Faculty Bibliography

How does coordinated activity between distinct brain regions implement a set of learning rules to sculpt information processing in a given neural circuit? Using interneuron cell-type-specific optical activation and pharmacogenetic silencing in vitro, we show that temporally precise pairing of direct entorhinal perforant path (PP) and hippocampal Schaffer collateral (SC) inputs to CA1 pyramidal cells selectively suppresses SC-associated perisomatic inhibition from cholecystokinin (CCK)-expressing interneurons. The CCK interneurons provide a surprisingly strong feedforward inhibitory drive to effectively control the coincident excitation of CA1 pyramidal neurons by convergent inputs. Thus, in-phase cortico-hippocampal activity provides a powerful heterosynaptic learning rule for long-term gating of information flow through the hippocampal excitatory macrocircuit by the silencing of the CCK inhibitory microcircuit.

The current report represents a further advancement of our previously reported technology termed Fluorescent transfer RNA (tRNA) for Translation Monitoring (FtTM), for monitoring of active global protein synthesis sites in single live cells. FtTM measures Forster resonance energy transfer (FRET) signals, generated when fluorescent tRNAs (fl-tRNAs), separately labeled as a FRET pair, occupy adjacent sites on the ribosome. The current technology, termed DiCodon Monitoring of Protein Synthesis (DiCoMPS), was developed for monitoring active synthesis of a specific protein. In DiCoMPS, specific fl-tRNA pair combinations are selected for transfection, based on the degree of enrichment of a dicodon sequence to which they bind in the mRNA of interest, relative to the background transcriptome of the cell in which the assay is performed. In this study, we used cells infected with the Epizootic Hemorrhagic Disease Virus 2-Ibaraki and measured, through DiCoMPS, the synthesis of the viral non-structural protein 3 (NS3), which is enriched in the AUA:AUA dicodon. fl-tRNA(Ile)UAU-generated FRET signals were specifically enhanced in infected cells, increased in the course of infection and were diminished on siRNA-mediated knockdown of NS3. Our results establish an experimental approach for the single-cell measurement of the levels of synthesis of a specific viral protein.

Monitoring protein synthesis is essential to our understanding of gene expression regulation, as protein abundance is thought to be predominantly controlled at the level of translation. Mass-spectrometric and RNA sequencing methods have been recently developed for investigating mRNA translation at a global level, but these still involve technical limitations and are not widely applicable. In this study, we describe a novel system-wide proteomic approach for direct monitoring of translation, termed puromycin-associated nascent chain proteomics (PUNCH-P), which is based on incorporation of biotinylated puromycin into newly synthesized proteins under cell-free conditions followed by streptavidin affinity purification and liquid chromatography-tandem mass spectrometry analysis. Using PUNCH-P, we measured cell cycle-specific fluctuations in synthesis for >5000 proteins in mammalian cells, identified proteins not previously implicated in cell cycle processes, and generated the first translational profile of a whole mouse brain. This simple and economical technique is broadly applicable to any cell type and tissue, enabling the identification and quantification of rapid proteome responses under various biological conditions.

A common single-nucleotide polymorphism (SNP) in the human brain-derived neurotrophic factor (BDNF) gene results in a Val66Met substitution in the BDNF prodomain region. This SNP is associated with alterations in memory and with enhanced risk to develop depression and anxiety disorders in humans. Here we show that the isolated BDNF prodomain is detected in the hippocampus and that it can be secreted from neurons in an activity-dependent manner. Using nuclear magnetic resonance spectroscopy and circular dichroism, we find that the prodomain is intrinsically disordered, and the Val66Met substitution induces structural changes. Surprisingly, application of Met66 (but not Val66) BDNF prodomain induces acute growth cone retraction and a decrease in Rac activity in hippocampal neurons. Expression of p75(NTR) and differential engagement of the Met66 prodomain to the SorCS2 receptor are required for this effect. These results identify the Met66 prodomain as a new active ligand, which modulates neuronal morphology.

The automated touchscreen operant chamber for rats and mice allows for the assessment of multiple cognitive domains within the same testing environment. This protocol presents the location discrimination (LD) task and the trial-unique delayed nonmatching-to-location (TUNL) task, which both assess memory for location. During these tasks, animals are trained to a predefined criterion during approximately 20-40 daily sessions. In LD sessions, touching the same location on the screen is rewarded on consecutive trials, followed by a reversal of location-reward contingencies. TUNL, a working memory task, requires animals to 'nonmatch' to a sample location after a delay. In both the LD and TUNL tasks, spatial similarity can be varied, allowing assessment of pattern separation ability, a function that is thought to be performed by the dentate gyrus (DG). These tasks are therefore particularly useful in animal models of hippocampal, and specifically DG, function, but they additionally permit discernment of changes in pattern separation from those in working memory.

