In summary, we have identified a regulatory network linking stress stimuli with CRH transcription. As depicted in Figure 7E, our model suggests that in response to various stressors, Otp and the short PAC1 splice variant modulate

transcriptional activation of CRH to adapt to the changes in homeostasis. Otp may contribute to the termination of CRH transcription by regulating the splicing factor A2BP1, which in turn promotes the formation of the long PAC1-hop splice variant. Generation of the long PAC1-hop splice variant terminates both stress-induced CRH transcription and HPA activation by means yet to be uncovered. Stress occurs when an animal’s state of homeostasis is threatened or perceived to be so (Chrousos, 1998, Chrousos, 2009, Engelmann et al., 2004 and Selye, 1936). The adaptive response to most stressors involves the release of CRH followed by rapid changes in its transcription (Aguilera, learn more 1998, Vale et al., 1981 and Yao and Denver, 2007). However, the exact intracellular signaling pathways that modulate CRH synthesis during stress adaptation remain unclear. To date, regulation of CRH transcription has only been addressed using either in vitro cell transfection assays or application of pharmacological agents in animal models. Our study provides pioneering in vivo evidence for a new molecular mechanism of stress adaptation. We show that stress-induced CRH levels are regulated

by the transcription factor Otp and the transmembrane neuropeptide receptor PAC1. We have also demonstrated beta-catenin tumor that the generation of a PAC1 splice variant by means of alternative Linifanib (ABT-869) splicing causes a signaling switch that terminates CRH transcription in response to stress and leads to dysregulated HPA axis response. The activation of CRH transcription following stressful stimuli is a biological response shared by all vertebrate species (Bernier et al., 2009, Burbach, 2002 and Yao and Denver, 2007). This speaks

to the importance of this pathway and implies that an evolutionarily conserved biochemical cascade controls the synthesis of CRH. In this respect, the hypothalamic neuroendocrine-specific factor Otp was an obvious candidate mediator of stress. Otp is expressed in the mature CRH neurons of fish and mouse. Otp deficient mice display impaired development of all hypothalamic neuroendocrine cell types (Acampora et al., 1999 and Wang and Lufkin, 2000). In contrast, we found that the zebrafish otpam866 mutant fish, which carries the partially redundant otpb duplicated paralog, displays normal development of CRH-containing neurons. This has allowed us to examine the role of Otp in mediation of stress response and to demonstrate that it regulates CRH synthesis during stress adaptation. A major finding of this study is that stress response is modulated by a mechanism that involves activity-dependent alternative splicing.