Classes Of Interneurons Research Articles

Functional, molecular and morphological heterogeneity of superficial interneurons in the larval zebrafish tectum.

The superficial interneurons, SINs, of the zebrafish tectum, have been implicated in a range of visual functions, including size discrimination, directional selectivity, and looming-evoked escape. This raises the question if SIN subpopulations, despite their morphological similarities and shared anatomical position in the retinotectal processing stream, carry out diverse, task-specific functions in visual processing, or if they have simple tuning properties in common. Here we have further characterized the SINs through functional imaging, electrophysiological recordings, and neurotransmitter typing in two transgenic lines, the widely used Gal4s1156t and the recently reported LCRRH2-RH2-2:GFP. We found that about a third of the SINs strongly responded to changes in whole-field light levels, with a strong preference for OFF over ON stimuli. Interestingly, individual SINs were selectively tuned to a diverse range of narrow luminance decrements. Overall responses to whole-field luminance steps did not vary with the position of the SIN cell body along the depth of the tectal neuropil or with the orientation of its neurites. We ruled out the possibility that intrinsic photosensitivity of Gal4s1156t+ SINs contribute to the measured visual responses. We found that, while most SINs express GABAergic markers, a substantial minority express an excitatory neuronal marker, the vesicular glutamate transporter, expanding the possible roles of SIN function in the tectal circuitry. In conclusion, SINs represent a molecularly, morphologically, and functionally heterogeneous class of interneurons, with subpopulations that detect a range of specific visual features, to which we have now added narrow luminance decrements.

Sustained Activation of PV+ Interneurons in Core Auditory Cortex Enables Robust Divisive Gain Control for Complex and Naturalistic Stimuli.

Sensory cortices must flexibly adapt their operations to internal states and external requirements. Sustained modulation of activity levels in different inhibitory interneuron populations may provide network-level mechanisms for adjustment of sensory cortical processing on behaviorally relevant timescales. However, understanding of the computational roles of inhibitory interneuron modulation has mostly been restricted to effects at short timescales, through the use of phasic optogenetic activation and transient stimuli. Here, we investigated how modulation of inhibitory interneurons affects cortical computation on longer timescales, by using sustained, network-wide optogenetic activation of parvalbumin-positive interneurons (the largest class of cortical inhibitory interneurons) to study modulation of auditory cortical responses to prolonged and naturalistic as well as transient stimuli. We found highly conserved spectral and temporal tuning in auditory cortical neurons, despite a profound reduction in overall network activity. This reduction was predominantly divisive, and consistent across simple, complex, and naturalistic stimuli. A recurrent network model with power-law input-output functions replicated our results. We conclude that modulation of parvalbumin-positive interneurons on timescales typical of sustained neuromodulation may provide a means for robust divisive gain control conserving stimulus representations.

In situ labeling of non-accommodating interneurons based on metabolic rates

Maintaining high frequency firing of narrow action potentials puts a large metabolic load on fast spiking (FS), perisomatic-inhibitory interneurons compared to their slow-spiking, dendrite targeting counterparts. Although the relationship of action potential (AP) firing and metabolism is firmly established, there is no single method to differentiate interneurons in situ based on their firing properties.In this study, we explore a novel strategy to easily identify the metabolically active FS cells among different classes of interneurons.We found that the oxidation of the fluorescent free radical marker 2,7-dichlorodihydrofluorescein (H2DCF) preferentially occurs in interneurons both in slice cultures and acute brain slices. Despite their morphological heterogeneity, almost all DCF-positive (DCF+) neurons belonged to the cluster of non-accommodating FS interneurons. Furthermore, all FS interneurons expressing parvalbumin (PV) both in slice cultures and in acute slices from tdTomato-PVCre transgenic mice were also DCF+. However, only half of the recorded DCF + cells were also PV+, indicating that H2DCF-oxidation occurs in different interneuron classes characterized by non-accomodating AP-firing. Comprehensively enhancing spontaneous neuronal activity led to mitochondrial oxidation of DCF in pyramidal cells as well as interneurons, suggesting that the apparent selectivity towards interneurons represents differences in the underlying metabolic load.While radical-scavenging, inhibition of APs or NO-synthesis, and iron chelation had no effect on the staining pattern, exposure to the complex-I inhibitor, rotenone, prevented interneuronal DCF accumulation. We conclude that H2DCF oxidation is independent of free radicals but correlates with the intensive oxidative energy metabolism and high mitochondrial mass in interneurons sharing the non-accommodating FS phenotype.