This protocol details a subset of assays developed within the touchscreen platform to measure various aspects of executive function in rodents. Three main procedures are included: extinction, measuring the rate and extent of curtailing a response that was previously, but is no longer, associated with reward; reversal learning, measuring the rate and extent of switching a response toward a visual stimulus that was previously not, but has become, associated with reward (and away from a visual stimulus that was previously, but is no longer, rewarded); and the 5-choice serial reaction time (5-CSRT) task, gauging the ability to selectively detect and appropriately respond to briefly presented, spatially unpredictable visual stimuli. These protocols were designed to assess both complementary and overlapping constructs including selective and divided visual attention, inhibitory control, flexibility, impulsivity and compulsivity. The procedures comprise part of a wider touchscreen test battery assessing cognition in rodents with high potential for translation to human studies.

The present review article summarizes and expands upon the discussions that were initiated during a meeting of the Cognitive Neuroscience Treatment Research to Improve Cognition in Schizophrenia (CNTRICS; http://cntrics.ucdavis.edu) meeting. A major goal of the CNTRICS meeting was to identify experimental procedures and measures that can be used in laboratory animals to assess psychological constructs that are related to the psychopathology of schizophrenia. The issues discussed in this review reflect the deliberations of the Motivation Working Group of the CNTRICS meeting, which included most of the authors of this article as well as additional participants. After receiving task nominations from the general research community, this working group was asked to identify experimental procedures in laboratory animals that can assess aspects of reinforcement learning and motivation that may be relevant for research on the negative symptoms of schizophrenia, as well as other disorders characterized by deficits in reinforcement learning and motivation. The tasks described here that assess reinforcement learning are the Autoshaping Task, Probabilistic Reward Learning Tasks, and the Response Bias Probabilistic Reward Task. The tasks described here that assess motivation are Outcome Devaluation and Contingency Degradation Tasks and Effort-Based Tasks. In addition to describing such methods and procedures, the present article provides a working vocabulary for research and theory in this field, as well as an industry perspective about how such tasks may be used in drug discovery. It is hoped that this review can aid investigators who are conducting research in this complex area, promote translational studies by highlighting shared research goals and fostering a common vocabulary across basic and clinical fields, and facilitate the development of medications for the treatment of symptoms mediated by reinforcement learning and motivational deficits.

PURPOSE: Patients with medial temporal lobe epilepsy (MTLE) exhibit structural brain damage involving gray matter (GM) and white matter (WM). The mechanisms underlying tissue loss in MTLE are unclear and may be associated with a combination of seizure excitotoxicity and WM vulnerability. The goal of this study was to investigate whether late-myelinating WM tracts are more vulnerable to injury in MTLE compared with early myelinating tracts. METHODS: Diffusional kurtosis imaging scans were obtained from 25 patients with MTLE and from 36 matched healthy controls. Diffusion measures from regions of interest (ROIs) for both late- and early myelinating WM tracts were analyzed. Regional Z-scores were computed with respect to normal controls to compare WM in early myelinating tracts versus late-myelinating tracts. KEY FINDINGS: We observed that late-myelinating tracts exhibited a larger decrease in mean, axial, and radial kurtosis compared with early myelinating tracts. We also observed that the change in radial kurtosis was more pronounced in late-myelinating tracts ipsilateral to the side of seizure onset. SIGNIFICANCE: These results suggest a developmentally based preferential susceptibility of late-myelinating WM tracts to damage in MTLE. Brain injury in epilepsy may be due to the pathologic effects of seizures in combination with regional WM vulnerability.

There is emerging evidence that mild chronic hyponatremia (MCH), highly prevalent in the elderly and once considered asymptomatic, is a major independent risk factor for falls, fall-related fractures (independent of osteoporosis, age, and sex), impaired attention and gait, reductions in bone mineral density (BMD), and even death. Although research on MCH and bone health is emerging and ongoing, it has not been recognized in orthopedics. Orthopedic surgeons must be educated regarding the impact of hyponatremia on bone, as osteoporotic fractures have enormous socioeconomic consequences, and the problem will worsen. Orthopedic surgeons should also be included in research, in education, and in the establishment of diagnostic and treatment protocols. In this article, we review the current concepts of MCH and its impacts on the skeletal system.