Functionally distinct dopamine domains in the hippocampus.

Numerous studies have identified dopamine signaling in the hippocampus as necessary for certain types of learning and memory. Since dopamine in the striatum is strongly tied to rewards, dopamine in the hippocampus is thought to reinforce reward learning. Despite the critical influence of dopamine on hippocampal function, little is known about dopamine release in the hippocampus or the specific ways dopamine can influence hippocampal function. Based on the functional complexity of hippocampal circuitry, we hypothesized the existence of multiple dopamine signaling domains. Using optical dopamine sensors, two-photon imaging, and head-fixed behaviors, we identified two functionally and spatially distinct dopamine domains in the hippocampus. The "superficial" domain (cell somata and apical dendrites) showed reward-related dopamine transients early in Pavlovian conditioning but were replaced by "deep" domain transients (basal dendritic layer) with experience. These two domains also play distinct roles in a hippocampal-dependent, goal-directed virtual reality task where mice use exploratory licks to discover the location of a hidden reward zone. Here, positive dopamine ramps appeared in the superficial domain as mice approached the reward zone, similar to those seen in the striatum. At the same time, the deep domain showed strong reward-related transients. These results reveal small-scale, anatomically segregated, dopamine domains in the hippocampus. Furthermore dopamine domain activity had temporal-specificity for different phases of behavior. Finally, the subcellular scale of dopamine domains suggests specialized postsynaptic pathways for processing and integrating functionally distinct dopaminergic influences.

Tmem178 Negatively Regulates IL-1ß Production Through Inhibition of the NLRP3 Inflammasome.

OBJECTIVE: Inflammasomes modulate the release of bioactive interleukin (IL)-1ß. Excessive IL-1ß levels are detected in patients with systemic juvenile idiopathic arthritis (sJIA) and cytokine storm syndrome (CSS) with mutated and unmutated inflammasome components, raising questions on the mechanisms of IL-1ß regulation in these disorders. METHODS: To investigate how the NLRP3 inflammasome is modulated in sJIA, we focused on Transmembrane protein 178 (Tmem178), a negative regulator of calcium levels in macrophages, and measured IL-1ß and caspase-1 activation in wild-type (WT) and Tmem178-/- macrophages after calcium chelators, silencing of Stim1, a component of store-operated calcium entry (SOCE), or by expressing a Tmem178 mutant lacking the Stromal Interaction Molecule 1 (Stim1) binding site. Mitochondrial function in both genotypes was assessed by measuring oxidative respiration, mitochondrial reactive oxygen species (mtROS), and mitochondrial damage. CSS development was analyzed in Perforin-/- /Tmem178-/- mice infected with lymphocytic choriomeningitis virus (LCMV) in which inflammasome or IL-1ß signaling was pharmacologically inhibited. Human TMEM178 and IL1B transcripts were analyzed in data sets of whole blood and peripheral blood monocytes from healthy controls and patients with active sJIA. RESULTS: TMEM178 levels are reduced in whole blood and monocytes from patients with sJIA while IL1B levels are increased. Accordingly, Tmem178-/- macrophages produce elevated IL-1ß compared with WT cells. The elevated intracellular calcium levels after SOCE activation in Tmem178-/- macrophages induce mitochondrial damage, release mtROS, and ultimately promote NLRP3 inflammasome activation. In vivo, inhibition of inflammasome or IL-1ß neutralization prolongs Tmem178-/- mouse survival in LCMV-induced CSS. CONCLUSION: Down-regulation of TMEM178 levels may represent a marker of disease activity and help identify patients who could benefit from inflammasome targeting.

Long non-coding RNA SNHG8 drives stress granule formation in tauopathies.

Tauopathies are a heterogenous group of neurodegenerative disorders characterized by tau aggregation in the brain. In a subset of tauopathies, rare mutations in the MAPT gene, which encodes the tau protein, are sufficient to cause disease; however, the events downstream of MAPT mutations are poorly understood. Here, we investigate the role of long non-coding RNAs (lncRNAs), transcripts >200 nucleotides with low/no coding potential that regulate transcription and translation, and their role in tauopathy. Using stem cell derived neurons from patients carrying a MAPT p.P301L, IVS10 + 16, or p.R406W mutation and CRISPR-corrected isogenic controls, we identified transcriptomic changes that occur as a function of the MAPT mutant allele. We identified 15 lncRNAs that were commonly differentially expressed across the three MAPT mutations. The commonly differentially expressed lncRNAs interact with RNA-binding proteins that regulate stress granule formation. Among these lncRNAs, SNHG8 was significantly reduced in a mouse model of tauopathy and in FTLD-tau, progressive supranuclear palsy, and Alzheimer's disease brains. We show that SNHG8 interacts with tau and stress granule-associated RNA-binding protein TIA1. Overexpression of mutant tau in vitro is sufficient to reduce SNHG8 expression and induce stress granule formation. Rescuing SNHG8 expression leads to reduced stress granule formation and reduced TIA1 levels in immortalized cells and in MAPT mutant neurons, suggesting that dysregulation of this non-coding RNA is a causal factor driving stress granule formation via TIA1 in tauopathies.