Large-Scale 3D Two-Photon Imaging of Molecularly Identified CA1 Interneuron Dynamics in Behaving Mice

Cortical computations are critically reliant on their local circuit, GABAergic cells. In the hippocampus, a large body of work has identified an unprecedented diversity of GABAergic interneurons with pronounced anatomical, molecular, and physiological differences. Yet little is known about the functional properties and activity dynamics of the major hippocampal interneuron classes in behaving animals. Here we use fast, targeted, three-dimensional (3D) two-photon calcium imaging coupled with immunohistochemistry-based molecular identification to retrospectively map invivo activity onto multiple classes of interneurons in the mouse hippocampal area CA1 during head-fixed exploration and goal-directed learning. We find examples of preferential subtype recruitment with quantitative differences in response properties and feature selectivity during key behavioral tasks and states. These results provide new insights into the collective organization of local inhibitory circuits supporting navigational and mnemonic functions of the hippocampus.

Generation of Sharp Wave-Ripple Events by Disinhibition

Sharp wave-ripple complexes (SWRs) are hippocampal network phenomena involved in memory consolidation. To date, the mechanisms underlying their occurrence remain obscure. Here, we show how the interactions between pyramidal cells, parvalbumin-positive (PV+) basket cells, and an unidentified class of anti-SWR interneurons can contribute to the initiation and termination of SWRs. Using a biophysically constrained model of a network of spiking neurons and a rate-model approximation, we demonstrate that SWRs emerge as a result of the competition between two interneuron populations and the resulting disinhibition of pyramidal cells. Our models explain how the activation of pyramidal cells or PV+ cells can trigger SWRs, as shown in vitro, and suggests that PV+ cell-mediated short-term synaptic depression influences the experimentally reported dynamics of SWR events. Furthermore, we predict that the silencing of anti-SWR interneurons can trigger SWRs. These results broaden our understanding of the microcircuits supporting the generation of memory-related network dynamics.SIGNIFICANCE STATEMENT The hippocampus is a part of the mammalian brain that is crucial for episodic memories. During periods of sleep and inactive waking, the extracellular activity of the hippocampus is dominated by sharp wave-ripple events (SWRs), which have been shown to be important for memory consolidation. The mechanisms regulating the emergence of these events are still unclear. We developed a computational model to study the emergence of SWRs and to explain the roles of different cell types in regulating them. The model accounts for several previously unexplained features of SWRs and thus advances the understanding of memory-related dynamics.

PlexinA4-Semaphorin3A Mediated Crosstalk between Main Cortical Interneuron Classes is Required for Superficial Interneurons Lamination

In the mammalian cerebral cortex, the developmental events governing the allocation of different classes of inhibitory interneurons (INs) into distinct cortical layers are poorly understood. Here we report that the guidance receptor PlexinA4 (PLXNA4) is upregulated in serotonin receptor 3a-expressing ( HTR3A+) cortical INs (hINs) as they invade the cortical plate and that it regulates their laminar allocation to superficial cortical layers. We find that the PLXNA4 ligand Semaphorin3A (SEMA3A) acts as a chemorepulsive factor on hINs migrating into the nascent cortex and demonstrate that SEMA3A specifically controls their laminar positioning through PLXNA4. We identify that deep layer INs constitute a major source of SEMA3A in the developing cortex and demonstrate that cell-type specific genetic deletion of SEMA3A in these INs specifically affects the laminar allocation of hINs. These data demonstrate that in the neocortex, deep layer INs control the laminar allocation of hINs into superficial layers.

