Albert Chen

Coordinate Regulation of Motor Neuron Subtype Identity and Pan-Neuronal Properties by the bHLH Repressor Olig2

Within the developing vertebrate nervous system, the mechanisms that coordinate neuronal subtype identity with generic features of neuronal differentiation are poorly defined. We show here that a bHLH protein, Olig2, is expressed selectively by motor neuron progenitors and has a key role in specifying the subtype identity and pan-neuronal properties of developing motor neurons. The role of Olig2 in the specification of motor neuron subtype identity depends on regulatory interactions with progenitor homeodomain proteins, whereas its role in promoting pan-neuronal properties is associated with expression of another bHLH protein, Ngn2. Both aspects of Olig2 function appear to depend on its activity as a transcriptional repressor. Together, these studies show that Olig2 has a critical role in integrating diverse features of motor neuron differentiation in the developing spinal cord.

Graded activity of transcription factor Runx3 specifies the laminar termination pattern of sensory axons in the developing spinal cord

Different functional classes of dorsal root ganglion sensory neurons project their axons to distinct target zones within the developing spinal cord. To explore the mechanisms that link sensory neuron subtype identity and axonal projection pattern, we analyzed the roles of Runx and ETS transcription factors in the laminar targeting of sensory afferents. Gain- and loss-of-function studies in chick embryos reveal that the status of Runx3 expression is a major determinant of the dorso-ventral position of termination of proprioceptive and cutaneous sensory axons. In addition, the level of expression and/or activity of Runx3 in individual proprioceptive sensory neurons appears to specify whether their axons terminate in intermediate or ventral regions. Our findings suggest that the selectivity of Runx3 expression, and its level of activity, control sensory afferent targeting in the developing spinal cord.

TE-Averaged two-dimensional proton spectroscopic imaging of glutamate at 3 T

Glutamate and glutamine are important neurochemicals in the central nervous system and the neurotoxic properties of excess glutamate have been associated with several neurodegenerative diseases. The TE-Averaged PRESS technique has been shown by our group to detect an unobstructed glutamate signal at 3 T that is resolved from glutamine and NAA at 2.35 ppm. TE-Averaged PRESS therefore provides an unambiguous measurement of glutamate as well as other metabolites such as NAA, choline, creatine, and myo-inositol. In this study, we extend the single voxel TE-Averaged PRESS technique for two-dimensional (2D) spectroscopic imaging (TE-Averaged MRSI) to generate 2D glutamate maps. To facilitate TE-Averaged MRSI within a reasonable time, a fast encoding trajectory was used. This enabled rapid acquisition of TE-Averaged spectral arrays with good spectral bandwidth (977 Hz) and resolution (approximately 2 Hz). MRSI data arrays of 10 x 16 were acquired with 1.8 cm3 spatial resolution over a approximately 110 cm3 volume in a scan time of approximately 21 min. Two-dimensional metabolite maps were obtained with good SNR and clear differentiation in glutamate levels was observed between gray and white matter with significantly higher glutamate in gray matter relative to white matter as anticipated.

TrkB (Tropomyosin-Related Kinase B) Controls the Assembly and Maintenance of GABAergic Synapses in the Cerebellar Cortex

Inhibitory interneurons play a critical role in coordinating the activity of neural circuits. To explore the mechanisms that direct the organization of inhibitory circuits, we analyzed the involvement of tropomyosin-related kinase B (TrkB) in the assembly and maintenance of GABAergic inhibitory synapses between Golgi and granule cells in the mouse cerebellar cortex. We show that TrkB acts directly within each cell-type to regulate synaptic differentiation. TrkB is required not only for assembly, but also maintenance of these synapses and acts, primarily, by regulating the localization of synaptic constituents. Postsynaptically, TrkB controls the localization of a scaffolding protein, gephyrin, but acts at a step subsequent to the localization of a cell adhesion molecule, Neuroligin-2. Importantly, TrkB is required for the localization of an Ig superfamily cell adhesion molecule, Contactin-1, in Golgi and granule cells and the absence of Contactin-1 also results in deficits in inhibitory synaptic development. Thus, our findings demonstrate that TrkB controls the assembly and maintenance of GABAergic synapses and suggest that TrkB functions, in part, through promoting synaptic adhesion.

Glutamatergic axon-derived BDNF controls GABAergic synaptic differentiation in the cerebellum

To study mechanisms that regulate the construction of inhibitory circuits, we examined the role of brain-derived neurotrophic factor (BDNF) in the assembly of GABAergic inhibitory synapses in the mouse cerebellar cortex. We show that within the cerebellum, BDNF-expressing cells are restricted to the internal granular layer (IGL), but that the BDNF protein is present within mossy fibers which originate from cells located outside of the cerebellum. In contrast to deletion of TrkB, the cognate receptor for BDNF, deletion of Bdnf from cerebellar cell bodies alone did not perturb the localization of pre- or postsynaptic constituents at the GABAergic synapses formed by Golgi cell axons on granule cell dendrites within the IGL. Instead, we found that BDNF derived from excitatory mossy fiber endings controls their differentiation. Our findings thus indicate that cerebellar BDNF is derived primarily from excitatory neurons—precerebellar nuclei/spinal cord neurons that give rise to mossy fibers—and promotes GABAergic synapse formation as a result of release from axons. Thus, within the cerebellum the preferential localization of BDNF to axons enhances the specificity through which BDNF promotes GABAergic synaptic differentiation.

