Faculty Bibliography

Down syndrome (DS) is the most prevalent form of mental retardation caused by genetic abnormalities in humans. This has been successfully modeled in mice to generate the Ts65Dn mouse, a genetic model of DS. This transgenic mouse model shares a number of physical and functional abnormalities with people with DS, including changes in the structure and function of neuronal circuits. Significant abnormalities in noradrenergic (NE-ergic) afferents from the locus coeruleus to the hippocampus, as well as deficits in NE-ergic neurotransmission are detected in these animals. In the current study we characterized in detail the behavioral phenotype of Ts65Dn mice, in addition to using pharmacological tools for identification of target receptors mediating the learning and memory deficits observed in this model of DS. We undertook a comprehensive approach to mouse phenotyping using a battery of standard and novel tests encompassing: (i) locomotion (Activity Chamber, PhenoTyper, and CatWalk), (ii) learning and memory (spontaneous alternation, delayed matching-to-place water maze, fear conditioning, and Intellicage), and (iii) social behavior. Ts65Dn mice showed increased locomotor activity in novel and home cage environments. There were significant and reproducible deficits in learning and memory tests including spontaneous alternation, delayed matching-to-place water maze, Intellicage place avoidance and contextual fear conditioning. Although Ts65Dn mice showed no deficit in sociability in the 3-chamber test, a marked impairment in social memory was detected. Xamoterol, a beta1-adrenergic receptor (beta1-ADR) agonist, effectively restored the memory deficit in contextual fear conditioning, spontaneous alternation and novel object recognition. These behavioral improvements were reversed by betaxolol, a selective beta1-ADR antagonist. In conclusion, our results demonstrate that this mouse model of Down syndrome displays cognitive deficits which are mediated by an imbalance in the noradrenergic system. In this experimental model of Down syndrome a selective activation of beta1-ADR does restore some of these behavioral deficits. Further mechanistic studies will be needed to investigate the failure of noradrenergic system and the role of beta1-ADR in cognitive deficit and pathogenesis of DS in people. Restoring NE neurotransmission or a selective activation of beta1)-ADR needs to be further investigated for the development of any potential therapeutic strategy for symptomatic relief of memory deficit in DS. Furthermore, due to the significant involvement of noradrenergic system in the cardiovascular function further safety and translational studies will be needed to ensure the safety and efficacy of this approach

Prolonged blockade of AMPA-type glutamate receptors in hippocampal neuron cultures leads to homeostatic enhancements of pre- and postsynaptic function that appear correlated at individual synapses, suggesting some form of transsynaptic coordination. The respective modifications are important for overall synaptic strength but their interrelationship, dynamics, and molecular underpinnings are unclear. Here we demonstrate that adaptation begins postsynaptically but is ultimately communicated to presynaptic terminals and expressed as an accelerated turnover of synaptic vesicles. Critical postsynaptic modifications occur over hours, but enable retrograde communication within minutes once AMPA receptor (AMPAR) blockade is removed, causing elevation of both spontaneous and evoked vesicle fusion. The retrograde signaling does not require spiking activity and can be interrupted by NBQX, philanthotoxin, postsynaptic BAPTA, or external sequestration of BDNF, consistent with the acute release of retrograde messenger, triggered by postsynaptic Ca(2+) elevation via Ca(2+)-permeable AMPARs

GABA, the principal inhibitory neurotransmitter in the adult brain, has a parallel inhibitory role in the immune system. We demonstrate that immune cells synthesize GABA and have the machinery for GABA catabolism. Antigen-presenting cells (APCs) express functional GABA receptors and respond electrophysiologically to GABA. Thus, the immune system harbors all of the necessary constituents for GABA signaling, and GABA itself may function as a paracrine or autocrine factor. These observations led us to ask further whether manipulation of the GABA pathway influences an animal model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). Increasing GABAergic activity ameliorates ongoing paralysis in EAE via inhibition of inflammation. GABAergic agents act directly on APCs, decreasing MAPK signals and diminishing subsequent adaptive inflammatory responses to myelin proteins

Miniature synaptic events, resulting from spontaneous presynaptic release of neurotransmitter in the absence of an action potential, are often used to assay neural connectivity and are thought to play a pivotal role in the development and maintenance of neuronal circuits. In this issue of Neuron, Trigo et al. identify a new class of miniature synaptic event, called 'preminis,' that originate from and are subsequently detected by the presynaptic terminals of GABAergic neurons in the molecular layer of cerebellum. Remarkably, these events easily outnumber conventional minis. Their restriction to a relatively narrow time window (<15 days after birth) is a clue that they may play a critical role in wiring up interneurons within the developing cerebellar circuitry

Synaptic transmission at CNS synapses is often mediated by joint actions of multiple Ca(2+) channel subtypes, most prominently, P/Q- and N-type. We have proposed that P/Q-type Ca(2+) channels saturate type-preferring slots at presynaptic terminals, which impose a ceiling on the synaptic efficacy of the channels. To test for analogous interactions for presynaptic N-type Ca(2+) channels, we overexpressed their pore-forming Ca(V)2.2 subunit in cultured mouse hippocampal neurons, recorded excitatory synaptic transmission from transfected cells, and dissected the contributions of N-, P/Q-, and R-type channels with subtype-specific blockers. Overexpression of Ca(V)2.2 did not increase the absolute size of the EPSC even though somatic N-type current was augmented by severalfold. Thus, the strength of neurotransmission is saturated with regard to levels of Ca(2+) channel expression for both N-type and P/Q-type channels. Overexpression of Ca(2+)-impermeable Ca(V)2.2 subunits decreased EPSC size, corroborating competition for channel slots. Striking asymmetries between N- and P/Q-type channels emerged when their relative contributions were compared with channel overexpression. Overexpressed N-type channels could competitively displace P/Q-type channels from P/Q-preferring slots and take over the role of supporting transmission. The converse was not found with overexpression of P/Q-type channels, regardless of their C-terminal domain. We interpret these findings in terms of two different kinds of presynaptic slots at excitatory synapses, one accepting N-type channels but rejecting P/Q-type (N(specific)) and the other preferring P/Q-type but also accepting N-type (PQ(preferring)). The interaction between channels and slots governs the respective contributions of multiple channel types to neurotransmission and, in turn, the ability of transmission to respond to various stimulus patterns and neuromodulators

