A Central Amygdala-Ventrolateral Periaqueductal Gray Matter Pathway for Pain in a Mouse Model of Depression-like Behavior.

BACKGROUND: The mechanisms underlying depression-associated pain remain poorly understood. Using a mouse model of depression, the authors hypothesized that the central amygdala-periaqueductal gray circuitry is involved in pathologic nociception associated with depressive states. METHODS: The authors used chronic restraint stress to create a mouse model of nociception with depressive-like behaviors. They then used retrograde tracing strategies to dissect the pathway from the central nucleus of the amygdala to the ventrolateral periaqueductal gray. The authors performed optogenetic and chemogenetic experiments to manipulate the activity of this pathway to explore its roles for nociception. RESULTS: The authors found that γ-aminobutyric acid-mediated (GABAergic) neurons from the central amygdala project onto GABAergic neurons of the ventrolateral periaqueductal gray, which, in turn, locally innervate their adjacent glutamatergic neurons. After chronic restraint stress, male mice displayed reliable nociception (control, mean ± SD: 0.34 ± 0.11 g, n = 7 mice; chronic restraint stress, 0.18 ± 0.11 g, n = 9 mice, P = 0.011). Comparable nociception phenotypes were observed in female mice. After chronic restraint stress, increased circuit activity was generated by disinhibition of glutamatergic neurons of the ventrolateral periaqueductal gray by local GABAergic interneurons via receiving enhanced central amygdala GABAergic inputs. Inhibition of this circuit increased nociception in chronic restraint stress mice (median [25th, 75th percentiles]: 0.16 [0.16, 0.16] g to 0.07 [0.04, 0.16] g, n = 7 mice per group, P < 0.001). In contrast, activation of this pathway reduced nociception (mean ± SD: 0.16 ± 0.08 g to 0.34 ± 0.13 g, n = 7 mice per group, P < 0.001). CONCLUSIONS: These findings indicate that the central amygdala-ventrolateral periaqueductal gray pathway may mediate some aspects of pain symptoms under depression conditions.

Activation of the neural pathway from the dorsolateral bed nucleus of the stria terminalis to the central amygdala induces anxiety-like behaviors.

The bed nucleus of the stria terminalis (BNST) and the central amygdala (CeA) comprise a forebrain unit that has been described as the "extended amygdala". These two nuclei send dense projections to each other and have been implicated in the regulation of negative emotional states, including anxiety and fear. The present study employed an optogenetic technique to examine whether stimulation of CeA-projecting dorsolateral BNST (dlBNST) neuron terminals would influence anxiety-like behaviors in male Sprague-Dawley rats. Photostimulation of CeA-projecting dlBNST neuron terminals produced anxiogenic effects in an elevated plus maze test. This finding is inconsistent with previous reports showing that optogenetic stimulation of BNST neurons projecting to the lateral hypothalamus (LH) and ventral tegmental area (VTA) produces anxiolytic rather than anxiogenic effects. To address this issue, electrophysiological analyses were conducted to characterize dlBNST neurons projecting to the CeA, LH, and VTA. dlBNST neurons can be electrophysiologically classified into three distinct cell types (types I-III) according to their responses to depolarizing and hyperpolarizing current injections. Whole-cell patch-clamp recordings revealed that more than 60% of the CeA-projecting dlBNST neurons were type II, whereas approximately 80% of the LH- and VTA-projecting dlBNST neurons were type III. These electrophysiological results will help elucidate the mechanisms underlying the heterogeneity of BNST neurons during the regulation of anxiety-like behaviors.

Differential efferent projections of GABAergic neurons in the basolateral and central nucleus of amygdala in mice.

The Basolateral amygdala (BLA) and central nucleus of the amygdala (CEA) have been proved to play a key role in the control of anxiety, stress and fear-related behaviors. BLA is a cortex-like complex consisting of both γ-aminobutyric acidergic (GABAergic) interneurons and glutamatergic neurons. The CEA is a striatum-like output of the amygdala, consisting almost exclusively of GABAergic medium spiny neurons. In this study, we explored the morphology and axonal projections of the GABAergic neurons in BLA and CEA, using conditional anterograde axonal tracing, immunohistochemistry, and VGAT-Cre transgenic mice to further understand their functional roles. We found that the axonal projections of GABAergic neurons from the BLA mainly distributed to the forebrain, whilst GABAergic neurons from the CEA distributed to the forebrain, midbrain and brainstem. In the forebrain, the axonal projections of GABAergic neurons from the BLA projected to the anterior olfactory nucleus, the cerebral cortex, the septum, the striatum, the thalamus, the amygdala and the hippocampus. The axonal projections of GABAergic neurons from the CEA distributed to the nuclei of the prefrontal cortex, the bed nucleus of the stria terminalis, the hypothalamus and the thalamus. In the midbrain and brainstem, the axonal projections of GABAergic neurons from the CEA were found in the periaqueductal gray, the substantia nigra, and the locus coeruleus. These data reveal the neuroanatomical basis for exploring the function of GABAergic neurons in the BLA and CEA, particularly during the processing of fear-related behavior.

The paraventricular thalamus input to central amygdala controls depression-related behaviors.

The dysregulation of neuronal networks may contribute to the etiology of major depressive disorder (MDD). However, the neural connections underlying the symptoms of MDD have yet to be elucidated. Here, we observed that glutamatergic neurons in the paraventricular thalamus (PVT) were activated by chronic unpredictable stress (CUS) with higher expression numbers of ΔFosB-labeled neurons and protein expression levels, activation of PVT neurons caused depressive-like phenotypes, whereas suppression of PVT neuronal activity induced an antidepressant effect in male, but not female mice, which were achieved by using a chemogenetic approach. Moreover, we found that PVT glutamatergic neurons showed strong neuronal projections to the central amygdala (CeA), activation of the CeA-projecting neurons in PVT or the neuronal terminals of PVT-CeA projection neurons induced depression-related behaviors or showed enhanced stress-induced susceptibility. These results suggest that PVT is a key depression-controlling nucleus, and PVT-CeA projection regulates depression-related behaviors in a sex-dependent manner, which could be served as an essential pathway for morbidity and treatment of depression.

Genetic recombination in disgust-associated bitter taste-responsive neurons of the central nucleus of amygdala in male mice.

A bitter substance induces specific orofacial and somatic behavioral reactions such as gapes in mice as well as monkeys and humans. These reactions have been proposed to represent affective disgust, and therefore, understanding the neuronal basis of the reactions would pave the way to understand affective disgust. It is crucial to identify and access the specific neuronal ensembles that are activated by bitter substances, such as quinine, the intake of which induces disgust reactions. However, the method to access the quinine-activated neurons has not been fully established yet. Here, we show evidence that a targeted recombination in active populations (TRAP) method, induces genetic recombination in the quinine-activated neurons in the central nucleus of the amygdala (CeA). CeA is one of the well-known emotional centers of the brain. We found that the intraoral quinine infusion, that resulted in disgust reactions, increased both cFos-positive cells and Arc-positive cells in the CeA. By using Arc-CreER;Ai3 TRAP mice, we induced genetic recombination in the quinine-activated neurons and labelled them with fluorescent protein. We confirmed that the quinine-TRAPed fluorescently-labelled cells preferentially coexpressed Arc after quinine infusion. Our results suggest that the TRAP method can be used to access specific functional neurons in the CeA.