Design of cosine modulated very selective suppression pulses for MR spectroscopic imaging at 3T.

The advantages of using a 3 Tesla (T) scanner for MR spectroscopic imaging (MRSI) of brain tissue include improved spectral resolution and increased sensitivity. Very selective saturation (VSS) pulses are important for maximizing selectivity for PRESS MRSI and minimizing chemical shift misregistration by saturating signals from outside the selected region. Although three‐dimensional (3D) PRESS MRSI is able to provide excellent quality metabolic data for patients with brain tumors and has been shown to be important for defining tumor burden, the method is currently limited by how much of the anatomic lesion can be covered within a single examination. In this study we designed and implemented cosine modulated VSS pulses that were optimized for 3T MRSI acquisitions. This provided improved coverage and suppression of unwanted lipid signals with a smaller number of pulses. The use of the improved pulse sequence was validated in volunteer studies, and in clinical 3D MRSI exams of brain tumors. Magn Reson Med, 2009.

Fast 3D 1 H MRSI of the Corticospinal Tract in Pediatric Brain

To develop a 1H magnetic resonance spectroscopic imaging (MRSI) sequence that can be used to image infants/children at 3T and by combining it with diffusion tensor imaging (DTI) tractography, extract relevant metabolic information corresponding to the corticospinal tract (CST).

Distinct Activities of Tfap2A and Tfap2B in the Specification of GABAergic Interneurons in the Developing Cerebellum

GABAergic inhibitory neurons in the cerebellum are subdivided into Purkinje cells and distinct subtypes of interneurons from the same pool of progenitors, but the determinants of this diversification process are not well defined. To explore the transcriptional regulation of the development of cerebellar inhibitory neurons, we examined the role of Tfap2A and Tfap2B in the specification of GABAergic neuronal subtypes in mice. We show that Tfap2A and Tfap2B are expressed in inhibitory precursors during embryonic development and that their expression persists into adulthood. The onset of their expression follows Ptf1a and Olig2, key determinants of GABAergic neuronal fate in the cerebellum; and, their expression precedes Pax2, an interneuron-specific factor. Tfap2A is expressed by all GABAergic neurons, whereas Tfap2B is selectively expressed by interneurons. Genetic manipulation via in utero electroporation (IUE) reveals that Tfap2B is necessary for interneuron specification and is capable of suppressing the generation of excitatory cells. Tfap2A, but not Tfap2B, is capable of inducing the generation of interneurons when misexpressed in the ventricular neuroepithelium. Together, our results demonstrate that the differential expression of Tfap2A and Tfap2B defines subtypes of GABAergic neurons and plays specific, but complementary roles in the specification of interneurons in the developing cerebellum.

Conditional deletion of Cadherin 13 perturbs Golgi cells and disrupts social and cognitive behaviors.

Inhibitory interneurons mediate the gating of synaptic transmission and modulate the activities of neural circuits. Disruption of the function of inhibitory networks in the forebrain is linked to impairment of social and cognitive behaviors, but the involvement of inhibitory interneurons in the cerebellum has not been assessed. We found that Cadherin 13 (Cdh13), a gene implicated in autism spectrum disorder and attention‐deficit hyperactivity disorder, is specifically expressed in Golgi cells within the cerebellar cortex. To assess the function of Cdh13 and utilize the manipulation of Cdh13 expression in Golgi cells as an entry point to examine cerebellar‐mediated function, we generated mice carrying Cdh13‐floxed alleles and conditionally deleted Cdh13 with GlyT2::Cre mice. Loss of Cdh13 results in a decrease in the expression/localization of GAD67 and reduces spontaneous inhibitory postsynaptic current (IPSC) in cerebellar Golgi cells without disrupting spontaneous excitatory postsynaptic current (EPSC). At the behavioral level, loss of Cdh13 in the cerebellum, piriform cortex and endopiriform claustrum have no impact on gross motor coordination or general locomotor behaviors, but leads to deficits in cognitive and social abilities. Mice lacking Cdh13 exhibit reduced cognitive flexibility and loss of preference for contact region concomitant with increased reciprocal social interactions. Together, our findings show that Cdh13 is critical for inhibitory function of Golgi cells, and that GlyT2::Cre‐mediated deletion of Cdh13 in non‐executive centers of the brain, such as the cerebellum, may contribute to cognitive and social behavioral deficits linked to neurological disorders.

Precision of Discrete and Rhythmic Forelimb Movements Requires a Distinct Neuronal Subpopulation in the Interposed Anterior Nucleus

The deep cerebellar nuclei (DCN) represent output channels of the cerebellum, and they transmit integrated sensorimotor signals to modulate limb movements. But the functional relevance of identifiable neuronal subpopulations within the DCN remains unclear. Here, we examine a genetically tractable population of neurons in the mouse interposed anterior nucleus (IntA). We show that these neurons represent a subset of glutamatergic neurons in the IntA and constitute a specific element of an internal feedback circuit within the cerebellar cortex and cerebello-thalamo-cortical pathway associated with limb control. Ablation and optogenetic stimulation of these neurons disrupt efficacy of skilled reach and locomotor movement and reveal that they control positioning and timing of the forelimb and hindlimb. Together, our findings uncover the function of a distinct neuronal subpopulation in the deep cerebellum and delineate the anatomical substrates and kinematic parameters through which it modulates precision of discrete and rhythmic limb movements.