Unique molecular features and cellular responses differentiate two populations of motor cortical layer 5b neurons in a preclinical model of ALS.

Many neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS), lead to the selective degeneration of discrete cell types in the CNS despite the ubiquitous expression of many genes linked to disease. Therapeutic advancement depends on understanding the unique cellular adaptations that underlie pathology of vulnerable cells in the context of disease-causing mutations. Here, we employ bacTRAP molecular profiling to elucidate cell type-specific molecular responses of cortical upper motor neurons in a preclinical ALS model. Using two bacTRAP mouse lines that label distinct vulnerable or resilient projection neuron populations in motor cortex, we show that the regulation of oxidative phosphorylation (Oxphos) pathways is a common response in both cell types. However, differences in the baseline expression of genes involved in Stem and the handling of reactive oxygen species likely lead to the selective degeneration of the vulnerable cells. These results provide a framework to identify cell-type-specific processes in neurodegenerative disease.

Dopamine neuron dependent behaviors mediated by glutamate cotransmission.

Dopamine neurons in the ventral tegmental area use glutamate as a cotransmitter. To elucidate the behavioral role of the cotransmission, we targeted the glutamate-recycling enzyme glutaminase (gene Gls1). In mice with a dopamine transporter (Slc6a3)-driven conditional heterozygous (cHET) reduction of Gls1 in their dopamine neurons, dopamine neuron survival and transmission were unaffected, while glutamate cotransmission at phasic firing frequencies was reduced, enabling a selective focus on the cotransmission. The mice showed normal emotional and motor behaviors, and an unaffected response to acute amphetamine. Strikingly, amphetamine sensitization was reduced and latent inhibition potentiated. These behavioral effects, also seen in global GLS1 HETs with a schizophrenia resilience phenotype, were not seen in mice with an Emx1-driven forebrain reduction affecting most brain glutamatergic neurons. Thus, a reduction in dopamine neuron glutamate cotransmission appears to mediate significant components of the GLS1 HET schizophrenia resilience phenotype, and glutamate cotransmission appears to be important in attribution of motivational salience.

Rapid Molecular Profiling of Defined Cell Types Using Viral TRAP.

Translational profiling methodologies enable the systematic characterization of cell types in complex tissues, such as the mammalian brain, where neuronal isolation is exceptionally difficult. Here, we report a versatile strategy for profiling CNS cell types in a spatiotemporally restricted fashion by engineering a Cre-dependent adeno-associated virus expressing an EGFP-tagged ribosomal protein (AAV-FLEX-EGFPL10a) to access translating mRNAs by translating ribosome affinity purification (TRAP). We demonstrate the utility of this AAV to target a variety of genetically and anatomically defined neural populations expressing Cre recombinase and illustrate the ability of this viral TRAP (vTRAP) approach to recapitulate the molecular profiles obtained by bacTRAP in corticothalamic neurons across multiple serotypes. Furthermore, spatially restricting adeno-associated virus (AAV) injections enabled the elucidation of regional differences in gene expression within this cell type. Altogether, these results establish the broad applicability of the vTRAP strategy for the molecular dissection of any CNS or peripheral cell type that can be engineered to express Cre.