Regulating anxiety with extrasynaptic inhibition

Aversive experiences can lead to complex behavioral adaptations including increased levels of anxiety and fear generalization. The neuronal mechanisms underlying such maladaptive behavioral changes, however, are poorly understood. Here, using a combination of behavioral, physiological and optogenetic approaches in mouse, we identify a specific subpopulation of central amygdala neurons expressing protein kinase C δ (PKCδ) as key elements of the neuronal circuitry controlling anxiety. Moreover, we show that aversive experiences induce anxiety and fear generalization by regulating the activity of PKCδ+ neurons via extrasynaptic inhibition mediated by α5 subunit-containing GABAA receptors. Our findings reveal that the neuronal circuits that mediate fear and anxiety overlap at the level of defined subpopulations of central amygdala neurons and demonstrate that persistent changes in the excitability of a single cell type can orchestrate complex behavioral changes

Long-Term Memory for Pavlovian Fear Conditioning Requires Dopamine in the Nucleus Accumbens and Basolateral Amygdala

The neurotransmitter dopamine (DA) is essential for learning in a Pavlovian fear conditioning paradigm known as fear-potentiated startle (FPS). Mice lacking the ability to synthesize DA fail to learn the association between the conditioned stimulus and the fear-inducing footshock. Previously, we demonstrated that restoration of DA synthesis to neurons of the ventral tegmental area (VTA) was sufficient to restore FPS. Here, we used a target-selective viral restoration approach to determine which mesocorticolimbic brain regions receiving DA signaling from the VTA require DA for FPS. We demonstrate that restoration of DA synthesis to both the basolateral amygdala (BLA) and nucleus accumbens (NAc) is required for long-term memory of FPS. These data provide crucial insight into the dopamine-dependent circuitry involved in the formation of fear-related memory

Transbilayer Phospholipid Movements in ABCA1-Deficient Cells

Tangier disease is an inherited disorder that results in a deficiency in circulating levels of HDL. Although the disease is known to be caused by mutations in the ABCA1 gene, the mechanism by which lesions in the ABCA1 ATPase effect this outcome is not known. The inability of ABCA1 knockout mice (ABCA1−/−) to load cholesterol and phospholipids onto apoA1 led to a proposal that ABCA1 mediates the transbilayer externalization of phospholipids, an activity integral not only to the formation of HDL particles but also to another, distinct process: the recognition and clearance of apoptotic cells by macrophages. Expression of phosphatidylserine (PS) on the surface of both macrophages and their apoptotic targets is required for efficient engulfment of the apoptotic cells, and it has been proposed that ABCA1 is required for transbilayer externalization of PS to the surface of both cell types. To determine whether ABCA1 is responsible for any of the catalytic activities known to control transbilayer phospholipid movements, these activities were measured in cells from ABCA1−/− mice and from Tangier individuals as well as ABCA1-expressing HeLa cells. Phospholipid movements in either normal or apoptotic lymphocytes or in macrophages were not inhibited when cells from knockout and wildtype mice or immortalized cells from Tangier individuals vs normal individuals were compared. Exposure of PS on the surface of normal thymocytes, apoptotic thymocytes and elicited peritoneal macrophages from wildtype and knockout mice or B lymphocytes from normal and Tangier individuals, as measured by annexin V binding, was also unchanged. No evidence was found of ABCA1-stimulated active PS export, and spontaneous PS movement to the outer leaflet in the presence or absence of apoA1 was unaffected by the presence or absence of ABCA1. Normal or Tangier B lymphocytes and macrophages were also identical in their ability to serve as targets or phagocytes, respectively, in apoptotic cell clearance assays. No evidence was found to support the suggestion that ABCA1 is involved in transport to the macrophage cell surface of annexins I and II, known to enhance phagocytosis of apoptotic cells. These results show that mutations in ABCA1 do not measurably reduce the rate of transbilayer movements of phospholipids in either the engulfing macrophage or the apoptotic target, thus discounting catalysis of transbilayer movements of phospholipids as the mechanism by which ABCA1 facilitates loading of phospholipids and cholesterol onto apoA1

Requirement of dopamine signaling in the amygdala and striatum for learning and maintenance of a conditioned avoidance response

Two-way active avoidance (2WAA) involves learning Pavlovian (association of a sound cue with a foot shock) and instrumental (shock avoidance) contingencies. To identify regions where dopamine (DA) is involved in mediating 2WAA, we restored DA signaling in specific brain areas of dopamine-deficient (DD) mice by local reactivation of conditionally inactivated Th genes using viral gene therapy. Among all targeted areas—prefrontal cortex (PFC), amygdala, ventral striatum, dorsal striatum, and whole striatum—only restoration of DA signaling to both the whole striatum together with the amygdala enabled DD mice to acquire 2WAA. However, after prolonged overtraining during which DD mice had DA synthesis systemically reconstituted pharmacologically with L-3,4-dihydroxyphenylalanine (L-Dopa), DA signaling in the striatum alone was sufficient to maintain 2WAA, whereas DA signaling in the PFC together with the amygdala was insufficient to maintain 2WAA. Our results indicate that learning 2WAA requires DA signaling in both the amygdala and the entire striatum; however, after sufficient training, DA signaling in the striatum alone can maintain the learned avoidance behavior

Temporal dynamics of affect in the brain: Evidence from human imaging and animal models

•Affective states are understood to emerge and persist through patterns of neural activity.•Human and animal neuroscience research on how the brain directs the unfolding of emotion over time are scarcely integrated.•We highlight the specific neural mechanisms and modulators that arbitrate emotional state rise-time, intensity, and duration. Emotions are time-varying internal states that promote survival in the face of dynamic environments and shifting homeostatic needs. Research in non-human organisms has recently afforded specific insights into the neural mechanisms that support the emergence, persistence, and decay of affective states. Concurrently, a separate affective neuroscience literature has begun to dissect the neural bases of affective dynamics in humans. However, the circuit-level mechanisms identified in animals lack a clear mapping to the human neuroscience literature. As a result, critical questions pertaining to the neural bases of affective dynamics in humans remain unanswered. To address these shortcomings, the present review integrates findings from humans and non-human organisms to highlight the neural mechanisms that govern the temporal features of emotional states. Using the theory of affective chronometry as an organizing framework, we describe the specific neural mechanisms and modulatory factors that arbitrate the rise-time, intensity, and duration of emotional states

Selective restoration of TH in virally-rescued DD mice.