Long non-coding RNA SNHG8 drives stress granule formation in tauopathies.

Tauopathies are a heterogenous group of neurodegenerative disorders characterized by tau aggregation in the brain. In a subset of tauopathies, rare mutations in the MAPT gene, which encodes the tau protein, are sufficient to cause disease; however, the events downstream of MAPT mutations are poorly understood. Here, we investigate the role of long non-coding RNAs (lncRNAs), transcripts >200 nucleotides with low/no coding potential that regulate transcription and translation, and their role in tauopathy. Using stem cell derived neurons from patients carrying a MAPT p.P301L, IVS10+16, or p.R406W mutation, and CRISPR-corrected isogenic controls, we identified transcriptomic changes that occur as a function of the MAPT mutant allele. We identified 15 lncRNAs that were commonly differentially expressed across the three MAPT mutations. The commonly differentially expressed lncRNAs interact with RNA-binding proteins that regulate stress granule formation. Among these lncRNAs, SNHG8 was significantly reduced in a mouse model of tauopathy and in FTLD-tau, progressive supranuclear palsy, and Alzheimer’s disease brains. We show that SNHG8 interacts with tau and stress granule-associated RNA-binding protein TIA1. Overexpression of mutant tau in vitro is sufficient to reduce SNHG8 expression and induce stress granule formation. Rescuing SNHG8 expression leads to reduced stress granule formation and reduced TIA1 levels, suggesting that dysregulation of this non-coding RNA is a causal factor driving stress granule formation via TIA1 in tauopathies.

Tmem178 negatively regulates IL-1ß production through inhibition of the NLRP3 inflammasome.

Objective: Inflammasomes modulate the release of bioactive IL-1ß. Excessive IL-1ß levels are detected in patients with systemic juvenile idiopathic arthritis (sJIA) and cytokine storm syndrome (CSS) with mutated and unmutated inflammasome components, raising questions on the mechanisms of IL-1ß regulation in these disorders. Methods: To investigate how the NLRP3 inflammasome is modulated in sJIA, we focused on Tmem178, a negative regulator of calcium levels in macrophages, and measured IL-1ß and caspase-1 activation in wild-type (WT) and Tmem178 -/- macrophages following calcium chelators, silencing of Stim1, a component of store-operated calcium entry (SOCE), or by expressing a Tmem178 mutant lacking Stim1 binding site. Mitochondrial function in both genotypes was assessed by measuring oxidative respiration, mitochondrial reactive oxygen species (mtROS), and mitochondrial damage. CSS development was analyzed in Perforin -/- /Tmem178 -/- mice infected with LCMV in which inflammasome or IL-1 signaling was pharmacologically inhibited. Human TMEM178 and IL-1B transcripts were analyzed in a dataset of peripheral blood monocytes from healthy controls and active sJIA patients. Results: TMEM178 levels are reduced in monocytes from sJIA patients while IL-1B show increased levels. Accordingly, Tmem178 -/- macrophages produce elevated IL-1ß compared to WT cells. The elevated intracellular calcium levels following SOCE activation in Tmem178 -/- macrophages induce mitochondrial damage, release mtROS, and ultimately, promote NLRP3 inflammasome activation. In vivo , inhibition of inflammasome or IL-1 neutralization prolongs Tmem178 -/- mouse survival to LCMV-induced CSS. Conclusion: Downregulation of Tmem178 levels may represent a new biomarker to identify sJIA/CSS patients that could benefit from receiving drugs targeting inflammasome signaling.

Separate Functional Subnetworks of Excitatory Neurons Show Preference to Periodic and Random Sound Structures.

