© 2016 Nature America, Inc. All rights reserved. NATURE NEUROSCIENCE ADVANCE ONLINE PUBLICATION 1 ARTICLES Innate fear is a basic and natural mechanism by which animals and humans avoid danger. This emotion is trigged by a threat perceived through sensory stimuli, which usually causes a quick response, such as freezing, flight or hiding, and thus has a profound role in survival and health 1,2 . Abnormal innate fear in humans, particularly phobias and panic disorders 1–3 , is strongly associated with anxiety disorders. The features of heritability and natural acquisition distinguish innate fear from conditioned fear. Unlike the innate fear circuitry, extensive studies have shown that the amygdala, hippocampus and prefrontal cortex are crucial for conditioned fear 4–6 . Emerging evidence sug- gests that distinct neuronal circuits are responsible for fear induced by different cues. For example, the lateral and central nuclei of the amygdala have a crucial role in conditioned fear, but not in the innate fear induced by an olfactory cue 7,8 . Although the cortical area of the amygdala has recently been shown to be responsible for innate fear induced by an olfactory cue 9 , and a number of studies have reported the expression pattern of the immediate-early gene c-fos in the rodent brain in response to TMT or the odor of cat 10 , the underlying neuronal circuitry is still largely unknown. RESULTS Activation of GABAergic interneurons in the LDT produces fear To explore the neuronal circuitry for olfactory cue–induced innate fear in rodents, we first studied c-Fos protein expression in the whole brain when provoked by the pungent odor TMT. Previous studies have shown that TMT effectively induces fear behavior–like freezing in naive mice and can therefore be used to study the neurobiology of innate fear 8 . As a result of its unpleasant smell, we assumed that TMT would activate neuronal circuits in addition to those for innate fear, such as olfaction. Thus, as well as saline controls, we used β-mercaptoethanol (ME), which also has a pungent odor, but does not induce freezing behavior in mice, as a positive control. Compared with saline and ME stimuli, we found that TMT exposure specifically induced massive c-Fos expression in the LDT (Fig. 1a), the lateral hypotha- lamic area (LH), the dorsomedial part of the ventromedial hypotha- lamic nucleus (VMHDM) and the LHb (Supplementary Fig. 1a–d). We noted that the olfactory bulb was activated by both TMT and ME, with a greater increase of c-Fos–positive neurons in response to TMT (Supplementary Fig. 1a,d). Previous studies have shown that the hypothalamus is critical for regulating the autonomic nervous system, including heart rate and hormone release 1,11 , as well as being involved in innate defensive behaviors 11–14 . We then focused on the LDT, as no evidence has yet linked this nucleus to fear. We found that the majority of c-Fos–positive neurons in the LDT expressed GAD67, suggesting that they are GABAergic neurons (Fig. 1a,b). Next, we determined whether selectively activating GABAergic transmission in the LDT using optogenetics is sufficient to induce fear-like behavior, as judged by four criteria we summarized from pre- vious studies 1,15–17 : defensive behaviors (typically freezing), changing autonomic function such as heart rate and defecation, release of corticosterone, and anxiety-like behavior after prolonged stimula- tion. In VGAT-ChR2(H134R)-eYFP (abbreviated to VGAT-ChR2) 1 Department of Neurobiology, Key Laboratory of Medical Neurobiology (Ministry of Health of China), Key Laboratory of Neurobiology of Zhejiang Province, Zhejiang School of Medicine, Hangzhou, China. 2 Department of Neurology, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China. 3 Key Laboratory of Magnetic Resonance in Biological Systems and State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Institute of Physics and Mathematics, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Wuhan, China. 4 Wuhan National Laboratory for Optoelectronics, Wuhan, China. 5 These authors contributed equally to this work. Correspondence should be addressed to S.D. (duanshumin@zju.edu.cn) or H.W. (haowang@zju.edu.cn). Received 30 July 2015; accepted 30 November 2015; published online 4 January 2016; doi:10.1038/nn.4208 Laterodorsal tegmentum interneuron subtypes oppositely regulate olfactory cue-induced innate fear Hongbin Yang 1,5 , Junhua Yang 1,5 , Wang Xi 1 , Sijia Hao 1 , Benyan Luo 2 , Xiaobin He 3 , Liya Zhu 1 , Huifang Lou 1 , Yan-qin Yu 1 , Fuqiang Xu 3,4 , Shumin Duan 1 & Hao Wang 1 Innate fear has a critical role in survival of animals. Unlike conditioned fear, the neuronal circuitry underlying innate fear is largely unknown. We found that the laterodorsal tegmentum (LDT) and lateral habenula (LHb) are specifically activated by the mouse predator odorant trimethylthiazoline (TMT). Using optogenetics to selectively stimulate GABAergic neurons in the LDT immediately produced fear-like responses (freezing, accelerated heart rate and increased serum corticosterone), whereas prolonged stimulation caused anxiety-like behaviors. Notably, although selective stimulation of parvalbumin (PV)-positive interneurons similarly induced fear-like responses, stimulation of somatostatin-positive interneurons or inhibition of PV neurons in the LDT suppressed TMT-induced fear-like responses without affecting conditioned fear. Finally, activation of LHb glutamatergic inputs to LDT interneurons was sufficient to generate fear-like responses. Thus, the LHb-LDT pathway is important for regulating olfactory cue–induced innate fear. Our results provide a potential target for therapeutic intervention for anxiety disorder.