Contrasting sex-dependent adaptations to synaptic physiology and membrane properties of prefrontal cortex interneuron subtypes in a mouse model of binge drinking

Alcohol use disorder (AUD) affects all sexes, however women who develop AUD may be particularly susceptible to cravings and other components of the disease. While many brain regions are involved in AUD etiology, proper prefrontal cortex (PFC) function is particularly important for top-down craving management and the moderation of drinking behaviors. Essential regulation of PFC output is provided by local inhibitory interneurons, yet how drinking affects interneuron physiology remains poorly understood, particularly in female individuals. To address this gap, we generated fluorescent reporter transgenic mice to label the two major classes of interneuron in deep layer prelimbic PFC, based on expression of parvalbumin (PV-IN) or somatostatin (SST-IN). We then interrogated PV-IN and SST-IN membrane and synaptic physiology in a rodent model of binge drinking. Beginning in late adolescence, mice received 3–4 weeks of intermittent access (IA) ethanol. We prepared acute brain slices one day after the last drinking session. PV-INs but not SST-INs from IA ethanol mice displayed increased excitability relative to controls, regardless of sex. On the contrary, synaptic adaptations to PV-INs differed based on sex. While drinking decreased excitatory synaptic strength onto PV-INs from female mice, PV-INs from IA ethanol male mice exhibited potentiated excitatory transmission relative to controls. In contrast, decreased synaptic strength onto SST-INs was observed following IA ethanol in all groups of mice. Together, these findings illustrate novel sex differences in drinking-related PFC pathophysiology. Discovering means to restore PV-IN and SST-IN dysfunction following extended drinking provides opportunities for developing new treatments for all AUD patients.

A Quantitative Comparison of Inhibitory Interneuron Size and Distribution between Mouse and Macaque V1, Using Calcium-Binding Proteins.

The mouse is a useful and popular model for studying of visual cortical function. To facilitate the translation of results from mice to primates, it is important to establish the extent of cortical organization equivalence between species and to identify possible differences. We focused on the different types of interneurons as defined by calcium-binding protein (CBP) expression in the layers of primary visual cortex (V1) in mouse and rhesus macaque. CBPs parvalbumin (PV), calbindin (CB), and calretinin (CR) provide a standard, largely nonoverlapping, labeling scheme in macaque, with preserved corresponding morphologies in mouse, despite a slightly higher overlap. Other protein markers, which are relevant in mouse, are not preserved in macaque. We fluorescently tagged CBPs in V1 of both species, using antibodies raised against preserved aminoacid sequences. Our data demonstrate important similarities between the expression patterns of interneuron classes in the different layers between rodents and primates. However, in macaque, expression of PV and CB is more abundant, CR expression is lower, and the laminar distribution of interneuron populations is more differentiated. Our results reveal an integrated view of interneuron types that provides a basis for translating results from rodents to primates, and suggest a reconciliation of previous results.

Transcriptional and imaging-genetic association of cortical interneurons, brain function, and schizophrenia risk

Inhibitory interneurons orchestrate information flow across the cortex and are implicated in psychiatric illness. Although interneuron classes have unique functional properties and spatial distributions, the influence of interneuron subtypes on brain function, cortical specialization, and illness risk remains elusive. Here, we demonstrate stereotyped negative correlation of somatostatin and parvalbumin transcripts within human and non-human primates. Cortical distributions of somatostatin and parvalbumin cell gene markers are strongly coupled to regional differences in functional MRI variability. In the general population (n = 9,713), parvalbumin-linked genes account for an enriched proportion of heritable variance in in-vivo functional MRI signal amplitude. Single-marker and polygenic cell deconvolution establish that this relationship is spatially dependent, following the topography of parvalbumin expression in post-mortem brain tissue. Finally, schizophrenia genetic risk is enriched among interneuron-linked genes and predicts cortical signal amplitude in parvalbumin-biased regions. These data indicate that the molecular-genetic basis of brain function is shaped by interneuron-related transcripts and may capture individual differences in schizophrenia risk.

Paradoxically Sparse Chemosensory Tuning in Broadly Integrating External Granule Cells in the Mouse Accessory Olfactory Bulb.