Vesicular secretion of neurotransmitter is essential for neuronal communication. Kiss-and-run is a mode of membrane fusion and retrieval without the full collapse of the vesicle into the plasma membrane and de novo regeneration. The importance of kiss-and-run during efficient neurotransmission has remained in doubt. We developed an approach for loading individual synaptic vesicles with single quantum dots. Their size and pH-dependent photoluminescence change allowed us to distinguish kiss-and-run from full-collapse fusion and to track single vesicles through multiple rounds of kiss-and-run and reuse, without perturbing vesicle cycling. Kiss-and-run dominated at the beginning of stimulus trains, reflecting the preference of vesicles with high release probability. Its incidence was increased by rapid firing, a response appropriate to shape the kinetics of neurotransmission during a wide range of firing patterns

N-methyl-D-aspartate receptors (NMDARs) play important functions in neural development. NR2B is the predominant NR2 subunit of NMDAR in the developing brain. Here we use mosaic analysis with double markers (MADM) to knock out NR2B in isolated single cells and analyze its cell-autonomous function in dendrite development. NR2B mutant dentate gyrus granule cells (dGCs) and barrel cortex layer 4 spiny stellate cells (bSCs) have similar dendritic growth rates, total length, and branch number as control cells. However, mutant dGCs maintain supernumerary primary dendrites resulting from a pruning defect. Furthermore, while control bSCs restrict dendritic growth to a single barrel, mutant bSCs maintain dendritic growth in multiple barrels. Thus, NR2B functions cell autonomously to regulate dendrite patterning to ensure that sensory information is properly represented in the cortex. Our study also indicates that molecular mechanisms that regulate activity-dependent dendrite patterning can be separated from those that control general dendrite growth and branching

Communication between cell surface proteins and the nucleus is integral to many cellular adaptations. In the case of ion channels in excitable cells, the dynamics of signaling to the nucleus are particularly important because the natural stimulus, surface membrane depolarization, is rapidly pulsatile. To better understand excitation-transcription coupling we characterized the dependence of cAMP response element-binding protein phosphorylation, a critical step in neuronal plasticity, on the level and duration of membrane depolarization. We find that signaling strength is steeply dependent on depolarization, with sensitivity far greater than hitherto recognized. In contrast, graded blockade of the Ca(2+) channel pore has a remarkably mild effect, although some Ca(2+) entry is absolutely required. Our data indicate that Ca(2+)/CaM-dependent protein kinase II acting near the channel couples local Ca(2+) rises to signal transduction, encoding the frequency of Ca(2+) channel openings rather than integrated Ca(2+) flux-a form of digital logic

Calcium entry into excitable cells is an important physiological signal, supported by and highly sensitive to the activity of voltage-gated Ca2+ channels. After membrane depolarization, Ca2+ channels first open but then undergo various forms of negative feedback regulation including voltage- and calcium-dependent inactivation (VDI and CDI, respectively). Inactivation of Ca2+ channel activity is perturbed in a rare yet devastating disorder known as Timothy syndrome (TS), whose features include autism or autism spectrum disorder along with severe cardiac arrhythmia and developmental abnormalities. Most cases of TS arise from a sporadic single nucleotide change that generates a mutation (G406R) in the pore-forming subunit of the L-type Ca2+ channel Ca(V)1.2. We found that the TS mutation powerfully and selectively slows VDI while sparing or possibly speeding the kinetics of CDI. The deceleration of VDI was observed when the L-type channels were expressed with beta1 subunits prominent in brain, as well as beta2 subunits of importance for the heart. Dissociation of VDI and CDI was further substantiated by measurements of Ca2+ channel gating currents and by analysis of another channel mutation (I1624A) that hastens VDI, acting upstream of the step involving Gly406. As highlighted by the TS mutation, CDI does not proceed to completeness but levels off at approximately 50%, consistent with a change in gating modes and not an absorbing inactivation process. Thus, the TS mutation offers a unique perspective on mechanisms of inactivation as well as a promising starting point for exploring the underlying pathophysiology of autism

Spinocerebellar ataxia type 6 (SCA6) is a neurodegenerative disorder caused by CAG repeat expansions within the voltage-gated calcium (Ca(V)) 2.1 channel gene. It remains controversial whether the mutation exerts neurotoxicity by changing the function of Ca(V)2.1 channel or through a gain-of-function mechanism associated with accumulation of the expanded polyglutamine protein. We generated three strains of knockin (KI) mice carrying normal, expanded, or hyperexpanded CAG repeat tracts in the Cacna1a locus. The mice expressing hyperexpanded polyglutamine (Sca6(84Q)) developed progressive motor impairment and aggregation of mutant Ca(V)2.1 channels. Electrophysiological analysis of cerebellar Purkinje cells revealed similar Ca(2+) channel current density among the three KI models. Neither voltage sensitivity of activation nor inactivation was altered in the Sca6(84Q) neurons, suggesting that expanded CAG repeat per se does not affect the intrinsic electrophysiological properties of the channels. The pathogenesis of SCA6 is apparently linked to an age-dependent process accompanied by accumulation of mutant Ca(V)2.1 channels