Secretin Prevents Apoptosis in the Developing Cerebellum Through Bcl-2 and Bcl-xL

  • Lei Wang
  • Li ZhangEmail author
  • Billy K. C. ChowEmail author


Secretin (SCT) is involved in a variety of physiological processes and has been implicated in preventing apoptosis during brain development. However, little is known about the molecular mechanism underlying its neuroprotective effects. The B cell lymphoma 2 (Bcl-2) family proteins, such as Bcl-2 and Bcl-xL, determine the commitment of neurons to apoptosis. In SCT knockout mice, we found reduced transcript levels of anti-apoptotic genes Bcl-2 and Bcl-xL, but not of pro-apoptotic gene Bax, in the developing cerebellum. SCT treatment on ex vivo cultured cerebellar slices triggered a time-dependent increase of Bcl-2 and Bcl-xL expression. This SCT-induced transcriptional regulation of Bcl-2 and Bcl-xL was dependent on the cyclic AMP (cAMP) response element-binding protein (CREB), which is a key survival factor at the convergence of multiple signaling cascades. We further demonstrated that activation of CREB by SCT was mediated by cAMP/protein kinase A (PKA) and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase 1/2 (ERK1/2) cascades. These findings, collectively, provide an uncharacterized signaling cascade for SCT-mediated neuronal survival, in which SCT promotes the key anti-apoptotic elements Bcl-2 and Bcl-xL in the intrinsic death pathway through PKA- and ERK-regulated CREB phosphorylation.


Secretin Apoptosis Bcl-2 family proteins CREB PKA ERK1/2 


Funding Information

This work is supported by Hong Kong Research Grant Council grant GRF 765113M to Professor B.K.C. Chow, and National Natural Science Foundation of China (#31500842) to Dr. L. Zhang.

Compliance with Ethical Standards

All animal experiments were conducted according to the protocols approved by Committee on the Use of Living Animals in Teaching and Research at the University of Hong Kong.

