Alzheimer's Disease-Related Dementias Summit 2016: National research priorities.

Goal 1 of the National Plan to Address Alzheimer's Disease is to prevent and effectively treat Alzheimer disease and Alzheimer disease-related dementias by 2025. To help inform the research agenda toward achieving this goal, the NIH hosts periodic summits that set and refine relevant research priorities for the subsequent 5 to 10 years. This proceedings article summarizes the 2016 Alzheimer's Disease-Related Dementias Summit, including discussion of scientific progress, challenges, and opportunities in major areas of dementia research, including mixed-etiology dementias, Lewy body dementia, frontotemporal degeneration, vascular contributions to cognitive impairment and dementia, dementia disparities, and dementia nomenclature.

The National Institutes of Health Microphysiological Systems Program focuses on a critical challenge in the drug discovery pipeline.

The National Institutes of Health has partnered with the US Food and Drug Administration and the Defense Advanced Research Projects Agency to accelerate the development of human microphysiological systems (MPS) that address challenges faced in predictive toxicity assessment and efficacy analysis of new molecular entities during the preclinical phase of drug development. Use of human MPS could provide better models for predicting the efficacy of new molecular entities in clinical trials. It is also anticipated that improvements in predicting drug toxicities early in the drug development process through the use of MPS or human organs-on-a-chip will decrease the need to withdraw new therapies from the market and minimize or eliminate deaths due to unidentified drug toxicities.

Deletion of galectin-3 exacerbates microglial activation and accelerates disease progression and demise in a SOD1(G93A) mouse model of amyotrophic lateral sclerosis.

Galectins are pleiotropic carbohydrate-binding lectins involved in inflammation, growth/differentiation, and tissue remodeling. The functional role of galectins in amyotrophic lateral sclerosis (ALS) is unknown. Expression studies revealed increases in galectin-1 mRNA and protein in spinal cords from SOD1(G93A) mice, and in galectin-3 and -9 mRNAs and proteins in spinal cords of both SOD1(G93A) mice and sporadic ALS patients. As the increase in galectin-3 appeared in early presymptomatic stages and increased progressively through to end stage of disease in the mouse, it was selected for additional study, where it was found to be mainly expressed by microglia. Galectin-3 antagonists are not selective and do not readily cross the blood-brain barrier; therefore, we generated SOD1(G93A)/Gal-3(-/-) transgenic mice to evaluate galectin-3 deletion in a widely used mouse model of ALS. Disease progression, neurological symptoms, survival, and inflammation were assessed to determine the effect of galectin-3 deletion on the SOD1(G93A) disease phenotype. Galectin-3 deletion did not change disease onset, but resulted in more rapid progression through functionally defined disease stages, more severely impaired neurological symptoms at all stages of disease, and expiration, on average, 25 days earlier than SOD1(G93A)/Gal-3(+/+) cohorts. In addition, microglial staining, as well as TNF-α, and oxidative injury were increased in SOD1(G93A)/Gal-3(-/-) mice compared with SOD1(G93A)/Gal-3(+/+) cohorts. These data support an important functional role for microglial galectin-3 in neuroinflammation during chronic neurodegenerative disease. We suggest that elevations in galectin-3 by microglia as disease progresses may represent a protective, anti-inflammatory innate immune response to chronic motor neuron degeneration.

Executive functions in the heterozygous reeler mouse model of schizophrenia.

Deficits in working memory and executive functions are now considered among the most reliable endophenotypes for schizophrenia. To determine whether cognitive deficits exist in mouse models of the disease, the authors trained heterozygous reeler (+/rl) mice on a series of visual discriminations similar to those used to test executive abilities in primates. These mice resemble schizophrenia patients in that both have reduced levels of reelin protein and altered gamma aminobutyric acid neurotransmission in the prefrontal cortex. The +/rl mice showed a selective deficit in reversal learning, with a pattern of errors that suggested impaired visual attention rather than a deficiency in perseveration and inhibitory control. These results show that cognitive dysfunction may serve as a useful biomarker in mouse models of neuropsychiatric disease.

Glutamate transporter cluster formation in astrocytic processes regulates glutamate uptake activity.

Glutamate is the predominant excitatory neurotransmitter in the CNS, and it is removed from the synaptic cleft by sodium-dependent glutamate transport activity. Glutamate transporter-1 (GLT-1) is expressed predominantly in astroglial cells and is responsible for the largest proportion of glutamate transport in the adult forebrain. In the present study, we demonstrate the ability of endogenous and recombinant GLT-1 to form clusters in astrocytic processes and characterize the mobility and physiological importance of these clusters in the regulation of GLT-1 activity in the presence or absence of neurons. At the distal end of C6 glioma cell processes, GLT-1 clusters undergo rapid morphological changes in both shape and size, and these changes are inhibited by cytochalasin D treatment, suggesting that the morphogenesis of GLT-1 clusters is highly dependent on the actin network. Treatment of astrocytes with phorbol 12-myristate 13-acetate (PMA) quickly and preferentially decreases GLT-1 localization on the process membrane, leading to de novo generation of GLT-1 clusters along the process shaft. Pretreatment with the PKC inhibitor bisindolylmaleimide II (Bis II), with sucrose (0.4 m), or through the expression of a dominant-negative form of dynamin prevents PMA-induced GLT-1 internalization and cluster formation. In terms of glutamate transporter function, PMA treatment elicits a significant decrease in GLT-1 activity that is prevented by preexposure to either Bis II or hypertonic treatment. Together, these data indicate that GLT-1 trafficking and cluster formation in glial cell processes are dynamic events that play important roles in regulating glutamate uptake in astrocytes and glioma cells.

