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Neurons activated by learning have been ascribed the unique potential to encode memory, but the functional contribution of discrete cell types remains poorly understood. In particular, it is unclear whether learning engages specific GABAergic interneurons and, if so, whether they differ functionally from interneurons recruited by other experiences. Here, we show that fear conditioning activates a heterogeneous neuronal population in the medial prefrontal cortex (mPFC) that is largely comprised of somatostatin-expressing interneurons (SST-INs). Using intersectional genetic approaches, we demonstrate that fear-learning-activated SST-INs exhibit distinct circuit properties and are selectively reactivated to mediate cue-evoked memory expression. In contrast, an orthogonal population of SST-INs activated by morphine experience exerts opposing control over fear and supports reward-like motivational effects. These results outline an important role for discrete subsets of GABAergic cells in emotional learning and point to an unappreciated capacity for functional specialization among SST-INs.

The paradigm of fear conditioning is largely responsible for our current understanding of how memories are encoded at the cellular level. Its most fundamental underlying mechanism is considered to be plasticity of synaptic connections between excitatory projection neurons (PNs). However, recent studies suggest that while PNs execute critical memory functions, their activity at key stages of learning and recall is extensively orchestrated by a diverse array of GABAergic interneurons (INs). Here we review the contributions of genetically-defined INs to processing of threat-related stimuli in fear conditioning, with a particular focus on how synaptic interactions within interconnected networks of INs modulates PN activity through both inhibition and disinhibition. Furthermore, we discuss accumulating evidence that GABAergic microcircuits are an important locus for synaptic plasticity during fear learning and therefore a viable substrate for long-term memory. These findings suggest that further investigation of INs could unlock unique conceptual insights into the organization and function of fear memory networks.

A novel clinical assay for the detection and quantitation of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was adapted from an in-house, research-based enzyme-linked immunosorbent assay (ELISA). Development and validation were performed under regulatory guidelines, and the test obtained emergency use authorization (EUA) from the New York State Department of Health (NYSDOH) and the Food and Drug Administration (FDA). The Mount Sinai coronavirus disease 2019 (COVID-19) antibody assay is an orthogonal, quantitative direct ELISA test which detects antibodies reactive to the receptor binding domain (RBD) and the spike protein of the novel SARS-CoV-2. The assay is performed on 96-well plates coated with either SARS-CoV-2 recombinant RBD or spike proteins. The test is divided into two stages, a qualitative screening assay against RBD and a quantitative assay against the full-length spike protein. The test uses pooled high titer serum as a reference standard. Negative pre-COVID-19 and positive post-COVID-19, PCR-confirmed specimens were incorporated in each ELISA test run, and the assays were performed independently at two different locations. The Mount Sinai COVID-19 serology performed with high sensitivity and specificity, 92.5% (95% CI: 0.785–0.980) and 100% (CI: 0.939–1.000) respectively. Between-run precision was assessed with a single run repeated over 22 days; and within-run precision was assessed with 10 replicates per day over 22 days. Both were within reported acceptance criteria (CV ≤ 20%). This population-based study reveals the applicability and reliability of this novel orthogonal COVID-19 serology test for the detection and quantitation of antibodies against SARS-CoV-2, allowing a broad set of clinical applications, including the broad evaluation of SARS-CoV-2 seroprevalence and antibody profiling in different population subsets., Coronavirus disease 2019; Severe acute respiratory syndrome coronavirus-2 (SARSCoV-2); Antibody tests; Diagnosis of COVID-19; ELISA; IgG antibody assay.

A brain network comprising the medial prefrontal cortex (mPFC) and amygdala plays important roles in developmentally regulated cognitive and emotional processes. However, very little is known about the maturation of mPFC-amygdala circuitry. We conducted anatomical tracing of mPFC projections and optogenetic interrogation of their synaptic connections with neurons in the basolateral amygdala (BLA) at neonatal to adult developmental stages in mice. Results indicate that mPFC-BLA projections exhibit delayed emergence relative to other mPFC pathways and establish synaptic transmission with BLA excitatory and inhibitory neurons in late infancy, events that coincide with a massive increase in overall synaptic drive. During subsequent adolescence, mPFC-BLA circuits are further modified by excitatory synaptic strengthening as well as a transient surge in feedforward inhibition. The latter was correlated with increased spontaneous inhibitory currents in excitatory neurons, suggesting that mPFC-BLA circuit maturation culminates in a period of exuberant GABAergic transmission. These findings establish a time course for the onset and refinement of mPFC-BLA transmission and point to potential sensitive periods in the development of this critical network.SIGNIFICANCE STATEMENTHuman mPFC-amygdala functional connectivity is developmentally regulated and figures prominently in numerous psychiatric disorders with a high incidence of adolescent onset. However, it remains unclear when synaptic connections between these structures emerge or how their properties change with age. Our work establishes developmental windows and cellular substrates for synapse maturation in this pathway involving both excitatory and inhibitory circuits. The engagement of these substrates by early life experience may support the ontogeny of fundamental behaviors but could also lead to inappropriate circuit refinement and psychopathology in adverse situations.