Auditory cortex (ACX) neurons are sensitive to spectro-temporal sound patterns and violations in patterns induced by rare stimuli embedded within streams of sounds. We investigate the auditory cortical representation of repeated presentations of sequences of sounds with standard stimuli (common) with an embedded deviant (rare) stimulus in two conditions, Periodic (Fixed deviant position) or Random (Random deviant position). We used extracellular single-unit and two-photon Ca2+ imaging recordings in layer 2/3 neurons of the mouse (Mus musculus) ACX of either sex. Population single-unit average responses increased over repetitions in the Random condition and were suppressed or did not change in the Periodic condition, showing general irregularity preference. A subset of neurons showed the opposite behavior, indicating regularity preference. Furthermore, pairwise noise correlations were higher in the Random condition than in the Periodic condition, suggesting a role of recurrent connections in the observed differential adaptation. Functional two-photon Ca2+ imaging showed that excitatory (EX), and inhibitory (IN) neurons [parvalbumin-positive (PV) and somatostatin-positive (SOM)] also had different categories of long-term adaptation as observed with single-units. However, examination of functional connectivity between pairs of neurons of different categories showed that EX-PV connected pairs behaved opposite to the EX-EX and EX-SOM pairs, with more connections outside category in Random condition than Periodic condition. Finally, considering Regularity, Irregularity, and no preference of connected pairs of neurons showed that EX-EX and EX-SOM pairs were in largely separate functional subnetworks with different preferences, not EX-PV pairs. Thus, separate subnetworks underlie coding of periodic and random sound sequences.SIGNIFICANCE STATEMENT Studying how the auditory cortex (ACX) neurons respond to streams of sound sequences help us understand the importance of changes in dynamic acoustic noisy scenes around us. Humans and animals are sensitive to regularity and its violations in sound sequences. Psychophysical tasks in humans show that the auditory brain differentially responds to Periodic and Random structures, independent of the listener's attentional states. Here, we show that mouse ACX L2/3 neurons detect changes and respond differently to patterns over long-time scales. The differential functional connectivity profile obtained in response to two different sound contexts suggests the vital role of recurrent connections in the auditory cortical network. Furthermore, the excitatory-inhibitory neuronal interactions can contribute to detecting the changing sound patterns.

Earliest Experience of a Relatively Rare Sound But Not a Frequent Sound Causes Long-Term Changes in the Adult Auditory Cortex.

Sensory experience during a critical period alters sensory cortical responses and organization. We find that the earliest sound-driven activity in the mouse auditory cortex (ACX) starts before ear-canal opening (ECO). The effects of auditory experience before ECO on ACX development are unknown. We find that in mouse ACX subplate neurons (SPNs), crucial in thalamocortical maturation, respond to sounds before ECO showing oddball selectivity. Before ECO, SPNs are more selective to oddball sounds in auditory streams than thalamo-recipient layer 4 (L4) neurons and not after ECO. We hypothesize that SPN's oddball selectivity can direct the development of L4 responses before ECO. Exposing mice, of either sex, before ECO to a rarely occurring tone in a stream of another tone occurring frequently leads to strengthening the adult cortical representation of the rare tone, but not that of the frequent tone. Results of control exposure experiments at multiple developmental windows that also use only a single tone corroborate the observations. We further explain the strengthening of deviant inputs before ECO and not after ECO using a binary network model mimicking the hierarchical structure of subplate and L4 neurons and response properties derived from our data, with synapses following Hebbian spike time-dependent plasticity learning rule. Information-theoretic analysis with sparse coding assumptions also predicts the observations. Thus, relatively salient low probability sounds in the earliest auditory environment cause long-term changes in the ACX.SIGNIFICANCE STATEMENT Early auditory experience can change the organization and responses of the auditory cortex in adulthood. However, little is known about how auditory experience at prenatal ages changes neural circuits and response properties. In mice at equivalent early developmental stages, we find that auditory experience of a particular kind, with a less frequently occurring sound in a stream of another sound, alters adult cortical responsiveness, specifically of the less frequent sound. However, at the previously known critical period of development, the opposite is observed, where the more frequent sound's representation is strengthened in the adult compared with the less frequent sound. We thus show that a specific type of auditory environment can influence adult auditory processing at the earliest ages.

One- and Two-Photon Uncaging: Carbazole Fused o-Hydroxycinnamate Platform for Dual Release of Alcohols (Same or Different) with Real-Time Monitoring.

A one- and two-photon activated photoremovable protecting group (PRPG) was designed based on a carbazole fused o-hydroxycinnamate platform for the dual (same or different) release of alcohols. The mechanism for the dual release proceeds through a stepwise pathway and also monitors the first and second photorelease in real time by an increase in fluorescence intensity and color change, respectively. Further, its application in staining live neurons and ex vivo imaging with two-photon excitation is shown.