The accessory olfactory bulb (AOB), the first neural circuit in the mouse accessory olfactory system, is critical for interpreting social chemosignals. Despite its importance, AOB information processing is poorly understood compared with the main olfactory bulb (MOB). Here, we sought to fill gaps in the understanding of AOB interneuron function. We used 2-photon GCaMP6f Ca2+ imaging in an ex vivo preparation to study chemosensory tuning in AOB external granule cells (EGCs), interneurons hypothesized to broadly inhibit activity in excitatory mitral cells (MCs). In ex vivo preparations from mice of both sexes, we measured MC and EGC tuning to natural chemosignal blends and monomolecular ligands, finding that EGC tuning was sparser, not broader, than upstream MCs. Simultaneous electrophysiological recording and Ca2+ imaging showed no differences in GCaMP6f-to-spiking relationships in these cell types during simulated sensory stimulation, suggesting that measured EGC sparseness was not due to cell type-dependent variability in GCaMP6f performance. Ex vivo patch-clamp recordings revealed that EGC subthreshold responsivity was far broader than indicated by GCaMP6f Ca2+ imaging, and that monomolecular ligands rarely elicited EGC spiking. These results indicate that EGCs are selectively engaged by chemosensory blends, suggesting different roles for EGCs than analogous interneurons in the MOB.SIGNIFICANCE STATEMENT The mouse accessory olfactory system (AOS) interprets social chemosignals, but we poorly understand AOS information processing. Here, we investigate the functional properties of external granule cells (EGCs), a major class of interneurons in the accessory olfactory bulb (AOB). We hypothesized that EGCs, which are densely innervated by excitatory mitral cells (MCs), would show broad chemosensory tuning, suggesting a role in divisive normalization. Using ex vivo GCaMP6f imaging, we found that EGCs were instead more sparsely tuned than MCs. This was not due to weaker GCaMP6f signaling in EGCs than in MCs. Instead, we found that many MC-activating chemosignals caused only subthreshold EGC responses. This indicates a different role for AOB EGCs compared with analogous cells in the main olfactory bulb.

Afadin Signaling at the Spinal Neuroepithelium Regulates Central Canal Formation and Gait Selection

Afadin, a scaffold protein controlling the activity of the nectin family of cell adhesion molecules, regulates important morphogenetic processes during development. In the central nervous system, afadin has critical roles in neuronal migration, axonal elongation, and synapse formation. Here we examine the role of afadin in development of spinal motor circuits. Afadin elimination in motor neuron progenitors results in striking locomotor behavior: left-right limb alternation is substituted by synchronous activation, characteristic of bound gait. We find that afadin function at the neuroepithelium is required for structural organization of the spinal midline and central canal morphogenesis. Perturbation of afadin results in formation of two central canals, aberrant contralateral wiring of different classes of spinal premotor interneurons, and loss of left-right limb alternation, highlighting important developmental principles controlling the assembly of spinal motor circuits.

Mouse Retinal Cell Atlas: Molecular Identification of over Sixty Amacrine Cell Types.

Amacrine cells (ACs) are a diverse class of interneurons that modulate input from photoreceptors to retinal ganglion cells (RGCs), rendering each RGC type selectively sensitive to particular visual features, which are then relayed to the brain. While many AC types have been identified morphologically and physiologically, they have not been comprehensively classified or molecularly characterized. We used high-throughput single-cell RNA sequencing to profile >32,000 ACs from mice of both sexes and applied computational methods to identify 63 AC types. We identified molecular markers for each type and used them to characterize the morphology of multiple types. We show that they include nearly all previously known AC types as well as many that had not been described. Consistent with previous studies, most of the AC types expressed markers for the canonical inhibitory neurotransmitters GABA or glycine, but several expressed neither or both. In addition, many expressed one or more neuropeptides, and two expressed glutamatergic markers. We also explored transcriptomic relationships among AC types and identified transcription factors expressed by individual or multiple closely related types. Noteworthy among these were Meis2 and Tcf4, expressed by most GABAergic and most glycinergic types, respectively. Together, these results provide a foundation for developmental and functional studies of ACs, as well as means for genetically accessing them. Along with previous molecular, physiological, and morphologic analyses, they establish the existence of at least 130 neuronal types and nearly 140 cell types in the mouse retina.SIGNIFICANCE STATEMENT The mouse retina is a leading model for analyzing the development, structure, function, and pathology of neural circuits. A complete molecular atlas of retinal cell types provides an important foundation for these studies. We used high-throughput single-cell RNA sequencing to characterize the most heterogeneous class of retinal interneurons, amacrine cells, identifying 63 distinct types. The atlas includes types identified previously as well as many novel types. We provide evidence for the use of multiple neurotransmitters and neuropeptides, and identify transcription factors expressed by groups of closely related types. Combining these results with those obtained previously, we proposed that the mouse retina contains ∼130 neuronal types and is therefore comparable in complexity to other regions of the brain.

