinking to intoxication. Different physiological pathways mediate high ethanol drinking as compared to water or sugar water drinking in DID. DID may be a useful screening tool to find new alcoholism medications and to discover genetic and neurobiological mechanisms relevant to the human disorder.

Results suggest pharmacokinetics may make a small contribution to differential stimulation between adolescents and adult mice, but are unlikely the only factor. Developmental differences within the brain that effect pharmacodynamic properties of psychostimulants (e.g., number of receptor or transporters) represent alternatives.

These results support the hypothesis that mGluR5 signaling plays a role in excessive ethanol intake in DID and suggest DID may have value for screening novel compounds that reduce overactive glutamate signaling for potential pharmaceutical treatment of excessive ethanol drinking behavior.

Adolescence is a time period when major changes occur in the brain with long-term consequences for behavior. One ramification is altered responses to drugs of abuse, but the specific brain mechanisms and implications for mental health are poorly understood. Here, we used a mouse model in which adolescents display dramatically reduced sensitivity to the acute locomotor stimulating effects of cocaine and methamphetamine. The goal was to identify key brain regions or circuits involved in the differential behavior. Male adolescent and young adult (PN 69-74) C57BL/6J mice were administered an intraperitoneal injection of cocaine (0, 15, 30 mg/kg) or methamphetamine (0, 2, 4 mg/kg) and euthanized 90 minutes later. Locomotor activity was monitored continuously in the home cage by video tracking. Immunohistochemical detection of Fos protein was used to quantify neuronal activation in 16 different brain regions. As expected, adolescents were less sensitive to the locomotor stimulating effects of cocaine and methamphetamine as indicated by a rightward shift in the dose response relationship. After a saline injection, adolescents showed similar levels of Fos as adults in all regions except the dorsal and lateral caudate where levels were lower in adolescents. Cocaine and methamphetamine dose dependently increased Fos in all brain regions sampled in both adolescents and adults, but Fos levels were similar in both age groups for a majority of regions and doses. Locomotor activity was correlated with Fos in several brain areas within adolescent and adult groups, and adolescents had a significantly greater induction of Fos for a given amount of locomotor activity in key brain regions including the caudate where they showed reduced Fos under baseline conditions. Future research will identify the molecular and cellular events that are responsible for the differential psychostimulant-induced patterns of brain activation and behavior observed in adolescent versus adult mice. Keywords adolescent; cocaine; methamphetamine; c-fos; locomotor stimulation; mice The literature on acute locomotor stimulation from cocaine and amphetamines in animal models dates back more than 80 years (Tatum and Seevers, 1929 Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. caudate, nucleus accumbens), neural circuits (e.g., natural reward, basal ganglia, motor), and specific cellular and molecular events (e.g., dopamine neurotransmission, DARPP-32 signaling) have been identified that contribute to increased physical activity, arousal, reward and other behaviors induced by these drugs (Gold et al., 1989, Uhl...