AAV11 permits efficient retrograde targeting of projection neurons

Viral tracers that permit efficient retrograde targeting of projection neurons are powerful vehicles for structural and functional dissections of the neural circuit and for the treatment of brain diseases. Recombinant adeno-associated viruses (rAAVs) are the most potential candidates because they are low-toxic with high-level transgene expression and minimal host immune responses. Currently, some rAAVs based on capsid engineering for retrograde tracing have been widely used in the analysis and manipulation of neural circuits, but suffer from brain area selectivity and inefficient retrograde transduction in certain neural connections. Here, we discovered that the recombinant adeno-associated virus 11 (rAAV11) exhibits potent retrograde labeling of projection neurons with enhanced efficiency to rAAV2-retro in some neural connections. Combined with calcium recording technology, rAAV11 can be used to monitor neuronal activities by expressing Cre recombinase or calcium-sensitive functional probe. In addition, we further showed the suitability of rAAV11 for astrocyte targeting. These properties make rAAV11 a promising tool for the mapping and manipulation of neural circuits and gene therapy of some neurological and neurodegenerative disorders.

Neural Circuits and Some New Factors Involved in Hippocampal Memory

Humans and other primates have memory, and the hippocampus plays a critical role in this process. The neural circuitry is one of the structural foundations for the hippocampus in exerting memory function. To understand the relationship between the hippocampus and memory, we need to understand neural circuits. Past research has identified several classical neural circuits involved in memory. Although there are challenges with the study of hippocampal neural circuits, research on this topic has continued, and some progress has been made. Here, we discuss recent advances in our understanding of hippocampal neural circuit mechanisms and some of the newly discovered factors that affect memory. Substantial progress has been made regarding hippocampal memory circuits and Alzheimer’s disease. However, it is unclear whether these novel findings regarding hippocampal memory circuits hold promise for human memory studies. Additional research on this topic is needed.

Developmental iron deficiency dysregulates TET activity and DNA hydroxymethylation in the rat hippocampus and cerebellum

Iron deficiency (ID) during neurodevelopment is associated with lasting cognitive and socioemotional deficits, and increased risk for neuropsychiatric disease throughout the lifespan. These neurophenotypical changes are underlain by gene dysregulation in the brain that outlasts the period of ID; however, the mechanisms by which ID establishes and maintains gene expression changes are incompletely understood. The epigenetic modification 5-hydroxymethylcytosine (5hmC), or DNA hydroxymethylation, is one candidate mechanism because of its dependence on iron-containing TET enzymes. The aim of the present study was to determine the effect of fetal-neonatal ID on regional brain TET activity, Tet expression, and 5hmC in the developing rat hippocampus and cerebellum, and to determine whether changes are reversible with dietary iron treatment. Timed pregnant Sprague-Dawley rats were fed iron deficient diet (ID; 4 mg/kg Fe) from gestational day (G)2 to generate iron deficient anemic (IDA) offspring. Control dams were fed iron sufficient diet (IS; 200 mg/kg Fe). At postnatal day (P)7, a subset of ID-fed litters was randomized to IS diet, generating treated IDA (TIDA) offspring. At P15, hippocampus and cerebellum were isolated for subsequent analysis. TET activity was quantified by ELISA from nuclear proteins. Expression of Tet1, Tet2, and Tet3 was quantified by qPCR from total RNA. Global %5hmC was quantified by ELISA from genomic DNA. ID increased DNA hydroxymethylation (p=0.0105), with a corresponding increase in TET activity (p<0.0001) and Tet3 expression (p<0.0001) in the P15 hippocampus. In contrast, ID reduced TET activity (p=0.0016) in the P15 cerebellum, with minimal effect on DNA hydroxymethylation. Neonatal dietary iron treatment resulted in partial normalization of these changes in both brain regions. These results demonstrate that the TET/DNA hydroxymethylation system is disrupted by developmental ID in a brain region-specific manner. Differential regional disruption of this epigenetic system may contribute to the lasting neural circuit dysfunction and neurobehavioral dysfunction associated with developmental ID.

Determinants of synapse diversity revealed by super-resolution quantal transmission and active zone imaging

Neural circuit function depends on the pattern of synaptic connections between neurons and the strength of those connections. Synaptic strength is determined by both postsynaptic sensitivity to neurotransmitter and the presynaptic probability of action potential evoked transmitter release (Pr). Whereas morphology and neurotransmitter receptor number indicate postsynaptic sensitivity, presynaptic indicators and the mechanism that sets Pr remain to be defined. To address this, we developed QuaSOR, a super-resolution method for determining Pr from quantal synaptic transmission imaging at hundreds of glutamatergic synapses at a time. We mapped the Pr onto super-resolution 3D molecular reconstructions of the presynaptic active zones (AZs) of the same synapses at the Drosophila larval neuromuscular junction (NMJ). We find that Pr varies greatly between synapses made by a single axon, quantify the contribution of key AZ proteins to Pr diversity and find that one of these, Complexin, suppresses spontaneous and evoked transmission differentially, thereby generating a spatial and quantitative mismatch between release modes. Transmission is thus regulated by the balance and nanoscale distribution of release-enhancing and suppressing presynaptic proteins to generate high signal-to-noise evoked transmission.