Compartmentalized Input–Output Organization of Lugaro Cells in the Cerebellar Cortex

Purkinje cells (PCs) are principal cerebellar neurons, and several classes of interneurons modulate their activity. Lugaro cells (LCs) are one such inhibitory interneuron with distinctive cytology and location, but still most enigmatic among cerebellar neurons. Here we serendipitously produced a novel transgenic mouse line, where a half of Yellow Cameleon (YC)(+) cells in the cerebellar cortex were judged to be LCs, and YC(+) LCs were estimated to constitute one-third of the total LC populations. Neurochemically, two-thirds of YC(+) LCs were dually GABAergic/glycinergic, with the rest being GABAergic. Beneath the PC layer, they extended a sheet of somatodendritic meshwork interconnected with neighboring LCs by adherens junctions, and received various inputs from climbing fibers, mossy fibers, granule cell axons, recurrent PC axons, Golgi cell axons, LC axons, and serotonergic fibers. Intriguingly, somatodendritic elements of individual LCs preferentially extended within a given cerebellar compartment defined by aldolase C expression. In turn, YC(+) LCs projected a dense lattice of ascending and transverse axons to the molecular layer, and innervated molecular layer interneurons (basket and stellate cells) and Golgi cells, but not PCs. Of note, ascending axons profusely innervated individual targets within a cerebellar compartment, while transverse axons ran across several compartments and innervated targets sparsely. This unique circuit configuration highlights that LCs integrate various excitatory, inhibitory, and modulatory inputs coming to the belonging cerebellar compartment and that, as an interneuron-selective interneuron, LCs can effectively disinhibit cerebellar cortical activities in a compartment-dependent manner through inhibition of inhibitory interneurons selectively targeting PCs and granule cells.

Axonal Tree Morphology and Signal Propagation Dynamics Improve Interneuron Classification.

Neurons are diverse and can be differentiated by their morphological, electrophysiological, and molecular properties. Current morphology-based classification approaches largely rely on the dendritic tree structure or on the overall axonal projection layout. Here, we use data from public databases of neuronal reconstructions and membrane properties to study the characteristics of the axonal and dendritic trees for interneuron classification. We show that combining signal propagation patterns observed by biophysical simulations of the activity along ramified axonal trees with morphological parameters of the axonal and dendritic trees, significantly improve classification results compared to previous approaches. The classification schemes introduced here can be utilized for robust neuronal classification. Our work paves the way for understanding and utilizing form-function principles in realistic neuronal reconstructions.

Postnatal maturation of spinal dynorphin circuits and their role in somatosensation.

Inhibitory interneurons in the adult spinal dorsal horn (DH) can be neurochemically classified into subpopulations that regulate distinct somatosensory modalities. Although inhibitory networks in the rodent DH undergo dramatic remodeling over the first weeks of life, little is known about the maturation of identified classes of GABAergic interneurons, or whether their role in somatosensation shifts during development. We investigated age-dependent changes in the connectivity and function of prodynorphin (DYN)-lineage neurons in the mouse DH that suppress mechanosensation and itch during adulthood. In vitro patch clamp recordings revealed a developmental increase in primary afferent drive to DYN interneurons and a transition from exclusive C-fiber monosynaptic input to mixed A-fiber and C-fiber innervation. Although most adult DYN interneurons exhibited tonic firing as expected from their inhibitory phenotype, neonatal and adolescent DYN cells were predominantly classified as phasic or single-spiking. Importantly, we also found that most of the inhibitory presynaptic terminals contacting lamina I spinoparabrachial projection neurons (PNs) originate from DYN neurons. Furthermore, inhibitory synaptic input from DYN interneurons onto PNs was weaker during the neonatal period, likely reflecting a lower number of GABAergic terminals and a reduced probability of GABA release compared to adults. Finally, spinal DYN interneurons attenuated mechanical sensitivity throughout development, but this population dampened acute nonhistaminergic itch only during adulthood. Collectively, these findings suggest that the spinal "gates" controlling sensory transmission to the brain may emerge in a modality-selective manner during early life due to the postnatal tuning of inhibitory synaptic circuits within the DH.

