Pituitary Adenylate Cyclase-Activating Polypeptide in the Ventromedial Hypothalamus Is Responsible for Food Intake Behavior by Modulating the Expression of Agouti-Related Peptide in Mice

  • Thanh Trung Nguyen
  • Yuki Kambe
  • Takashi Kurihara
  • Tomoya Nakamachi
  • Norihito Shintani
  • Hitoshi Hashimoto
  • Atsuro MiyataEmail author


Pituitary adenylate cyclase-activating polypeptide (PACAP) is abundantly expressed in the hypothalamus and contributes to hypothalamic functions, including appetite regulation. Although food intake is suggested to be decreased in PACAP (−/−) mice, the detailed mechanisms are still being discussed. We sought to investigate this link. The food consumption at 8 h after refeeding in the (−/−) mice who had fasted for 2 days was significantly lower than in the PACAP (+/+) mice. The nocturnal and daily food intake of (−/−) mice was significantly lower than those of (+/+) mice, but the diurnal food intake showed a tendency to increase. mRNA expression levels of agouti-related peptide (AgRP) were decreased, but those of proopiomelanocortin (POMC) were increased in the hypothalamus of (−/−) mice 4 h after refeeding. Furthermore, intracerebroventricular administration of a PACAP receptor antagonist, PACAP6–38 (1 nmol/4 μL/mouse), decreased food intake and body weight 1, 2, and 4 h after refeeding, as well as expression levels of AgRP at 4 h after refeeding in (+/+) mice. The selective overexpression of PACAP by the infection of an adeno-associated virus in the ventromedial hypothalamus (VMH) resulted in an increase in food intake and AgRP expression in the nocturnal period in addition to the increased food intake at 8 h after refeeding. These results suggest that food intake behavior in mice is triggered by the increase in PACAP expression in the VMH via modulation of AgRP expression in the hypothalamus, pointing to PACAP inhibition as a potential strategy for the development of anti-obesity drugs.


Pituitary adenylate cyclase-activating polypeptide Ventromedial hypothalamus Appetite Agouti-related peptide Food intake 



Agouti-related peptide


Bed nucleus of the stria terminalis


Central amygdala


Cocaine- and amphetamine-regulated transcript


Neuropeptide Y


Steroidogenic factor 1


Pituitary adenylate cyclase-activating polypeptide




Paraventricular hypothalamus


Ventromedial hypothalamus




Wild type



We would like to thank Ms. Izumi Fujisima and Mr. Tetsuya Kawamura for their technical contribution and all the staff members of the Joint Research Laboratory and the Division of Laboratory Animal Sciences, Kagoshima University for their help with animal care and the use of the facilities. We are also grateful to the Ministry of Agriculture and Rural Development, Vietnam, for the doctoral scholarship to T.T.N.

Author Contributions

T.T.N carried out the experiments, performed statistical analysis, and drafted the manuscript. YK carried out the experiments, performed behavioral studies, and wrote the manuscript. TK and AM conceived of and participated in the design of the study and wrote the manuscript. TN, NS, and HH participated in the design of the study and reviewed the manuscript. All authors read and approved of the final manuscript.

Funding Information

This work was supported by a Grant-in-Aid for Scientific Research (C), Japan Society for the Promotion of Science (JSPS) (JSPS KAKENHI Grant No. 17K08310, 17 K08599 and JP19K07121), a Grant-in-Aid for Scientific Research (B) (Grant No. JP17H03989), MEXT KAKENHI, (grant number JP18H05416), and AMED (grant No. JP19dm0107122h0004 and JP19dm0207061h0003).

Compliance with Ethical Standards

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. Specifically, all experiments in the present study were approved by the Experimental Animal Research Committee of Kagoshima University (Approval numbers: MD17054 and MD18105) and the Gene Recombination Experiment Safety Committee of Kagoshima University (Approval number: S28006).

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12035_2019_1864_MOESM1_ESM.png (98 kb)
ESM 1 PACAP (-/-) mice had reduced mesenteric and epididymal fat masses. Fat mass in mesenteric (a) and epididymal tissues (b) under the conditions indicated on the horizontal axis (n = 9 to 11 mice per group). *p<0.05; Student’s t-test (PNG 97 kb)
12035_2019_1864_MOESM2_ESM.png (601 kb)
ESM 2 Successful expression of PACAP after transient transfection of pAAV-CAG::PACAP-IRES-EGFP. (a) HEK293 cells were transiently transfected with pAAV-CAG::IRES-EGFP or pAAV-CAG::PACAP-IRES-EGFP, and mRNA expression levels of PACAP and GAPDH (reference) were quantified by RT-qPCR (n = 2 mice per group). (b) HEK293 cells were transiently transfected with pAAV-CAG::PACAP-IRES-EGFP, and expression of PACAP (red) and GFP (green) was confirmed by immunohistochemistry with each specific antibody. Arrow indicates PACAP immunoreactivity. Scale bars, 10 μm (PNG 601 kb)
12035_2019_1864_MOESM3_ESM.png (104 kb)
ESM 3 PACAP (-/-) mice exhibited enhanced locomotor activity and anti-anxiety-like behaviors compared with their PACAP (+/+) littermates. Distance travelled (a) and time spent in the center region (b) of (+/+) and (-/-) mice in the open-field test. The time spent in the open arms (c) of (+/+) and (-/-) mice in the elevated plus-maze test (n = 5 to 6 mice per group). *p<0.05; Student’s t-test (PNG 104 kb)
12035_2019_1864_MOESM4_ESM.docx (14 kb)
ESM 4 List of primer sequences used for genotyping (DOCX 13 kb)
12035_2019_1864_MOESM5_ESM.docx (16 kb)
ESM 5 List of primer sequences used for RT-qPCR (DOCX 15 kb)


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Copyright information

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Authors and Affiliations

  1. 1.Department of Pharmacology, Graduate School of Medical and Dental ScienceKagoshima UniversityKagoshimaJapan
  2. 2.Department of Pharmacology, Toxicology, Internal Medicine and Diagnostics, Faculty of Veterinary MedicineVietnam National University of AgricultureHanoiVietnam
  3. 3.Laboratory of Regulatory Biology, Graduate School of Science and EngineeringUniversity of ToyamaToyamaJapan
  4. 4.Laboratory of Molecular Neuropharmacology, Graduate School of Pharmaceutical SciencesOsaka UniversitySuitaJapan
  5. 5.Molecular Research Center for Children’s Mental Development, United Graduate School of Child DevelopmentOsaka UniversitySuitaJapan
  6. 6.Division of Bioscience, Institute for Datability ScienceOsaka UniversitySuitaJapan
  7. 7.Transdimensional Life Imaging Division, Institute for Open and Transdisciplinary Research InitiativesOsaka UniversitySuitaJapan

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