Layer- and cell type-selective co-transmission by a basal forebrain cholinergic projection to the olfactory bulb.

Cholinergic neurons in the basal forebrain project heavily to the main olfactory bulb, the first processing station in the olfactory pathway. The projections innervate multiple layers of the main olfactory bulb and strongly influence odor discrimination, detection, and learning. The precise underlying circuitry of this cholinergic input to the main olfactory bulb remains unclear, however. Here, we identify a specific basal forebrain cholinergic projection that innervates select neurons concentrated in the internal plexiform layer of the main olfactory bulb. Optogenetic activation of this projection elicits monosynaptic nicotinic and GABAergic currents in glomerular layer-projecting interneurons. Additionally, we show that the projection co-expresses markers for GABAergic neurotransmission. The data thus implicate neurotransmitter co-transmission in the basal forebrain regulation of this inhibitory olfactory microcircuit.Cholinergic neurons innervate multiple layers in the main olfactory bulb but the precise circuitry of this input is not known. Here the authors show that VGLUT3+ cholinergic neurons selectively innervate deep short axon cells in specific layers and elicit robust monosynaptic GABAergic and nicotinic postsynaptic currents.

Loss of VGLUT3 Produces Circadian-Dependent Hyperdopaminergia and Ameliorates Motor Dysfunction and l-Dopa-Mediated Dyskinesias in a Model of Parkinson's Disease.

The striatum is essential for many aspects of mammalian behavior, including motivation and movement, and is dysfunctional in motor disorders such as Parkinson's disease. The vesicular glutamate transporter 3 (VGLUT3) is expressed by striatal cholinergic interneurons (CINs) and is thus well positioned to regulate dopamine (DA) signaling and locomotor activity, a canonical measure of basal ganglia output. We now report that VGLUT3 knock-out (KO) mice show circadian-dependent hyperlocomotor activity that is restricted to the waking cycle and is due to an increase in striatal DA synthesis, packaging, and release. Using a conditional VGLUT3 KO mouse, we show that deletion of the transporter from CINs, surprisingly, does not alter evoked DA release in the dorsal striatum or baseline locomotor activity. The mice do, however, display changes in rearing behavior and sensorimotor gating. Elevation of DA release in the global KO raised the possibility that motor deficits in a Parkinson's disease model would be reduced. Remarkably, after a partial 6-hydroxydopamine (6-OHDA)-mediated DA depletion (∼70% in dorsal striatum), KO mice, in contrast to WT mice, showed normal motor behavior across the entire circadian cycle. l-3,4-dihydroxyphenylalanine-mediated dyskinesias were also significantly attenuated. These findings thus point to new mechanisms to regulate basal ganglia function and potentially treat Parkinson's disease and related disorders. SIGNIFICANCE STATEMENT: Dopaminergic signaling is critical for both motor and cognitive functions in the mammalian nervous system. Impairments, such as those found in Parkinson's disease patients, can lead to severe motor deficits. Vesicular glutamate transporter 3 (VGLUT3) loads glutamate into secretory vesicles for neurotransmission and is expressed by discrete neuron populations throughout the nervous system. Here, we report that the absence of VGLUT3 in mice leads to an upregulation of the midbrain dopamine system. Remarkably, in a Parkinson's disease model, the mice show normal motor behavior. They also show fewer abnormal motor behaviors (dyskinesias) in response to l-3,4-dihydroxyphenylalanine, the principal treatment for Parkinson's disease. The work thus suggests new avenues for the development of novel treatment strategies for Parkinson's disease and potentially other basal-ganglia-related disorders.

VGLUT3 does not synergize GABA/glycine release during functional refinement of an inhibitory auditory circuit.

The vesicular glutamate transporter 3 (VGLUT3) is expressed at several locations not normally associated with glutamate release. Although the function of this protein has been generally elusive, when expressed in non-glutamatergic synaptic terminals, VGLUT3 can not only allow glutamate co-transmission but also synergize the action of non-glutamate vesicular transporters. Interestingly, in the immature glycinergic projection between the medial nucleus of the trapezoid body (MNTB) and the lateral superior olive (LSO) of auditory brainstem, the transient early expression of VGLUT3 is required for normal developmental refinement. It has however been unknown whether the primary function of VGLUT3 in development of these inhibitory synapses is to enable glutamate release or to promote loading of inhibitory neurotransmitter through vesicular synergy. Using tissue from young mice in which Vglut3 had been genetically deleted, we evaluated inhibitory neurotransmission in the MNTB-LSO pathway. Our results show, in contrast to what has been seen at adult synapses, that VGLUT3 expression has little or no effect on vesicular synergy at the immature glycinergic synapse of brainstem. This finding supports the model that the primary function of increased VGLUT3 expression in the immature auditory brainstem is to enable glutamate release in a developing inhibitory circuit.