Abbreviated action potential kinetics in a mouse model of potassium channel overexpression during hippocampal development.

Loss or "gain" of function mutations in voltage-gated ion channels often results in an adverse neurological phenotype. We have examined the electrical characteristics of hippocampal pyramidal cells in a transgenic mouse model to determine how overexpression of a Shaker-type potassium channel subunit during early postnatal development might alter excitability properties of developing neurons. We found that in CA3 neurons potassium channel overexpression led to a transient shortening in duration of single action potentials during the first two postnatal weeks. There was an increase in maximal repolarization rate, without significant effect on the rate of rise. Transgenic CA3 neurons also showed a decrease of firing frequency in response to sustained depolarizing current injection. In contrast, repolarization of action potentials in CA1 neurons was not significantly altered by trangene expression. Western Blot Analysis of membrane-associated transgene protein indicated that transgene protein levels decreased during development, in agreement with functional measures of spike width. Our data indicate that the functional consequences of potassium channel transgene expression are dependent on cellular environment and developmental stage. A transient period of hypoexcitability during a critical period of development for CA3 neurons may contribute to the hyperexcitable phenotype observed in adult animals.

Loss of glial fibrillary acidic protein results in decreased glutamate transport and inhibition of PKA-induced EAAT2 cell surface trafficking.

Loss of the astrocyte-specific intermediate filament protein, glial fibrillary acidic protein (GFAP) results in an increased susceptibility to ischemic insult, enhanced hippocampal LTP, and decreased cerebellar long-term depression (LTD). Because glutamate receptor activation plays a key role in cell death and cellular plasticity responses, we wanted to determine if alterations in glial glutamate transport could contribute to the GFAP null phenotype. To address functional changes in glutamate transport, we measured glutamate uptake in cortical, cerebellar, and hippocampal synaptosomal preparations from age-matched adult wild type and GFAP null mice and demonstrated a 25-30% reduction in the V(max) for d-aspartate uptake in the cortex and hippocampus of GFAP null animals. Western blot analysis of cortical synaptosomal fractions from wild type and GFAP null animals demonstrated that loss of GFAP results in decreases in both astrocytic (EAAT1) and neuronal (EAAT3) glutamate transporter subtypes. Immunohistochemical analysis demonstrated a region-specific modification of neuronal glutamate transporter, EAAT3 trafficking in the GFAP null phenotype. Analysis of primary cortical astrocyte cultures prepared from GFAP null and wild type mice demonstrated that loss of GFAP results in an inability to traffic the glial glutamate transporter, EAAT2, to the surface of the cell following protein kinase A (PKA) stimulation by dibutyryl cAMP. Taken together, these results suggest that the intermediate filament protein, GFAP plays a key role in modulating astrocytic and neuronal glutamate transporter trafficking and function.

Mutation of the GABAA receptor M1 transmembrane proline increases GABA affinity and reduces barbiturate enhancement.

All GABA(A) receptor (GABAR) subunits include an invariant proline in a consensus motif in the first transmembrane segment (M1). In receptors containing bovine alpha1, beta1 and gamma2 subunits, we analyzed the effect of mutating this M1 proline to alanine in the alpha1 or beta1 subunit using 3 different expression systems. The beta1 subunit mutant, beta1(P228A), reduced the EC(50) for GABA about 10-fold in whole cell recordings in HEK293 cells and L929 fibroblasts. The corresponding alpha1 subunit mutant (alpha1(P233A)) also reduced the GABA EC(50) when expressed in Xenopus oocytes; alpha1(P233A)beta1gamma2S receptors failed to assemble in HEK293 cells. Binding of [(3)H]flumazenil and [(3)H]muscimol to transfected HEK293 cell membranes showed similar levels of receptor expression with GABARs containing beta1 or beta1(P228A) subunits and no change in the affinity for [(3)H]flumazenil; however, the affinity for [(3)H]muscimol was increased 6-fold in GABARs containing beta1(P228A) subunits. In L929 cells, presence of the beta1(P228A) subunit reduced enhancement by barbiturates without affecting enhancement by diazepam or alfaxalone. Single channel recordings from alpha1beta1gamma2S and alpha1beta1(P228A)gamma2L GABARs showed similar channel kinetics, but beta-mutant containing receptors opened at lower GABA concentrations. We conclude that the beta1 subunit M1 segment proline affects the linkage between GABA binding and channel gating and is critical for barbiturate enhancement. Mutation of the M1 proline in the alpha1 subunit also inhibited receptor assembly.