Histological characterization of interneurons in Alzheimer's disease reveals a loss of somatostatin interneurons in the temporal cortex.

Neuronal dysfunction and synaptic loss are major hallmarks of Alzheimer's disease (AD) which correlate with symptom severity. Impairment of the γ-aminobutyric acid (GABA)ergic inhibitory interneurons, which form around 20% of the total neuronal network, may be an early event contributing to neuronal circuit dysfunction in neurodegenerative diseases. This study examined the expression of two of the main classes of inhibitory interneurons, parvalbumin (PV) and somatostatin (SST) interneurons in the temporal cortex and hippocampus of AD and control cases, using immunohistochemistry. We report a significant regional variation in the number of PV and SST interneurons with a higher number identified per mm2 in the temporal cortex compared to the hippocampus. Fewer SST interneurons, but not PV interneurons, were identified per mm2 in the temporal cortex of AD cases compared to control subjects. Our results support regional neuroanatomical effects on selective interneuron classes in AD, and suggest that impairment of the interneuronal circuit may contribute to neuronal dysfunction and cognitive decline in AD.

Cholecystokinin-Expressing Interneurons of the Medial Prefrontal Cortex Mediate Working Memory Retrieval.

Distinct components of working memory are coordinated by different classes of inhibitory interneurons in the PFC, but the role of cholecystokinin (CCK)-positive interneurons remains enigmatic. In humans, this major population of interneurons shows histological abnormalities in schizophrenia, an illness in which deficient working memory is a core defining symptom and the best predictor of long-term functional outcome. Yet, CCK interneurons as a molecularly distinct class have proved intractable to examination by typical molecular methods due to widespread expression of CCK in the pyramidal neuron population. Using an intersectional approach in mice of both sexes, we have succeeded in labeling, interrogating, and manipulating CCK interneurons in the mPFC. Here, we describe the anatomical distribution, electrophysiological properties, and postsynaptic connectivity of CCK interneurons, and evaluate their role in cognition. We found that CCK interneurons comprise a larger proportion of the mPFC interneurons compared with parvalbumin interneurons, targeting a wide range of neuronal subtypes with a distinct connectivity pattern. Phase-specific optogenetic inhibition revealed that CCK, but not parvalbumin, interneurons play a critical role in the retrieval of working memory. These findings shine new light on the relationship between cortical CCK interneurons and cognition and offer a new set of tools to investigate interneuron dysfunction and cognitive impairments associated with schizophrenia.SIGNIFICANCE STATEMENT Cholecystokinin-expressing interneurons outnumber other interneuron populations in key brain areas involved in cognition and memory, including the mPFC. However, they have proved intractable to examination as experimental techniques have lacked the necessary selectivity. To the best of our knowledge, the present study is the first to report detailed properties of cortical cholecystokinin interneurons, revealing their anatomical organization, electrophysiological properties, postsynaptic connectivity, and behavioral function in working memory.

TrkB Signaling Influences Gene Expression in Cortistatin-Expressing Interneurons.

Brain-derived neurotrophic factor (BDNF) signals through its cognate receptor tropomyosin receptor kinase B (TrkB) to promote the function of several classes of inhibitory interneurons. We previously reported that loss of BDNF–TrkB signaling in cortistatin (Cort)-expressing interneurons leads to behavioral hyperactivity and spontaneous seizures in mice. We performed bulk RNA sequencing (RNA-seq) from the cortex of mice with disruption of BDNF–TrkB signaling in cortistatin interneurons, and identified differential expression of genes important for excitatory neuron function. Using translating ribosome affinity purification and RNA-seq, we define a molecular profile for Cort-expressing inhibitory neurons and subsequently compare the translatome of normal and TrkB-depleted Cort neurons, revealing alterations in calcium signaling and axon development. Several of the genes enriched in Cort neurons and differentially expressed in TrkB-depleted neurons are also implicated in autism and epilepsy. Our findings highlight TrkB-dependent molecular pathways as critical for the maturation of inhibitory interneurons and support the hypothesis that loss of BDNF signaling in Cort interneurons leads to altered excitatory/inhibitory balance.

