An intellectual-disability-associated mutation of the transcriptional regulator NACC1 impairs glutamatergic neurotransmission.

Advances in genome sequencing technologies have favored the identification of rare de novo mutations linked to neurological disorders in humans. Recently, a de novo autosomal dominant mutation in NACC1 was identified (NM_052876.3: c.892C > T, NP_443108.1; p.Arg298Trp), associated with severe neurological symptoms including intellectual disability, microcephaly, and epilepsy. As NACC1 had never before been associated with neurological diseases, we investigated how this mutation might lead to altered brain function. We examined neurotransmission in autaptic glutamatergic mouse neurons expressing the murine homolog of the human mutant NACC1, i.e., Nacc1-R284W. We observed that expression of Nacc1-R284W impaired glutamatergic neurotransmission in a cell-autonomous manner, likely through a dominant negative mechanism. Furthermore, by screening for Nacc1 interaction targets in the brain, we identified SynGAP1, GluK2A, and several SUMO E3 ligases as novel Nacc1 interaction partners. At a biochemical level, Nacc1-R284W exhibited reduced binding to SynGAP1 and GluK2A, and also showed greatly increased SUMOylation. Ablating the SUMOylation of Nacc1-R284W partially restored its interaction with SynGAP1 but did not restore binding to GluK2A. Overall, these data indicate a role for Nacc1 in regulating glutamatergic neurotransmission, which is substantially impaired by the expression of a disease-associated Nacc1 mutant. This study provides the first functional insights into potential deficits in neuronal function in patients expressing the de novo mutant NACC1 protein.

A fluorescent nanosensor paint detects dopamine release at axonal varicosities with high spatiotemporal resolution.

The neurotransmitter dopamine (DA) controls multiple behaviors and is perturbed in several major brain diseases. DA is released from large populations of specialized structures called axon varicosities. Determining the DA release mechanisms at such varicosities is essential for a detailed understanding of DA biology and pathobiology but has been limited by the low spatial resolution of DA detection methods. We used a near-infrared fluorescent DA nanosensor paint, adsorbed nanosensors detecting release of dopamine (AndromeDA), to detect DA secretion from cultured murine dopaminergic neurons with high spatial and temporal resolution. We found that AndromeDA detects discrete DA release events and extracellular DA diffusion and observed that DA release varies across varicosities. To systematically detect DA release hotspots, we developed a machine learning­based analysis tool. AndromeDA permitted the simultaneous visualization of DA release for up to 100 dopaminergic varicosities, showing that DA release hotspots are heterogeneous and occur at only ∼17% of all varicosities, indicating that many varicosities are functionally silent. Using AndromeDA, we determined that DA release requires Munc13-type vesicle priming proteins, validating the utility of AndromeDA as a tool to study the molecular and cellular mechanism of DA secretion.

Imaging of Monoamine Neurotransmitters with Fluorescent Nanoscale Sensors.

Cells use biomolecules to convey information. For instance, neurons communicate by releasing chemicals called neurotransmitters, including several monoamines. The information transmitted by neurons is, in part, coded in the type and amount of neurotransmitter released, the spatial distribution of release sites, the frequency of release events, and the diffusion range of the neurotransmitter. Therefore, quantitative information about neurotransmitters at the (sub)cellular level with high spatiotemporal resolution is needed to understand how complex cellular networks function. So far, various analytical methods have been developed and used to detect neurotransmitter secretion from cells. However, each method has limitations with respect to chemical, temporal and spatial resolution. In this review, we focus on emerging methods for optical detection of neurotransmitter release and discuss fluorescent sensors/probes for monoamine neurotransmitters such as dopamine and serotonin. We focus on the latest advances in near infrared fluorescent carbon nanotube-based sensors and engineered fluorescent proteins for monoamine imaging, which provide high spatial and temporal resolution suitable for examining the release of monoamines from cells in cellular networks.