Memory is a dynamic process that is continuously regulated by both synaptic and intrinsic neural mechanisms. While numerous studies have shown that synaptic plasticity is important in various types and phases of learning and memory, neuronal intrinsic excitability has received relatively less attention, especially regarding the dynamic nature of memory. In this review, we present evidence demonstrating the importance of intrinsic excitability in memory allocation, consolidation, and updating. We also consider the intricate interaction between intrinsic excitability and synaptic plasticity in shaping memory, supporting both memory stability and flexibility.

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate depends on properties intrinsic to each channel protein as well as the mechanism by which it is generated and controlled and represents an important area of current research. Information about the operational dynamics of ion channel proteins can be obtained by observing long stretches of current produced by a single molecule. Described here is a protocol for obtaining one-channel cell-attached patch-clamp current recordings for a ligand gated ion channel, the NMDA receptor, expressed heterologously in HEK293 cells or natively in cortical neurons. Also provided are instructions on how to adapt the method to other ion channels of interest by presenting the example of the mechano-sensitive channel PIEZO1. This method can provide data regarding the channel’s conductance properties and the temporal sequence of open-closed conformations that make up the channel’s activation mechanism, thus helping to understand their functions in health and disease.

N-Methyl-D-Aspartate receptors (NMDARs) are ligand-gated ion channels that mediate excitatory neurotransmission in the mammalian central nervous system. They are required for normal neuronal function and are a factor in several neuropathies including Alzheimer's disease and schizophrenia. Classical NMDARs require both glycine (NR1) and glutamate (NR2) bound for receptor activation. Reaction mechanisms for NMDA receptors have been developed for several isoforms, however these models assume saturation of glycine sites and a quantitative understanding of glycine association and dissociation kinetics is currently inadequate. We measured current responses with fast application of glycine (in 1 mM glutamate) onto outside-out patches containing several NR1/NR2A receptors and observed relatively slow kinetics, with a rise time of 4.9 ± 0.6ms and deactivation of 105 ± 8ms. Anticipating a slow dissociation rate for glycine, we first developed models for two low-affinity glycine-site full-agonists: L-serine (95% efficacy; EC50=0.21mM) and 3,3,3-trifluoro-DL-alanine (132% efficacy; EC50=2.2mM). We used these data to select best fitting multi-state kinetic models of several arrangements and further validated these with cycles of simulations and experimental measurements which included macroscopic responses to several stimulation patterns. These models will be used to estimate glycine association and dissociation rate constants. Work is in progress to validate these results with macroscopic measurements. Knowledge about how glycine gates NMDA receptors will offer insight to glycine-dependent NR1/NR2A kinetic mechanisms and contribute to a more comprehensive understanding of the activation of these physiologically important receptors.

GluN1 and GluN3 subunits of the N-methyl-D-aspartate receptor family form tetrameric cation-permeable channels that are gated by glycine alone and are insensitive to glutamate. They are expressed primarily during early development and their role in cellular physiology is unknown. One hypothesis states that their tonic activation by basal levels of brain glycine controls cellular excitability. Notably, GluN1/GluN3A receptors are selectively up-regulated following ischemia, while ‘classical’ GluN2A- and GluN2B-containing receptors are down-regulated and also strongly inhibited by ischemic acidification. We found that extracellular protons strongly potentiated peak glycine-elicited currents from recombinant GluN1/GluN3A receptors (Ipk, 10-fold at pH 6.8 versus pH 8.0) with a half maximal effect in the physiologic range (EC50 = pH 7.1 ± 0.03). The time-course of current desensitization was also significantly prolonged (2-fold at pH 6.8 versus pH 8.0) and the recovery from desensitization was accelerated (2-fold at pH 6.8 versus pH 7.4). In addition, extracellular protons decreased receptor sensitivity to the agonist glycine (EC50, 48 ± 6 µM at pH 7.4 versus 83 ± 3 µM at pH 6.8) and to the endogenous potentiator Zn2+ (EC50, 32 ± 3 µM at pH 7.4 versus 185 ± 26 µM at pH 6.8). Importantly, we found that extracellular acidification during glycine-elicited steady-state activity produced a large transient influx of positive charge (Ipk, 8-fold at pH 6.8 versus pH 7.4) and increased steady-state activity (Iss/Ipk, 0.03 ± 0.01 at pH 7.4 versus 0.5 ± 0.2 at pH 6.8) which depolarized the membrane substantially (−23 ± 5 mV at pH 7.4 versus −13 ± 3 mV at pH 6.8). Taken together, these results indicate that small pH fluctuations potently modulate GluN1/GluN3A receptor currents and that protons may play a novel positive modulatory role at GluN1/GluN3A receptors in vivo by increasing cellular excitability.