<p><i>A–E)</i> Immunohistochemistry (IHC) results for a control mouse. <i>A)</i> Tyrosine hydroxylase (TH) stain in the nucleus accumbens (NAc). <i>B)</i> 40x magnification of TH stain. <i>C)</i> Stain for dopamine transporter (DAT) in NAc. <i>D)</i> Merged image of TH and DAT stain showing extensive overlap of the two signals. <i>E)</i> TH stain in basolateral amygdala (BLA) shown at 20x. <i>F–J)</i> IHC results for a non-rescued DD mouse. <i>F)</i> Complete absence of TH in NAc. <i>G)</i> 40x magnification demonstrating lack of TH in NAc. <i>H)</i> DAT stain. <i>I)</i> Merged image of TH and DAT illustrating lack of TH in the NAc. <i>J)</i> TH in BLA (20x) is almost undetectable. <i>K–O)</i> Representative IHC from NAc-rescued DD mouse. <i>K)</i> TH was largely restored in the NAc. <i>L)</i> 40x magnification illustrating rescue of TH. <i>M)</i> DAT staining in NAc-rescued DD mouse. <i>N)</i> Merged image of TH and DAT illustrating large extent of TH restoration in NAc. <i>O)</i> TH in BLA (20x) remains at low, non-injected levels. <i>P–T)</i> Representative IHC from DD mouse injected into the NAc and BLA. <i>P)</i> Robust restoration of TH to the NAc. <i>Q)</i> 40x magnification demonstrating TH rescue. <i>R)</i> DAT staining. <i>S)</i> Merged image of TH and DAT showing extensive restoration of TH to NAc. <i>T)</i> TH is restored to higher levels in NAc and BLA rescued DD mice.</p

HPLC Quantification of DA, NE, and DA metabolites.

<p>DA, dopamine; HVA, homovanillic acid; DOPAC, 3.4-dihydroxyphenylacetic acid; 3-MT, 3-methoxytyramine. Data presented as means ± SEM.</p

Effects of footshock stress on social behavior and neuronal activation in the medial prefrontal cortex and amygdala of male and female mice.

Social behavior is complex and fundamental, and its deficits are common pathological features for several psychiatric disorders including anxiety, depression, and posttraumatic stress disorder. Acute stress may have a negative impact on social behavior, and these effects can vary based on sex. The aim of this study was to explore the effect of acute footshock stress, using analogous parameters to those commonly used in fear conditioning assays, on the sociability of male and female C57BL/6J mice in a standard social approach test. Animals were divided into two main groups of footshock stress (22 male, 24 female) and context exposed control (23 male and 22 female). Each group had mice that were treated intraperitoneally with either the benzodiazepine-alprazolam (control: 10 male, 10 female; stress: 11 male, 11 female), or vehicle (control: 13 male, 12 female; stress: 11 male, 13 female). In all groups, neuronal activation during social approach was assessed using immunohistochemistry against the immediate early gene product cFos. Although footshock stress did not significantly alter sociability or latency to approach a social stimulus, it did increase defensive tail-rattling behavior specifically in males (p = 0.0022). This stress-induced increase in tail-rattling was alleviated by alprazolam (p = 0.03), yet alprazolam had no effect on female tail-rattling behavior in the stress group. Alprazolam lowered cFos expression in the medial prefrontal cortex (p = 0.001 infralimbic area, p = 0.02 prelimbic area), and social approach induced sex-dependent differences in cFos activation in the ventromedial intercalated cell clusters (p = 0.04). Social approach following stress-induced cFos expression was positively correlated with latency to approach and negatively correlated with sociability in the prelimbic area and multiple amygdala subregions (all p < 0.05). Collectively, our results suggest that acute footshock stress induces sex-dependent alterations in defensiveness and differential patterns of cFos activation during social approach

Nat Commun.

Fear extinction is an adaptive process whereby defensive responses are attenuated following repeated experience of prior fear-related stimuli without harm. The formation of extinction memories involves interactions between various corticolimbic structures, resulting in reduced central amygdala (CEA) output. Recent studies show, however, the CEA is not merely an output relay of fear responses but contains multiple neuronal subpopulations that interact to calibrate levels of fear responding. Here, by integrating behavioural, in vivo electrophysiological, anatomical and optogenetic approaches in mice we demonstrate that fear extinction produces reversible, stimulus- and context-specific changes in neuronal responses to conditioned stimuli in functionally and genetically defined cell types in the lateral (CEl) and medial (CEm) CEA. Moreover, we show these alterations are absent when extinction is deficient and that selective silencing of protein kinase C delta-expressing (PKC?) CEl neurons impairs fear extinction. Our findings identify CEA inhibitory microcircuits that act as critical elements within the brain networks mediating fear extinction.Amygdala Circuits for Appetitive Conditionin