Conflict of Interest Statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


  1. Ballif BA, Blenis J (2001) Molecular mechanisms mediating mammalian mitogen-activated protein kinase (MAPK) kinase (MEK)-MAPK cell survival signals. Cell Growth Differ 12:397–408Google Scholar
  2. Bhave S, Hoffman P (2004) Phosphatidylinositol 3′-OH kinase and protein kinase a pathways mediate the anti-apoptotic effect of pituitary adenylyl cyclase-activating polypeptide in cultured cerebellar granule neurons: modulation by ethanol. J Neurochem 88:359–369. CrossRefGoogle Scholar
  3. Castorina A, Tiralongo A, Giunta S, Carnazza ML, Rasi G, D'Agata V (2008) PACAP and VIP prevent apoptosis in schwannoma cells. Brain Res 1241:29–35. CrossRefGoogle Scholar
  4. Cheng EHYA, Wei MC, Weiler S, Flavell RA, Mak TW, Lindsten T, Korsmeyer SJ (2001) BCL-2, BCL-XL sequester BH3 domain-only molecules preventing BAX- and BAK-mediated mitochondrial apoptosis. Mol Cell 8:705–711. CrossRefGoogle Scholar
  5. Czabotar PE, Lessene G, Strasser A, Adams JM (2014) Control of apoptosis by the BCL-2 protein family: implications for physiology and therapy. Nat Rev Mol Cell Biol 15:49–63CrossRefGoogle Scholar
  6. Dohi K, Mizushima H, Nakajo S, Ohtaki H, Matsunaga S, Aruga T, Shioda S (2002) Pituitary adenylate cyclase-activating polypeptide (PACAP) prevents hippocampal neurons from apoptosis by inhibiting JNK/SAPK and p38 signal transduction pathways. Regul Pept 109:83–88. CrossRefGoogle Scholar
  7. Du C, Fang M, Li Y, Li L, Wang X (2000) Smac, a mitochondrial protein that promotes cytochrome c–dependent caspase activation by eliminating IAP inhibition. Cell 102:33–42CrossRefGoogle Scholar
  8. Eskes R, Desagher S, Antonsson B, Martinou J-C (2000) Bid induces the oligomerization and insertion of Bax into the outer mitochondrial membrane. Mol Cell Biol 20:929–935CrossRefGoogle Scholar
  9. Falluel-Morel A, Aubert N, Vaudry D, Basille M, Fontaine M, Fournier A, Vaudry H, Gonzalez BJ (2004) Opposite regulation of the mitochondrial apoptotic pathway by C2-ceramide and PACAP through a MAP-kinase-dependent mechanism in cerebellar granule cells. J Neurochem 91:1231–1243CrossRefGoogle Scholar
  10. Fang X, Yu S, Eder A, Mao M, Bast RC Jr, Boyd D, Mills GB (1999) Regulation of BAD phosphorylation at serine 112 by the Ras-mitogen-activated protein kinase pathway. Oncogene 18:6635–6640CrossRefGoogle Scholar
  11. Fletcher JI, Meusburger S, Hawkins CJ, Riglar DT, Lee EF, Fairlie WD, Huang DCS, Adams JM (2008) Apoptosis is triggered when prosurvival Bcl-2 proteins cannot restrain Bax. Proc Natl Acad Sci U S A 105:18081–18087CrossRefGoogle Scholar
  12. Fuchs Y, Steller H (2011) Programmed cell death in animal development and disease. Cell 147:742–758. CrossRefGoogle Scholar
  13. Ghoumari AM, Wehrlé R, Bernard O, Sotelo C, Dusart I (2000) Implication of Bcl-2 and Caspase-3 in age-related Purkinje cell death in murine organotypic culture: an in vitro model to study apoptosis. Eur J Neurosci 12:2935–2949CrossRefGoogle Scholar
  14. Gutiérrez-Cañas I, Rodríguez-Henche N, Bolaños O, Carmena MJ, Prieto JC, Juarranz MG (2003) VIP and PACAP are autocrine factors that protect the androgen-independent prostate cancer cell line PC-3 from apoptosis induced by serum withdrawal. Brit J Pharmacol 139:1050–1058. CrossRefGoogle Scholar
  15. Heaton MB, Moore DB, Paiva M, Gibbs T, Bernard O (1999) Bcl-2 overexpression protects the neonatal cerebellum from ethanol neurotoxicity. Brain Res 817:13–18. CrossRefGoogle Scholar
  16. Hurtado de Mendoza T, Balana B, Slesinger PA, Verma IM (2011) Organotypic cerebellar cultures: apoptotic challenges and detection. J Vis Exp.
  17. Hwang DW, Givens B, Nishijima I (2009) Ethanol-induced developmental neurodegeneration in secretin receptor-deficient mice. Neuroreport 20:698–701. CrossRefGoogle Scholar
  18. Jukkola PI, Rogers JT, Kaspar BK, Weeber EJ, Nishijima I (2011) Secretin deficiency causes impairment in survival of neural progenitor cells in mice. Hum Mol Genet 20:1000–1007. CrossRefGoogle Scholar
  19. Kerr JF, Wyllie AH, Currie AR (1972) Apoptosis: a basic biological phenomenon with wideranging implications in tissue kinetics. Brit J Cancer 26:239–257CrossRefGoogle Scholar
  20. Kim HS, Yumkham S, Kim SH, Yea K, Shin YC, Ryu SH, Suh PG (2006) Secretin induces neurite outgrowth of PC12 through cAMP-mitogen-activated protein kinase pathway. Exp Mol Med 38:85–93. CrossRefGoogle Scholar
  21. Kim S-J, Nian C, Widenmaier S, McIntosh CH (2008) Glucose-dependent insulinotropic polypeptide-mediated up-regulation of β-cell antiapoptotic Bcl-2 gene expression is coordinated by cyclic AMP (cAMP) response element binding protein (CREB) and cAMP-responsive CREB coactivator 2. Mol Cell Biol 28:1644–1656CrossRefGoogle Scholar