Activation of neural cell fate programs toward direct conversion of adult human fibroblasts into tri-potent neural progenitors using OCT-4.

Several transcription factors and methods have been used to convert fibroblasts directly to neural fate and have provided insights into molecular mechanisms as to how each of these required factors orchestrate neural fate conversion. Here, we provide evidence and detailed characterization of the direct conversion process of primary adult human fibroblasts (hFib) to neural progenitor cells (NPC) using OCT4 alone. Factors previously associated with neural cell fate conversion were induced during hFib-NPC(OCT-4) generation, where OCT-4 alone was sufficient to induce neural fate conversion without the use of promiscuous small-molecule manipulation. Human Fib-NPC(OCT-4) proliferate, express neural stem/progenitor markers, and possess developmental potential that gives rise to all three major subtypes of neural cells: astrocytes, oligodendrocytes, and neurons with functional capacity. We propose a de-convoluted reprogramming approach for neural fate conversion in which OCT4 is sufficient for inducing neural conversion from hFib for disease modeling as well as the fundamental study of early neural fate induction.

Pre- and postsynaptic properties of glutamatergic transmission in the immature inhibitory MNTB-LSO pathway.

The lateral superior olive (LSO) integrates excitatory inputs driven by sound arriving at the ipsilateral ear with inhibitory inputs driven by sound arriving at the contralateral ear in order to compute interaural intensity differences needed for localizing high-frequency sound sources. Specific mechanisms necessary for developmental refinement of the inhibitory projection, which arises from the medial nucleus of the trapezoid body (MNTB), have only been partially deciphered. The demonstration that immature MNTB-LSO synapses release glutamate has led to a model in which early glutamate neurotransmission plays a major role in inhibitory plasticity. We used whole cell electrophysiology in acute auditory brain stem slices of neonatal rats to examine glutamatergic transmission in the developing MNTB-LSO pathway. Unexpectedly, AMPA receptor (AMPAR)-mediated responses were prevalent at the earliest ages. We found a salient developmental profile for NMDA receptor (NMDAR) activation, described both by the proportion of total glutamate current and by current durations, and we found evidence for distinct release probabilities for GABA/glycine and glutamate in the MNTB-LSO pathway. The developmental profile of NMDAR is consistent with the possibility that the inhibitory MNTB-LSO pathway experiences a sensitive period, driven by cochlear activity and mediated by GluN2B-containing NMDARs, between postnatal days 3 and 9. Differing neurotransmitter release probabilities could allow the synapse to switch between GABA/glycinergic transmission and mixed glutamate/GABA/glycinergic transmission in response to changing patterns of spiking activity.

Functional refinement in the projection from ventral cochlear nucleus to lateral superior olive precedes hearing onset in rat.

Principal neurons of the lateral superior olive (LSO) compute the interaural intensity differences necessary for localizing high-frequency sounds. To perform this computation, the LSO requires precisely tuned, converging excitatory and inhibitory inputs that are driven by the two ears and that are matched for stimulus frequency. In rodents, the inhibitory inputs, which arise from the medial nucleus of the trapezoid body (MNTB), undergo extensive functional refinement during the first postnatal week. Similar functional refinement of the ascending excitatory pathway, which arises in the anteroventral cochlear nucleus (AVCN), has been assumed but has not been well studied. Using whole-cell voltage clamp in acute brainstem slices of neonatal rats, we examined developmental changes in input strength and pre- and post-synaptic properties of the VCN-LSO pathway. A key question was whether functional refinement in one of the two major input pathways might precede and then guide refinement in the opposite pathway. We find that elimination and strengthening of VCN inputs to the LSO occurs over a similar period to that seen for the ascending inhibitory (MNTB-LSO) pathway. During this period, the fractional contribution provided by NMDA receptors (NMDARs) declines while the contribution from AMPA receptors (AMPARs) increases. In the NMDAR-mediated response, GluN2B-containing NMDARs predominate in the first postnatal week and decline sharply thereafter. Finally, the progressive decrease in paired-pulse depression between birth and hearing onset allows these synapses to follow progressively higher frequencies. Our data are consistent with a model in which the excitatory and inhibitory projections to LSO are functionally refined in parallel during the first postnatal week, and they further suggest that GluN2B-containing NMDARs may mediate early refinement in the VCN-LSO pathway.