SLC17A6/7/8 Vesicular Glutamate Transporter Homologs in Nematodes.

Members of the superfamily of solute carrier (SLC) transmembrane proteins transport diverse substrates across distinct cellular membranes. Three SLC protein families transport distinct neurotransmitters into synaptic vesicles to enable synaptic transmission in the nervous system. Among them is the SLC17A6/7/8 family of vesicular glutamate transporters, which endows specific neuronal cell types with the ability to use glutamate as a neurotransmitter. The genome of the nematode Caenorhabditis elegans encodes three SLC17A6/7/8 family members, one of which, eat-4/VGLUT, has been shown to be involved in glutamatergic neurotransmission. Here, we describe our analysis of the two remaining, previously uncharacterized SLC17A6/7/8 family members, vglu-2 and vglu-3 These two genes directly neighbor one another and are the result of a recent gene duplication event in C. elegans, but not in other Caenorhabditis species. Compared to EAT-4, the VGLU-2 and VGLU-3 protein sequences display a more distant similarity to canonical, vertebrate VGLUT proteins. We tagged both genomic loci with gfp and detected no expression of vglu-3 at any stage of development in any cell type of both C. elegans sexes. In contrast, vglu-2::gfp is dynamically expressed in a restricted set of distinct cell types. Within the nervous system, vglu-2::gfp is exclusively expressed in a single interneuron class, AIA, where it localizes to vesicular structures in the soma, but not along the axon, suggesting that VGLU-2 may not be involved in synaptic transport of glutamate. Nevertheless, vglu-2 mutants are partly defective in the function of the AIA neuron in olfactory behavior. Outside the nervous system, VGLU-2 is expressed in collagen secreting skin cells where VGLU-2 most prominently localizes to early endosomes, and to a lesser degree to apical clathrin-coated pits, the trans-Golgi network, and late endosomes. On early endosomes, VGLU-2 colocalizes most strongly with the recycling promoting factor SNX-1, a retromer component. Loss of vglu-2 affects the permeability of the collagen-containing cuticle of the worm, and based on the function of a vertebrate VGLUT1 protein in osteoclasts, we speculate that vglu-2 may have a role in collagen trafficking in the skin. We conclude that C. elegans SLC17A6/7/8 family members have diverse functions within and outside the nervous system.

Disrupted GABAergic facilitation of working memory performance in people with schizophrenia.

Gamma-Amiobutyric acid (GABA) is a primary inhibitory neurotransmitter that facilitates neural oscillations that coordinate neural activity between brain networks to facilitate cognition. The present magnetic resonance spectroscopy (MRS) study tests the hypothesis that GABAergic facilitation of working memory is disrupted in people with schizophrenia (PSZ). 51 healthy participants and 40 PSZ from the UC Davis Early Psychosis Program performed an item and temporal order working memory (WM) task and underwent resting MRS to measure GABA and glutamate concentrations in dorsolateral prefrontal (DLPFC) and anterior cingulate (ACC) regions of interest. MRS was acquired on a 3 Tesla Siemens scanner and GABA and glutamate concentrations were referenced to creatine. Percent correct on the WM task indexed performance and correlation coefficients examined GABAergic or Glutamatergic facilitation of WM, with Fisher's Z transformation testing for group differences. There were no group differences in GABA or glutamate concentrations, but WM correlations were reversed between groups. In patients, higher DLPFC GABA was associated with worse rather than better WM performance. This pattern was not observed for glutamate or in the ACC. Although under-powered, there was no indication of medication effects. Results cannot be explained by group differences in DLPFC GABA or glutamate concentrations but, instead, indicate that schizophrenia disrupts the GABAergic facilitation of WM seen in healthy individuals. Results appear to parallel post mortem findings in suggesting that schizophrenia alters the distribution of different classes of GABAergic interneurons rather than producing a general deficit across the total population of neurons.