Prefrontal brain areas are implicated in the control of fear behavior. However, how prefrontal circuits control fear response to innate threat is poorly understood. Here, we show that the anterior cingulate cortex (ACC) and its input to the basolateral nucleus of amygdala (BLA) contribute to innate fear response to a predator odor in mice. Optogenetic inactivation of the ACC enhances freezing response to fox urine without affecting conditioned freezing. Conversely, ACC stimulation robustly inhibits both innate and conditioned freezing. Circuit tracing and slice patch recordings demonstrate a monosynaptic glutamatergic connectivity of ACC-BLA but no or very sparse ACC input to the central amygdala. Finally, our optogenetic manipulations of the ACC-BLA projection suggest its inhibitory control of innate freezing response to predator odors. Together, our results reveal the role of the ACC and its projection to BLA in innate fear response to olfactory threat stimulus.

No abstract

Memory is thought to be sparsely encoded throughout multiple brain regions forming unique memory trace. Although evidence has established that the amygdala is a key brain site for memory storage and retrieval of auditory conditioned fear memory, it remains elusive whether the auditory brain regions may be involved in fear memory storage or retrieval. To investigate this possibility, we systematically imaged the brain activity patterns in the lateral amygdala, MGm/PIN, and AuV/TeA using activity-dependent induction of immediate early gene zif268 after recent and remote memory retrieval of auditory conditioned fear. Consistent with the critical role of the amygdala in fear memory, the zif268 activity in the lateral amygdala was significantly increased after both recent and remote memory retrieval. Interesting, however, the density of zif268 (+) neurons in both MGm/PIN and AuV/TeA, particularly in layers IV and VI, was increased only after remote but not recent fear memory retrieval compared to control groups. Further analysis of zif268 signals in AuV/TeA revealed that conditioned tone induced stronger zif268 induction compared to familiar tone in each individual zif268 (+) neuron after recent memory retrieval. Taken together, our results support that the lateral amygdala is a key brain site for permanent fear memory storage and suggest that MGm/PIN and AuV/TeA might play a role for remote memory storage or retrieval of auditory conditioned fear, or, alternatively, that these auditory brain regions might have a different way of processing for familiar or conditioned tone information at recent and remote time phases.

Alleviating pain with controlled breathing has been practiced throughout human history. Despite its wide use and long history, a neural circuit-based understanding of the pain-breathing interaction is largely lacking.Here we report a novel breathing circuit that regulates non-homeostatic breathing rhythm, as well as pain and anxiety. We identify that a cluster of neurons expressing the Oprm1 gene, which encodes the µ-opioid receptor (MOR) in the lateral subdivision of parabrachial nucleus (PBL Oprm1 ), directly regulates breathing rate in mice by conveying signals from the limbic areas to respiratory rhythm generating neurons in the medullary preBötzinger Complex (preBötC). In addition, we found that pain signals rapidly increase breathing rate by activating these neurons in both awake and anesthetized mice. Inactivating these neurons not only decreases the breathing rate, but it also substantially decreases anxiety-like behaviors and induces strong appetitive behaviors. Furthermore, PBL Oprm1 inactivation alleviates pain by attenuating the perception of the affective-motivational aspect of pain. These results suggest that PBL Oprm1 neurons play a critical role in the non-homeostatic regulation of breathing and in the regulation of pain and anxiety through breathing.

Neurons produce and release neuropeptides to communicate with one another. Despite their profound impact on critical brain functions, circuit-based mechanisms of peptidergic transmission are poorly understood, primarily due to the lack of tools for monitoring and manipulating neuropeptide release in vivo. Here, we report the development of two genetically encoded tools for investigating peptidergic transmission in behaving mice: a genetically encoded large dense core vesicle (LDCV) sensor that detects the neuropeptides release presynaptically, and a genetically encoded silencer that specifically degrades neuropeptides inside the LDCV. Monitoring and silencing peptidergic and glutamatergic transmissions from presynaptic terminals using our newly developed tools and existing genetic tools, respectively, reveal that neuropeptides, not glutamate, are the primary transmitter in encoding unconditioned stimulus during Pavlovian threat learning. These results show that our sensor and silencer for peptidergic transmission are reliable tools to investigate neuropeptidergic systems in awake behaving animals.