  22. Kluck RM, Bossy-Wetzel E, Green DR, Newmeyer DD (1997) The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science 275:1132–1136CrossRefGoogle Scholar
  23. Kluck RM, Esposti MD, Perkins G, Renken C, Kuwana T, Bossy-Wetzel E, Goldberg M, Allen T, Barber MJ, Green DR, Newmeyer DD (1999) The pro-apoptotic proteins, bid and Bax, cause a limited permeabilization of the mitochondrial outer membrane that is enhanced by cytosol. J Cell Biol 147:809–822CrossRefGoogle Scholar
  24. Kuwana T, Bouchier-Hayes L, Chipuk JE, Bonzon C, Sullivan BA, Green DR, Newmeyer DD (2005) BH3 domains of BH3-only proteins differentially regulate Bax-mediated mitochondrial membrane permeabilization both directly and indirectly. Mol Cell 17:525–535CrossRefGoogle Scholar
  25. Lee VH, Lee LT, Chu JY, Lam IP, Siu FK, Vaudry H, Chow BK (2010) An indispensable role of secretin in mediating the osmoregulatory functions of angiotensin II. FASEB J 24:5024–5032CrossRefGoogle Scholar
  26. Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES, Wang X (1997) Cytochrome c and dATP-dependent formation of Apaf-1/Caspase-9 complex initiates an apoptotic protease Cascade. Cell 91:479–489. CrossRefGoogle Scholar
  27. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods 25:402–408CrossRefGoogle Scholar
  28. Luciano F, Jacquel A, Colosetti P, Herrant M, Cagnol S, Pages G, Auberger P (2003) Phosphorylation of Bim-EL by Erk1/2 on serine 69 promotes its degradation via the proteasome pathway and regulates its proapoptotic function. Oncogene 22:6785–6793CrossRefGoogle Scholar
  29. May V, Lutz E, MacKenzie C, Schutz KC, Dozark K, Braas KM (2010) Pituitary adenylate cyclase-activating polypeptide (PACAP)/PAC1HOP1 receptor activation coordinates multiple neurotrophic signaling pathways: Akt activation through phosphatidylinositol 3-kinase gamma and vesicle endocytosis for neuronal survival. J Biol Chem 285:9749–9761. CrossRefGoogle Scholar
  30. Meier P, Finch A, Evan G (2000) Apoptosis in development. Nature 407:796–801CrossRefGoogle Scholar
  31. Merry DE, Veis DJ, Hickey WF, Korsmeyer SJ (1994) Bcl-2 protein expression is widespread in the developing nervous system and retained in the adult PNS. Development 120:301–311Google Scholar
  32. Opferman JT, Kothari A (2017) Anti-apoptotic BCL-2 family members in development. Cell Death Differ 25:37–45. CrossRefGoogle Scholar
  33. Riccio A, Ahn S, Davenport CM, Blendy JA, Ginty DD (1999) Mediation by a CREB family transcription factor of NGF-dependent survival of sympathetic neurons. Science 286:2358–2361CrossRefGoogle Scholar
  34. Virdee K, Parone PA, Tolkovsky AM (2000) Phosphorylation of the pro-apoptotic protein BAD on serine 155, a novel site, contributes to cell survival. Curr Biol 10:1151–1154. CrossRefGoogle Scholar
  35. Wang L, Zhang L, Chow BKC (2017) Secretin modulates the postnatal development of mouse cerebellar cortex via PKA- and ERK-dependent pathways. Front Cell Neurosci 11.
  36. Wei MC, Lindsten T, Mootha VK, Weiler S, Gross A, Ashiya M, Thompson CB, Korsmeyer SJ (2000) tBID, a membrane-targeted death ligand, oligomerizes BAK to release cytochrome c. Genes Dev 14:2060–2071Google Scholar
  37. Weston CR, Balmanno K, Chalmers C, Hadfield K, Molton SA, Ley R, Wagner EF, Cook SJ (2003) Activation of ERK1/2 by ΔRaf-1: ER* represses Bim expression independently of the JNK or PI3K pathways. Oncogene 22:1281–1293CrossRefGoogle Scholar
  38. Willis SN, Fletcher JI, Kaufmann T, van Delft MF, Chen L, Czabotar PE, Ierino H, Lee EF, Fairlie WD, Bouillet P, Strasser A, Kluck RM, Adams JM, Huang DCS (2007) Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak. Science 315:856–859CrossRefGoogle Scholar
  39. Wilson BE, Mochon E, Boxer LM (1996) Induction of bcl-2 expression by phosphorylated CREB proteins during B-cell activation and rescue from apoptosis. Mol Cell Biol 16:5546–5556CrossRefGoogle Scholar
  40. Yamaguchi Y, Miura M (2015) Programmed cell death in neurodevelopment. Dev Cell 32:478–490. CrossRefGoogle Scholar
  41. Yang J, Liu X, Bhalla K, Kim CN, Ibrado AM, Cai J, Peng TI, Jones DP, Wang X (1997) Prevention of apoptosis by Bcl-2: release of cytochrome c from mitochondria blocked. Science 275:1129–1132CrossRefGoogle Scholar
  42. Yung W-H, Leung P-S, Ng SS, Zhang J, Chan SC, Chow BK (2001) Secretin facilitates GABA transmission in the cerebellum. J Neurosci 21:7063–7068CrossRefGoogle Scholar
  43. Zanjani H, Vogel M, Delhaye-Bouchaud N, Martinou J, Mariani J (1996) Increased cerebellar Purkinje cell numbers in mice overexpressing a human Bcl-2 transgene. J Comp Neurol 374:332–341CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.School of Life SciencesGuangzhou UniversityGuangzhouChina
  2. 2.GHM Institute for CNS RegenerationJinan UniversityGuangzhouChina
  3. 3.School of Biological SciencesThe University of Hong KongHong KongHong Kong

Personalised recommendations