Low-cost and easy-fabrication lightweight drivable electrode array for multiple-regions electrophysiological recording in free-moving mice

Objective. Extracellular electrophysiology has been widely applied to neural circuit dissections. However, long-term multiregional recording in free-moving mice remains a challenge. Low-cost and easy-fabrication of elaborate drivable electrodes is required for their prevalence. Approach. A three-layer nested construct (OD ~1.80 mm, length ~10 mm, <0.1g) was recruited as a drivable component, which consisted of an ethylene-vinyl acetate copolymer (EVA) heat-shrinkable tube, non-closed loop ceramic bushing, and stainless ferrule with a bulge twining silver wire. The supporting and working components were equipped with drivable components to be assembled into a drivable microwire electrode array with a nested structure (drivable MEANS). Two drivable microwire electrode arrays were independently implanted for chronic recording in different brain areas at respective angles. An optic fiber was easily loaded into the drivable MEANS to achieve optogenetic modulation and electrophysiological recording simultaneously. Main results. The drivable MEANS had lightweight (~ 0.37 g), small (~ 15 mm ×15 mm × 4 mm), and low cost (≤ $64.62). Two drivable MEANS were simultaneously implanted in mice, and high-quality electrophysiological recordings could be applied ≥ 5 months after implantation in freely behaving animals. Electrophysiological recordings and analysis of the lateral septum (LS) and lateral hypothalamus (LH) in food-seeking behavior demonstrated that our drivable MEANS can be used to dissect the function of neural circuits. An optical fiber-integrated drivable MEANS (~ 0.47 g) was used to stimulate and record LS neurons, which suggested that changes in working components can achieve more functions than electrophysiological recordings, such as optical stimulation, drug release, and calcium imaging. Significance. Drivable MEANS is an easily fabricated, lightweight drivable microwire electrode array for multiple-region electrophysiological recording in free-moving mice. Our design is likely to be a valuable platform for both current and prospective users, as well as for developers of multifunctional electrodes for free-moving mice.

Neural Circuits Underlying Behavioral Flexibility: Insights From Drosophila

Behavioral flexibility is critical to survival. Animals must adapt their behavioral responses based on changes in the environmental context, internal state, or experience. Studies in Drosophila melanogaster have provided insight into the neural circuit mechanisms underlying behavioral flexibility. Here we discuss how Drosophila behavior is modulated by internal and behavioral state, environmental context, and learning. We describe general principles of neural circuit organization and modulation that underlie behavioral flexibility, principles that are likely to extend to other species.

A Conditional Glutamatergic Synaptic Vesicle Marker for Drosophila

Glutamate is a principal neurotransmitter used extensively by the nervous systems of all vertebrate and invertebrate animals. It is primarily an excitatory neurotransmitter that has been implicated in nervous system development as well as a myriad of brain functions from the simple transmission of information between neurons to more complex aspects of nervous system function including synaptic plasticity, learning, and memory. Identification of glutamatergic neurons and their sites of glutamate release are thus essential for understanding the mechanisms of neural circuit function and how information is processed to generate behavior. Here we describe and characterize smFLAG-vGlut, a conditional marker of glutamatergic synaptic vesicles for the Drosophila model system. smFLAG-vGlut is validated for functionality, conditional expression, and specificity for glutamatergic neurons and synaptic vesicles. The utility of smFLAG-vGlut is demonstrated by glutamatergic neurotransmitter phenotyping of 26 different central complex neuron types of which nine were established to be glutamatergic. This illumination of glutamate neurotransmitter usage will enhance the modeling of central complex neural circuitry and thereby our understanding of information processing by this region of the fly brain. The use of smFLAG for glutamatergic neurotransmitter phenotyping and identification of glutamate release sites can be extended to any Drosophila neuron(s) represented by a binary transcription system driver.

Endocannabinoid-Mediated Control of Neural Circuit Excitability and Epileptic Seizures

Research on endocannabinoid signaling has greatly advanced our understanding of how the excitability of neural circuits is controlled in health and disease. In general, endocannabinoid signaling at excitatory synapses suppresses excitability by inhibiting glutamate release, while that at inhibitory synapses promotes excitability by inhibiting GABA release, although there are some exceptions in genetically epileptic animal models. In the epileptic brain, the physiological distributions of endocannabinoid signaling molecules are disrupted during epileptogenesis, contributing to the occurrence of spontaneous seizures. However, it is still unknown how endocannabinoid signaling changes during seizures and how the redistribution of endocannabinoid signaling molecules proceeds during epileptogenesis. Recent development of cannabinoid sensors has enabled us to investigate endocannabinoid signaling in much greater spatial and temporal details than before. Application of cannabinoid sensors to epilepsy research has elucidated activity-dependent changes in endocannabinoid signaling during seizures. Furthermore, recent endocannabinoid research has paved the way for the clinical use of cannabidiol for the treatment of refractory epilepsy, such as Dravet syndrome, Lennox-Gastaut syndrome and tuberous sclerosis complex. Cannabidiol significantly reduces seizures and is considered to have comparable tolerability to conventional antiepileptic drugs. In this article, we introduce recent advances in research on the roles of endocannabinoid signaling in epileptic seizures and discuss future directions.

Neuronal cell-type and circuit basis for visual predictive processing

Bayesian Brain theory suggests brain utilises predictive processing framework to interpret the noisy world. Predictive processing is essential to perception, action, cognition and psychiatric disease, but underlying neural circuit mechanisms remain undelineated. Here we show the neuronal cell-type and circuit basis for visual predictive processing in awake, head-fixed mice during self-initiated running. Preceding running, vasoactive intestinal peptide (VIP)-expressing inhibitory interneurons (INs) in primary visual cortex (V1) are robustly activated in absence of structured visual stimuli. This pre-running activation is mediated via distal top-down projections from frontal, parietal and retrosplenial areas known for motion planning, but not local excitatory inputs associated with the bottom-up pathway. Somatostatin (SST) INs show pre-running suppression and post-running activation, indicating a VIP-SST motif. Differential VIP-SST peri-running dynamics anisotropically suppress neighbouring pyramidal (Pyr) neurons, preadapting Pyr neurons to the incoming running. Our data delineate key neuron types and circuit elements of predictive processing brain employs in action and perception.